CA1103257A - Proline derivatives and related compounds - Google Patents
Proline derivatives and related compoundsInfo
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- CA1103257A CA1103257A CA349,476A CA349476A CA1103257A CA 1103257 A CA1103257 A CA 1103257A CA 349476 A CA349476 A CA 349476A CA 1103257 A CA1103257 A CA 1103257A
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- proline
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Abstract
Abstract of the Disclosure New proline derivatives and related compounds which have the general formula are useful as angiotensin converting enzyme inhibitors.
Description
IIAl 3 Sb ~;13~
This invention relates to new proline derivatives and related compounds which have the ~eneral formula ~I) 3 ::
14 ll 21 (* )m R2- S ~ )n- C~ CO - N ~ COR
wherein R is hydro-;y, NH2 or lower alkoxy;
Rl and R~ each is hydrogen, lower alkyl, phenyl or phenyl-lower alkyl;
R2 is hydrogen, lower alkyl, phenyl, substituted phenyl wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, phenyl-lower alkyl, diphenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl, phenyl-lowe:r alkylthiometilyl, loMer alkanoMyl-1~ 1l 11 amidomethyl, R5-C-, R5-M-C-,~5-N~1-C-,R6-s- or 1~7;
R3 is hyclrogen,ilydrc)xy or lower alkyl;
R5 is lower alkyl, ~henyl or phenyl-lower ,.
~ .
. . .
~ liA135b alkyl;
R6 is lower alkyl, ~henyl, substituted phenyl, (wherein the phenyl subs-tituent is halo, lower alkyl or lower alkoxy), hydroxy-lower alkyl or amino(carboxy)-lower alkyl;
m * 1 2 Rl R~l R7 is R-OC-IIC-- N -- CO _ Cil - (CH) - S (O) ~1 is 0 or S; ,!
m is 1 to 3;
n and p each is 0 to 2, and to processes for making them.
The asterisks indicate asymmetric carbon atoms.
Each of the carbons bearing a substituent Rl, R3 and R4 is asymmetric when that substituent is other than hydrogen.
The invention in its broad aspects includes proline and related derivatives havincJ formula I above. Within this broad group, because of their properties, certain subgroups are preferred over others.
- Broadly preferred are those compounds of formula I
wherein R is hydroxy or lower alkoxy; Rl is hydro~en or lower alkyl; R2 is hydrogen, R5-CO, R6-S-, or R7; R3 and R4 each is hydrogen; R5 is lower alkyl, especially methyl or phenyl; R6 is lower alkyl, especially methyl or ethyl;
m is 2, n is 0, 1 or 2, especially 1; and R7 wherein R, Rl, 30 R3~ RL~ m and n have the same preferences as c~ove and p is 0.
~ ~ 3~5 ~g Especially prefer~ed are those compounds which have the formula (II)
This invention relates to new proline derivatives and related compounds which have the ~eneral formula ~I) 3 ::
14 ll 21 (* )m R2- S ~ )n- C~ CO - N ~ COR
wherein R is hydro-;y, NH2 or lower alkoxy;
Rl and R~ each is hydrogen, lower alkyl, phenyl or phenyl-lower alkyl;
R2 is hydrogen, lower alkyl, phenyl, substituted phenyl wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, phenyl-lower alkyl, diphenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl, phenyl-lowe:r alkylthiometilyl, loMer alkanoMyl-1~ 1l 11 amidomethyl, R5-C-, R5-M-C-,~5-N~1-C-,R6-s- or 1~7;
R3 is hyclrogen,ilydrc)xy or lower alkyl;
R5 is lower alkyl, ~henyl or phenyl-lower ,.
~ .
. . .
~ liA135b alkyl;
R6 is lower alkyl, ~henyl, substituted phenyl, (wherein the phenyl subs-tituent is halo, lower alkyl or lower alkoxy), hydroxy-lower alkyl or amino(carboxy)-lower alkyl;
m * 1 2 Rl R~l R7 is R-OC-IIC-- N -- CO _ Cil - (CH) - S (O) ~1 is 0 or S; ,!
m is 1 to 3;
n and p each is 0 to 2, and to processes for making them.
The asterisks indicate asymmetric carbon atoms.
Each of the carbons bearing a substituent Rl, R3 and R4 is asymmetric when that substituent is other than hydrogen.
The invention in its broad aspects includes proline and related derivatives havincJ formula I above. Within this broad group, because of their properties, certain subgroups are preferred over others.
- Broadly preferred are those compounds of formula I
wherein R is hydroxy or lower alkoxy; Rl is hydro~en or lower alkyl; R2 is hydrogen, R5-CO, R6-S-, or R7; R3 and R4 each is hydrogen; R5 is lower alkyl, especially methyl or phenyl; R6 is lower alkyl, especially methyl or ethyl;
m is 2, n is 0, 1 or 2, especially 1; and R7 wherein R, Rl, 30 R3~ RL~ m and n have the same preferences as c~ove and p is 0.
~ ~ 3~5 ~g Especially prefer~ed are those compounds which have the formula (II)
2 S (CH2)n CH -CO- N COR
* *
wherein R is hydroxy or lower alkoxy;
Rl is hydrogen or lower alkyl;
R2 is hydrogen, R5-CO-, R~-S- or R7;
R5 is lower alkyl or pheryl, especially the first;
R6 is lower alkyl;
and n is 0, 1 or 2.
Within the group of compounds represented by formula II, the following are still more preferred subgroups in the order (a to r) of increasing preference to the com-pounas which are especially preferred emhodiments:
a) R is hydroxy b) n is 1 c) R2 is hydrogen or lower alkanoyl d) R2 is hydrogen e) R2 is acetyl f) Rl is hydrogen or lower alkyl g) Rl is hydrogen or methyl h) R is hydroxy, Rl is hydrogen or methyl i) R is hydroxy, Rl is hydrogen or methyl, R2 is hydro-gen or acetyl and n is 0, 1 or 2 j) R is hydroxy, Rl and R2 each is hydrogen and n is 0 .~ ,, . ~
.
k) R is hydroxy, Rl is hydrogen, R2 is acetyl and n is 1) R is hydroxy, Rl is methyl, R2 is acetyl and n is 1 m) R is hydroxy, Rl and R2 each is hydrogen and n is 1 n) R is hydroxy, Rl is methyl, R2 is hydrogen and n is o) R is hydroxy, Rl is hydrogen, R2 is lower alkylthio and n is 1 p) R2 is ~ 11 ,, R-OC N -OC CH - (CH2~n-S-;
each R is hydroxy; Rl is hydrogen or lower alkyl, es-pecially hydrogen or methyl; and n is 0 to 2, es-pecially 1 M
q) R2 is R5-M-C- wherein M is O or S
M
r) R2 iS R5-NH-C- wherein M is O or S, preferably S.
It will be appreciated that combinations of the fore-goingr where applicable, are among the preferred groups.
The stereoisomers in which the proline is in the L-form are especially preferred.
The lower alkyl groups represented by any of the vari ables include straight and branched chain hydrocarbon radicals from methyl to heptyl, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, iso-pentyl and the like. The lower alkoxy groups are of the same kind having 1 to 7 carbons linked to oxygen, for ex-ample, methoxy, ethoxy, propoxy, isopropoxy, butoxyl iso-butoxy, t-butoxy and the like. The Cl-C4 members, espe-cially Cl and C2 members, of both ~ ~ 4 ~
~V ~ ~
IIA135b types are pr~ferred. Phenylmeth~l is thc ~re~erred phenyl-lower alkyl group.
The lower al~anoyl groups are those havillg the acyl radicals of the lower (C2-C7) ~-atty acic~s, for example, acetyl, propionyl, butyryl, isobutyryl and the like. Similarly, those lower alkanoyl groups havinc3 up to four carbons, and especially acetyl, are preferred.
The four cor,lmon haloyens are included by the term "halo" but chlorine and bromine are preferred. The substi-tuted phenyl groups preferably bear the substituent in the 4-position of the ring. The hydroxy-lower alkyl groups have a hydroxy group on an alkyl chain like those described above, preferably on the terminal carbon, e.g., hydroxymethyl, 2-hydroxyethyl, etc. The amino~carboxy)lower alkyl groups have one amino and one carbo~y on a lower alkyl group such as those described above, preferably both on one carbon, e.g., on the terminal carbon as in the preferred 2-amino-2-carboxyethyl group.
The products of formula I and the preferred subgroups can be produced by various methods of synthesis.
In general, the products of this invention are produced by acylating a compound of the formula (III) 2C (C~l)m HN Cll - COR
with an acid of the formula (IV) R~ II
1~2 5 (Cl~ C()OII
_ rj_ . . . _ ilA13Sb or its chemical equivalent.
Thus, the final product can be produced not only by direct acylation with an acid of formula IV hut also by~ intermediates such as ta) l~-haloalkanoic acids of the formula (V) 1 11 X - - (CH) -CH - COOH
wherein X is bromo, chloro or iodo, or (b) a tosyloxyalkanoic acid, i,e., X in formula V is tosyloxy~ (CH3 ~ -S2-) (c) a substituted acrylic acid of the formula 14 IRl (VI) CH - C COOH
The product of this acylation is then subjected to displacement or addition with the anion of a thiol or thioacid of the formula (VII) R2 SH
Acylation can also be effected with a thiolactone of the formula (VIII) (fH)n CH
S =O
wherein n is 1 or 2, or a mercaptoalkanoic acid of the formula IR4 IRl (IX) Y S -(CH)n CH COR
, ~IA~35i~
wherein Y is R2 or, in adclition, i~ d product of forml~la I
whereill R2 is hydro~en is desired, then Y can also be a protecting group such as ~a) C~i30~ C~l2-, (b) ~ , (c) CH3CONHC~l2, (cl) ~-O-C-C~I-(C~I) -S- or other sulfur protecting group. "Depro-tection" can be effected by conventional means such as treatment with hot trifluoroacetic acid, cold trifluoromethanesulfonic acid, mercuric acetate, sodium in liquid ammonia, zinc and hydrochloric acid or the like. For a review oE these methods see Methoden der Organischen Chemie (Houben-Weyl), Vol. XV, part I, page 736 et seq. (197~) When the acid oE forlllula IV is used as the acylatinc agent, th~ acylation can be effected in the presence of a coupling agent like dicyclohexycarbodiimide or the like, or the acid ean be activated by formation of its mixed annydride, sy~netrical anhydride, acid chloride, acid ester or use of Woodward reagent ~, N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline or the like. For a review of the methods for acylation, see Methoden der Orc~anischen Chemie 2Q (Houben-Weyl), Vol. XV, part II, page 1 et seq. (1974).
Compounds of formula III include, for example, proline, hydroxyproline, 4-methylproline, pipecolic acid, 5-hydroxypipecolic acid, azetidine-2-carboxylic acid, their lower alkyl esters and the like. The acylation of SUC}l eompounds is described in greater detail below.
Aeeording to a preferred method for producinc3 compounds of formula I, espeeially wherein R2 is R5-CO-, an acid or ester of formula III is coupled with a haloalkanoic acid of the formula _ .
-IIA135b (V) i~ L
X-(Cil)n- Ci~ COOii wherein X is a halocJen, prefera~ly chlorine or bromine. This can be effected by one of the kno~-~n procedures in which the acid IV is activated, prior to reaction with the acid III, involving formation of a mixed anhydride, symmetrical anhydride, acid chloride, active estcr, or use of ~ooclward reagent K, EEDQ
(N-ethoxycarbonyl-2-ethoxy-1,2-dihydroxyquinoline) or the like.
The product of this reaction is a compound of the formula (X) 13 14 1l 21 (Cil)m .
X - ~C~ -C~l- CO - N- CII-COR
The product of formula X is subjectcd to a ciis~laccmc.~nt reaction with the anion of a thioacid oE the formula (VII) R2-S~]
yielding a product of the formula (XI) 14 1l 112C- (Cll)m R2- S - (Cll)n- CII-CO - N - CII-COR
l~hen R2 is R5CO, this product can thcll bc convcrteci to the product (XII) R3 ~ 2C (Cii) liS - (Cll) - C'li - CO - N CII-COR
-------by ammonolysis. When R2 is a protecting ~roup, then the compound o formula XIl can be obtained by "deprotection"
as described above. When R is an ester group (i.e., R is lower alkoxy), the ester group can be removed, e.g., when R is tert. butoxy or tert. amyloxy, by treatment oE the ester of formula XI or XII with trifluoroacetic acid and anisole to give the corresponding free acid. When other alkoxy groups are present, alkaline hydrolysis will yield the corresponding acid.
A variation of this procedure involves the use of an acrylic acid of the formula (VI) IR4 Rll CH=C-COOH
as starting material. This acrylic acid is first conver-ted to the acid halide form then made to react with a compound of formula III to obtain a compound of the for-mula (XIII) 13 IR4 IRl H2C (CH)m CH=C - CO - N CH-COR
and this intermediate is subjected to the addi-tion reac-tion with the thiol or thioacid VII as described above.
A tosyloxyalkanoic acid of the formula (XIV) R14 IRl CH3- ~ _ S020-(CH)n CH - COOH
can also be used as the agent to acylate the acid of formula III, then the acylation product is subjected to the displacement reaction, etc., as described above.
The acrylic acid of formula VI can alternatively be first made to react with the thioacid of formula VII to '!
-obtain a product of the formula (XV) R4 Rl R -S-C~-CH-COOH
which is converted to its acid halide, e.g., with thionyl chloride, then coupled to the compound of formula III and the same sequence as above then fo:Llowed.
The acid or ester of formula :[II can also be acylated with a "proteeted" form of a ~-mercaptoalkanoic aeid of the formula (XVI) R4 Rl R8~S~(CH)n~CH~CH
whe ein R8 is the "protecting" group. Such "protecting"
groups can take the form deseribed above.
Following the aeylation, the produet ean be "depro-teeted" by one of the known methods referred to above.
Still another aeylating agent ean take the form of a thiolaetone, e.g., ~-propiothiolaetone, ~-methyl-~-pro-piothiolaetone or the like.
Additional details of preferred modes of produeing eompounds of this invention ean be found in the following and in the speeifie examples.
Aecording to a partieularly preferred modifieation, the aeid or ester of formula III is acylated with a halo-alkanoyl halide of the formula (XVII) R4 Rl X-(CH)n CH-COX
wherein eaeh X is independently a halogen, preferably ehlorine or bromine, Rl is hydrogen, lower alkyl or phenyl-lower alkyl and n is 0, 1 or 2. ~his reaction is effected in an alka-line medium, e.g., dilute alkali metal hyclroxide solution, alkali metal dicarbonate or alkali metal carbonate solution at a reduced temperature, e.g., about 0 to 15C. The re-action product is subjected to displacement with the anion of the thiol or thio acid of the formula ~II above, also in alkaline medium, preferably alkali metal carbonate so-lution, and then worked up in conventional manner. The product of this reaction, wherein R2 of formula I is R5-CO, is converted to the product wherein R2 is hydrogen by ammonolysis, e.g., alcoholic ammonia or concentrated ammonium hydroxide solution, or alkaline hydrolysis, e.g., with aqueous metal hydroxide. When an acid of formula III is used as starting material, the final product ob-tained as the free carboxylic acid can then be converted to its ester, for example by esterification with a dia-zoalkane, like diazomethane, l-alkyl-3-p-tolyl-triazene, like l-n-butyl-3-p-tolyltriazene or the like. l`reatment of an ester, preferably the methyl ester, with an alco-holic ammonia solution, converts the free acid to theamide, i.e., R is NH2.
According to another variation, an ester, prefer-ably the t-butyl ester, of formula III, in an anhydrous medium such as dichloromethane, tetrahydrofuran, dioxane or the like, is treated with a thioalkanoic acid of the formula (XVIII) 1l R2-S-(CH2)n - CH ~ COOH
in the presence of dicyclohexylcarbodiimide, N,N'-car-bonyl-bisimidazo]e, ethoxyacetylene, diphenylphosphoryl azide or similar coupling agents at a temperature in the range of about 0 to 10C. The ester group (R) can then be removed, for ., .~, --11--.
.3 ~
e.~ample, by treatment with trifluoroacetic acid and ani-sole at about room temperature.
When an ester of formula III (e.g., R is lower al-koxy, especially, t-butoxy) is acylated with a thiolactone, e.g., ~-propiothiolactone, ~-me-thyl-~-propiothiolactone or the like, the reaction can be eEfected in an anhydrous solvent like tetrahydrofuran, dioxane, methylene chloride or the like at about 0 C. to about room temperature. The ester group can be removed with anisole and trifluorace-tic acid as described above. M
In similar manner, when R2 is R5-M-C-, products of formula I having this substituent are formed by reacting a compound of formula XII with the halogenated compound (XIX) M
or alternatively reacting a compound of formula X with an alkali metal salt or alkaline earth metal salt of the formula (XX~ M
R5-M-C-S-Me wherein Me represents the alkali metal or alkaline earth metal. M
When R2 is R5-NH-C-, products of formula I having - this substituent are produced by reacting a compound of formula XII with the appropriately substituted isocyanate or isothiocyanate of the formula (XXI) R5-N=C=M
Alternatively, the same products can be produced by coup-ling an acid of the formula ~ -12-:i~l ~5~
HA135b (XXII) 1'1 R~ IRl R5-N}I-C-S-(CII)-- CH-COOH
with an amino acid of formula III.
Compounds oE formula I, wherein R2 is lower alkyl, phenyl, substituted phenyl, phenyl-lower alkyl, -triphenyl-lower alkyl, lower alkylthiomethyl or phenyl-lower alkylthiome-thyl are produced by reacting a compound or formula XII with the corresponding halide R2X or by reacting a compound of formula X
with the corresponding thiol R2SH in the same manner as described above.
When R2 is lower alkanoylamidomethyl, -the product of Eormula I is produced by condensing a compound of formula XII
with the corresponding hydroxymethyl-lower alkanoylamide of the formula ~XXIII) lower alkyl-CO-NI~CH2OH
in the presence of an acid catalyst like trifluoroacetic acid.
Products of formula I wherein R2 is R6-S can be prepared by any of the known methods for the synthesis of mi~ed disulfides, e.g., by the reaction of a compound of formula(XII)with a thiosulfinate(xxIv), thiosulfonate (XXV), sulfenyl halide (XXVI), thiosulfa-te (XXVII) or sulfenyl thiocyanate (XXVIII) O
tXXIV) R6-S-S-R6 , tXXV) R6-S-S-R6 , (XXVI) R6S-X, O i, (XXVII) R6-S-SO3~1, (XXVIII) R6-S-SCN
In the particular case wherein R7 is l4 IRl CH2 - (IcH)n -S(O)--- (CH) -- CH _ CO - N--- CH-COR, .
~ }IA135b R, Rl, R3 and R4 are t~e same as -the corresponding substituents in formula I and p is O, the symmetrical disulfides can be obtained by direct oxidation of a compound of formula XII
with iodine. When p is 1 or 2, such products are obtained by the stepwise oxidation of the corresponding compound wherein p is O. Mixed disulfides are obtained by the modification shown in the examples.
Products of formula I have one or more asymmetric carbons.
When Rl, R3 or R~ is other than hydrogen the carbon to which it is attached is asymmetric. These carbon atoms are indicated by an asterisk in formula I. The compounds accordingly exist in stereoisomeric forms or in racemic mixtures thereof. All of these are within the scope of the invention. The above described synthesis can utilize the racemate or one of the enantiomers as starting material.
When the racemic starting material is used in the synthetic procedure, the stereoisomers obtained in the product can be separated by conventional chromatographic or fractional crystallization methods. In general, the L-isomer with respect to the carbon of the amino acid constitutes the preferred isomeric form. Also the D-isomer with respect to the a-carbon in the acyl side chain (i.e., the carbon bearing Rl) is preferred.
The compounds of this invention form basic salts with various inorganic and organis bases which are also within the scope af the invention. Such salts include ammonium salts, alkali metal salts like sodium and potassium salts ~which are preferred), alkaline earth metal salts like the calcium and magnesium salts, salts with organic bases, e.g., dicyclohe~ylamine salt, benzathine, N-methyl-D-glucamine, ;
~A135b hydrabamine salts, salts with amino acids like arginine, I
lysine and the like. The non-toxic, physiologically acceptable salts are preferred, although other salts are also useful, e.g., in isolating or purifyinc~ the product, as illustrated in the examples in the case of tihe dicyclohexylamine salt.
The salts are formed in conventional manner by reacting the free acid form of the product with one or more equivalents of the appropriate base providing the desired cation in a solvent or medium in which the salt is insoluble, or in water and removing the water by freeze drying. By neutralizing the salt with an insoluble acid like a cation exchange resin in the hydrogen form (e.g., polystyrene sulfonic acid resin like Dowex 50) or with an aqueous acid and extraction with an organic solvent, e.g., ethyl acetate, dichloromethane or the like, the free acid form can be obtained, and, if desired, another salt formed.
. -- --Additional experimelltal details are found in the examples which are preferred embodiments and also serve as models for the preparation of other members of the ~roup.
The compounds of this invention inhibit the ~onver-sion of the decapeptide angiotensin I to angiotensin II
and therefore are useful in reducing or relieving angio-tensin related hypertension. The action of the enzyme re-nin on angiotensinogen, a pseudoglobulin in blood plasma, produces angiotensin I. Angiotensin I is converted by angiotensin converting enzyme (ACE) to angiotensin II.
The latter is an active pressor substance which has been implicated as the causative agent in various forms of hypertension in various mammalian species, e.g., rats and dogs. The compounds of this invention intervene in the angiotensin(renin)~angiotensin I ~angiotensin II sequence by inhibiting angiotensin converting enzyme and reducing or eliminating the formation of the pressor substance angiotensin II. Thus by the administration of a composi-tion containing one or a combination of compounds of for-mula I or physiologically acceptable salt thereof, angio-tensin dependent hypertension in the species of mammal suffering therefrom is alleviated. A single dose, or pre-~ferably two to four divided daily doses, provided on a ,~
basis of about 0.1 to 100 mg. per kilogram per day, pre-ferably about 1 to 50 mg. per kilogram per day is appro-priate to reduced blood pressure as indicated in the ani-mal model experiments described by S.L. Engel, T.R.
Schaeffer, M.H. Waugh and B. Rubin, Proc. Soc. Exp. Biol.
Med. 143, 483 (1973). The substance is preferably admin-istered orally, but parenteral routes such as subcutaneous,intramuscular, intravenous or -16- -~
intraperitoneal can also be employed.
~ ne cc~pounds of this invention can be utilized to aehieve the reduction of blood pressure by formulating in cc~positions sueh as tablets, eapsules or elixirs for oral acbministration or in s-terile solutions or suspensions for parenteral administration. About 10 to 500 mg. of a compound or mixture of compounds of formula I or physio-logieally acceptable salt is compounded with a physiologieally accep-table vehiele, carrier, excipient, binder, preservative, stabilizer, flavor, ete., in a unit dosage form as ealled for by accepted phar-maceutieal praetiee. m e amount of active substance in these cc~positions or preparations is sueh that a suitable dosage in the range indicated is obtained.
Illustrative of the adjuvants whieh may be ineorporated in tablets, eapsules and the like are the follc~ing: a binder such as gum tragaeanth, aeaeia, eorn stareh or gelatin; an exeipient such as diealeium phosphate; a disintegrating agent sueh as eorn stareh, potato stareh, alginie aeid and the like; a lubrieant sueh as magnesium stearate; a sweetening agent such as sucrose, laetose or ~ !
saeeharin; a flavoring agent such as peppermint, oil of winter-green or eherry. When the dosage unit form is a eapsule, it may eontain in addition to materials of the above type a liquid earrier sueh as a fatty oil. Various other materials may be present as eoatings or to otherwise modify the physieal form of the dosage unit. For instanee, tablets may be eoated with shellae, sugar or both. A syrup or elixir may eontain the aetive compound, suerose as a sweetening agent, methyl and propyl parabens as preservatives, a dye and a flavoring such as eherry or orange flavor.
Sterile eompositions for injeetion ean be formulated according to conventional pharmaceutical practice by dis-solving or suspending the active substance in a vehicle such as water for injection, a naturally occurring vege-table oil like sesame oil, coconut oil, peanut oil, cotton-seed oil, etc., or a synthetic Eatty vehicle like e-thyl oleate or the like. Buffers, preservatives, antioxidants and the like can be incorporated as required.
The following examples are illustrative of the in-ven~ion and cons-titute especially preferred embodiments t All temperatures are in degrees celsius.
~J
,,,~, ~ 3~
E~ample 1 _ 1-(2-Benzoylthioacetyl)-L-Proline L-Proline (5.75 g.) is dissolved in N sodium hydr-oxide (50 ml.) and the solution is chilled in an ice-water bath. Sodium hydroxide 2N (26 ml.~ and chloroacetyl chlo-ride (5.65 g.) are added and the mixture is stirred vigo-rously at room temperature for three hours. A suspension of thiobenzoic acid (7.5 g.) and potassium carbonate (4.8 g.) in water (50 ml.) is added. After 18 hours stirring at room temperature, the reaction mixture is acidified and extracted with ethyl acetate. The ethyl acetate layer is washed with water, dried over magnesium sulfate and con-centrated to dryness in vacuo. The residue (14.6 g.) is dissolved in ethyl acetate (150 ml.) and dicyclohexylamine (11 ml.) is added. The crystals are filtered and recry-stallized from ethyl acetate, yield 5.7 g. m.p. 151-152.
To convert the salt to the acid, the crystals are dissol-ved in a mixture of 5~ aqueous potassium bisulfate (100 ml.) and ethyl acetate (300 ml.). The organic phase is washed onee with water, dried over magnesium sulfate and eoncentrated to dryness in vacuo, yield 3.45 g.
Example 2 1-(2-Mercaptoacetyl)-L-Proline 1-(2-Benzoylthioacetyl)-L-proline (3O4 g.) is dis-solved in a mixture of water (10.5 ml.) and concentrated ammonia (6.4 ml.~. After one hour, the reaction mixture is diluted with water and filtered. The filtrate is ex-traeted with ethyl aeetate and then aeidified with eon-eentrated hydrochloric acid, saturated with sodium chlo-ride and extracted twice with ethyl acetate. The ethylacetate extraets are washed with saturated sodium ehlo-ride and eoneentrated to dryness, yield 1.5 g.
,~
The product, 1-(2-mercaptoacetyl)-L-proline is crystallized from ethyl acetate (m.p. 133-135) .
Exam~le 3 1-(2-Benzoylthioacetyl)-L-Proline Methyl Ester 1-(2-Benzoylthioacetyl)-L-proline o~tained in Example 1, is dissolved in methanol and an ethereal solution of diazomethane is added until there is a persistent yellow color. After 15 minutes, a few drops of acetic acid are added and the solvent is removed in vacuo to obtain 1-(2-benzoylthioacetyl)-L-proline methyl ester.
Example 4 1-(2-Mercaptoacetyl)-L-Proline Amide The product of Example 3 is dissolved in 10~ metha-nolic ammonia and the solution is stored at room tempera-ture in a pressure bottle. When thin layer chromatogra-phic analysis indicates that the two ester functions have been ammonolyzed, the reaction mixture is concentrated to dryness to obtain l-(2-mercaptoacetyl)-L-proline amide.
Example 5 1-(2-Benzoylthioacetyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example 1, 1-(2-benzoylthioacetyl~-L-hydroxyproline is obtained.
Example 6 1-(2-Mercaptoacetyl~-L-Hydroxyproline By treating the product of Example 5 with ammonia as in Example 2, 1-t2-mercaptoacetyl)-L-hydroxyproline is obtained.
Example 7 1-(2-Benzoylthioacetyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid for the L-proline in the procedure of Example 1, 1-(2-benzoylthio-acetyl)-L-azetidine-2-carboxylic acid is obtained.
Example 8 1-(2-Mercaptoace-tyl)-L-Aze idine-2-Carboxylic ~cid By treating the product of ~xample 7 with ammonia as in Example 2, 1-(2-mercaptoacetyl)-L-azetidine-2-car-boxylic acid is obtained.
Example 9 1-(2-Benzoylthioacetyl)-L-pipecolic Acid , .
By substituting L-pipecolic acid for the L-proline in the procedure of Example 1, 1-(2-benzoylthioacetyl)-L-pipecolic acid is obtained.
Example 10 1-(2-Mercaptoacetyl)-L-Pipecolic Acid By treating the product of Example 9 with ammonia as in Example 2, 1-(2-mercaptoacetyl)-L-pipecolic acid is obtained.
Example 11 l-t2-Benzoylthiopropanoyl)-L-Proline L-Proline (5.75 g.) is dissolved in aqueous N so-dium hydroxide (50 ml.) and the solution is chilled in an ice bath with stirring. 2N sodium hydroxide (25 ml.) and 2-bromopropionyl chloride (8.57 g.) are added in that order and the mixture is removed from the ice bath and stirred at room temperature for one hour. A mixture of thiobenzoic acid (7.5 g.) and potassium carbonate (4.8 g.) in water (50 ml.) is added and the mixture is stirred over-night at room temperature. After acidification with con-centrated hydrochloric acid, the aqueous solution is ex-tracted with ethy:l acetate and the organic phase is washed with water, dried and concentrated to dryness. The resid~le (14.7 g.) is chroma-to~raphed on a column of 440 g. o~ silica gel with a mix-ture of benzene-acetic acid (7:1). The fractions contain-ing the desired material are pooled, concentrated to dry-ness, and the residue is precipitated -twice with ether-hexane and converted to a dicyclohexylamine salt in ether-hexane, yield 9.4 g. m.p., (142) 148-156. The dicyclo-hexylamine salt is converted back to the acid as in Exam-ple 1, yield 5.7 g.
Example 12 1-(2-Mercaptopropanoyl)-L-Proline 1-(2-Benzoylthiopropanoyl)-L-proline (5.7 g.) is dissolved in a mixture of water (12 ml.) and concentrated ammonium hydroxide (9 ml.) with stirring. After one hour, the mixture is diluted with water (10 ml.) and filtered.
The filtrate is extracted twice with ethyl acetate, con-centrated to one-third of the original volume, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with satura-ted sodium chloride, dried and concentrated to dryness invacuo. The residue, 1-(2-mercaptopropanoyl)-L-proline, is crystallized from ethyl acetate-hexane, yield 3 g., m.p.
(105) 116-120.
Example 13 1-(3-Benzoylthiopropanoyl)-L-Proline L-Proline (5.75 g.) is dissolved in normal sodium hydroxide (50 ml.) and the solution is chilled in an ice bath. 3-Bromopropionyl chloride (8.5 g.) and 2N sodium hydroxide (27 ml.) are added and the mixture is stirred for 10 minutes in the ice bath and three hours at room temperature. A suspension of thiobenzoic acid (7.5 g.) and potassium ca:rbonate (4 5 g.) in water (50 ml.) is added and the mixture is stirred .
.
v~7 for 18 hours at room tempera-ture. After acidification with concentrated hydrochloric acid, the aqueous phase is ex-tracted twice with ethyl acetate. The organic layers are dried over magnesium sulEate and concentrated to dryness in vacuo to obtain l-(3-benzoylthiopropanoyl)-L-proline, yield 7.1 g., m.p. 101-102 (ethyl acetate-hexane).
Example 14 L-Proline tert.-butyl ester L-Proline (230 g.) is dissolved in a mixture of water (1 1.) and 5 N sodium hydroxide (400 ml.). The so-lution is chilled in an ice bath, and under vigorous stir-ring, 5 N sodium hydroxide (460 ml.) and benzyloxycarbonyl chloride (340 ml.) are added in five equal aliquots during a half hour period. After one hour stirring at room tem-perature, the mixture is extracted twice with ether and acidified with concentrated hydrochloric -acid. The preci-pitate is filtered and dried. Yield 442 g., m.p. 78-80.
The benzyloxycarbonyl-L-proline thus obtained (180 g.~ is dissolved in a mixture of dichloromethane (300 ml.), liquid isobutylene (800 ml.) and concentrated sulfuric acid (7.2 ml.). The solution is shaken in a pressure bottle for 72 hours. The pressure is released, the isobutylene is allowed to evaporate and the solution is washed with 5 sodium carbonate, water, dried over magnesium sulfate and concentrated to dryness in vacuo, to obtain benzyloxycar-bonyl-L-proline tert. butyl ester, yield 205 g.
Benzoyloxycarbonyl-L-proline tert. butyl ester (205 g.) is dissolved in absolute ethanol (1.2 1) and hy-drogenated at normal pressure with 10~ Pd on carbon (10 g.) until only a trace of carbon dioxide is observed in the hydrogen exit gas ~4 hours). The catalyst is filtered off and the Eiltrate is c~ncentrated in vacuo at 30 mm. H~. The residue is distilled in vacuo, to obtain L-proline tert.butyl ester, b.p. 50-51.
lmm Example 15 1-(3-~cetylthiopropanoyl)-L-Proline tert-butyl Ester L-Proline tert-butyl ester (5.13 g.) is dissolved in dichloromethane (40 ml.) and the solution is chilled in an ice-water bath. A solution of dicyclohexylcarbodiimide (6.18 g.) in dichloromethane (20 ml.) is added followed immediately by 3-acetylthiopropionic acid (4.45 g.).
After 15 minutes stirring in the ice-water bath and 16 hours at room temperature, the precipitate is filtered off and the filtrate is concentrated to dryness in vacuo.
The residue is dissolved in ethyl acetate and washed neu-tral. The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo to obtain 9.8 g. of 1-(3-acetylthiopropanoyl)-L-proline tert-butyl ester.
Example 16 1-(3-Acetylthiopropanoyl)-L-Proline 1-(3-Acetylthiopropanoyl)-L-proline-t-butyl ester (4.7 g.) is dissolved in a mixture of anisole (34 ml.) and trifluoroacetic acid (68 ml.) and the mixture is kept at room temperature for one hour. The solvents are removed in vacuo and the residue is precipitated from ether-hexane several times. The residue (3.5 g.) is dis-solved in acetonitrile (25 ml.) and dicyclohexylamine (2.8 ml.) is added. The crystalline salt is filtered and recrystallized from isopropanol. Yield 3.8 g.. m.p.
176-177. The salt is reconverted to 1-(3-acetylthio-propanoyl)-L-proline as in --?4--~ r~ ~
E.~ample 1, yield 1 25 g., m.p. 89-90 (ethyl acetate-he~ane).
Example 17 1-(3-Mercaptopropanoyl)-L-proline tert-butyl Ester To a solution of L-proline tert-butyl ester (3.42 g.) in dry tetrahydrofuran (10 ml.) chilled in an ice bath, propiothiolactone (1.76 g.) is added. After 5 minutes storage in the ice bath and three hours at room -temperature, the reaction mixture is diluted with ethyl acetate (200 ml.) and washed with 5~ potassium bisulfate, and water.
The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo. The residue 1-(3-mer-captopropanoyl)-L-proline tert-butyl ester is crystallized from ether-hexane, yield 3.7 g., m.p. 57-58.
Example 18 1-(3-~lercaptopropanoyl)-L-Proline Procedure A
1-(3-Benzoylthiopropanoyl)-L-proline (4.9 g.) is dissolved in a mixture of water (8 ml.) and concentrated ammonium hydroxide (5.6 ml.) and the solution is stored with stirring under argon for one hour. The reaction mix-ture is diluted with water, filtered, and the filtrate is extracted with ethyl acetate. The aqueous phase is acid-ified with concentrated hydrochloric acid, saturated with sodium chloride and extracted with ethyl acetate. The organic layers are washed with saturated sodium chloride, dried over magnesium sulfate, and concen-trated to dryness in vacuo. The residue, 1-(3-mercaptopropanoyl)-L-proline, is crystallized from ethyl acetate hexane, yield 2.5 g., m.p. 68-70.
Procedure B
.
l-t3-Acetylthiopropanoyl)-L-proline (0.8 g.) is dis-solved :: `
in 5.5 N methanolic ammonia (5 ml.) and the solution kept under ar~on at room temperature. After 2 hours the solvent is removed in vacuo, the residue is dissolved in water and applied to an ion exchange column on the H cycle [Dowex 50 (Analytical grade)] and eluted with water. The fractions that give thiol positive reaction are pooled and concentra-ted to dryness, yield 0.6 g. This product is crystallized from ethyl acetate-hexane as in Procedure A to obtain 1-(3-mercaptopropanoyl)-L-proline.
Procedure C
1-(3-Mercaptopropanoyl)-L-proline t-butyl ester (2.3 g.) is dissolved in a mixture of anisole (20 ml.) and tri-fluoroacetic acid (45 ml.). After one hour storage at room temperature under argon, the reaction mixture is con-centrated to dryness in vacuo and the residue precipitated from ethyl acetate-hexane several times. The residue (1.9 g.) is dissolved in ethyl acetate t30 ml.) and di-cyclohexylamine (1.85 ml.) is added. The crystalline salt is filtered and recrystallized from isopropanol, yield 2 g., m.p. 187-188.
The salt is converted to the acid as in Example 1, yield 1.3 g. The product is crystallized from ethyl ace-tate hexane as in Procedure A.
Salts sOaium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.) is dis-solved in a mixture of water (2.5 ml.) and N sodium hy-droxide (2.5 ml.). The solution is freeze dried to ob-tain the sodium salt.
Magnesium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.), mag-nesium oxide (49.5 mg.), and water ~10 ml.) are stirred ~ -26-with slight heating until complete solution is obtained.
Then the solvent is removed by freeze drying to obtain the magnesium salt.
Calcium 1-(3-Mercaptopropanoyl)-2-proline (S00 mg.) is dissolved in a mixture of calcium hydroxide (91 mg.) and water (10 ml.), and the solu-tion is freeze dried to obtain the calcium salt.
Potassium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.) is dissolved in a mixture of potassium bicarbonate (246 mg.) and water (10 ml.) and freeze dried to obtain the potassium salt.
N-Methyl-D-Glucamine 1-(3-Mercaptopropanoyl)-L-proline(500 mg.) and N-methyl-D-glucamine (480 mg.) are dissolved in water (10 ml.) - and freeze dried to obtain the N-methyl-D-glucamine salt.
Example 19 1-(3-Mercaptopropanoyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example 11 and then treating the pro-duct by Procedure A of Example 18, 1-(3-benzoylthiopropan-oyl)-L-hydroxyproline and 1-(3-mercaptopropanoyl)-L-hydr-, oxyproline, respectively, are obtained.
Example 20 1-(3-Mercaptopropanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid tert-butyl ester (prepaxed by substituting L-azetidine-2-carboxylic acid for the proline in Example 14) for the L-proline tert-butyl ester in the procedure of Example 15, ~AI
treating the product as in Example 16 and the 1-(3-acetyl-thiopropanoyl)-L-azetidine-2-carboxylic acid thus obtained by Procedure B of Example 18, 1-(3-acetylthiopropanoyl)-L-azetidine-2-carboxylic acid tert-butyl ester and 1-(3-mercaptopropanoyl)-L-azetidine-2-carboxylic acid, respec-tively, are obtained.
Example 21 1-(3-Mercaptopropanoyl)-L-Pipecolic Acid By substituting L-pipecolic acid tert-bu-tyl ester (prepared by substituting L-pipecolic acid for the L-pro-line in Example 14) for the L-proline tert-butyl ester in the procedure of Example 15 and treating the product by Procedure C of Example 18, 1-(3-mercaptopropanoyl)-L-pipecolic acid tert-butyl ester and l-(3-mercaptopropan-oyl)-L-pipecolic acid, respectively, are obtained.
Example 22 1-(3-Mercaptopropanoyl)-4-Methyl-L-Proline By substituting 4-methyl-L-proline for L-proline in the procedure of Example 13 and then treating the product by Procedure A of Example 18, 1-(3-benzoylthiopropanoyl)-4-methyl-L-proline and 1-(3-mercaptopropanoyl)-4-methyl-L-proline, are obtained.
Example 23 1-(3-Mercaptopropanoyl)-5-Hydroxy-L-Pipecolic Acid By substituting 5-hydroxy-L-pipecolic acid for L-proline in the procedure of Example 13 and then treating the product by the Procedure A of Example 18, 1-(3-ben-zoylthiopropanoyl)-5-hydroxy-L-pipecolic, and 1-(3-mercaptopropanoyl)-5-hydroxy-L-pipecolic acid are obtained.
Example 24 1-(3-Mercaptopropano~l)-D-Proline By substituting D-proline for L-proline in the pro-cedure of E~ample 13 and then treating the product by Pro-cedure A of Example 18, 1-(3-benzoylthiopropanoyl)-D-proline and l-(3-mercaptopropanoyl)-D-proline, m.p. 68-70, are ob-tained.
Example 25
* *
wherein R is hydroxy or lower alkoxy;
Rl is hydrogen or lower alkyl;
R2 is hydrogen, R5-CO-, R~-S- or R7;
R5 is lower alkyl or pheryl, especially the first;
R6 is lower alkyl;
and n is 0, 1 or 2.
Within the group of compounds represented by formula II, the following are still more preferred subgroups in the order (a to r) of increasing preference to the com-pounas which are especially preferred emhodiments:
a) R is hydroxy b) n is 1 c) R2 is hydrogen or lower alkanoyl d) R2 is hydrogen e) R2 is acetyl f) Rl is hydrogen or lower alkyl g) Rl is hydrogen or methyl h) R is hydroxy, Rl is hydrogen or methyl i) R is hydroxy, Rl is hydrogen or methyl, R2 is hydro-gen or acetyl and n is 0, 1 or 2 j) R is hydroxy, Rl and R2 each is hydrogen and n is 0 .~ ,, . ~
.
k) R is hydroxy, Rl is hydrogen, R2 is acetyl and n is 1) R is hydroxy, Rl is methyl, R2 is acetyl and n is 1 m) R is hydroxy, Rl and R2 each is hydrogen and n is 1 n) R is hydroxy, Rl is methyl, R2 is hydrogen and n is o) R is hydroxy, Rl is hydrogen, R2 is lower alkylthio and n is 1 p) R2 is ~ 11 ,, R-OC N -OC CH - (CH2~n-S-;
each R is hydroxy; Rl is hydrogen or lower alkyl, es-pecially hydrogen or methyl; and n is 0 to 2, es-pecially 1 M
q) R2 is R5-M-C- wherein M is O or S
M
r) R2 iS R5-NH-C- wherein M is O or S, preferably S.
It will be appreciated that combinations of the fore-goingr where applicable, are among the preferred groups.
The stereoisomers in which the proline is in the L-form are especially preferred.
The lower alkyl groups represented by any of the vari ables include straight and branched chain hydrocarbon radicals from methyl to heptyl, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, iso-pentyl and the like. The lower alkoxy groups are of the same kind having 1 to 7 carbons linked to oxygen, for ex-ample, methoxy, ethoxy, propoxy, isopropoxy, butoxyl iso-butoxy, t-butoxy and the like. The Cl-C4 members, espe-cially Cl and C2 members, of both ~ ~ 4 ~
~V ~ ~
IIA135b types are pr~ferred. Phenylmeth~l is thc ~re~erred phenyl-lower alkyl group.
The lower al~anoyl groups are those havillg the acyl radicals of the lower (C2-C7) ~-atty acic~s, for example, acetyl, propionyl, butyryl, isobutyryl and the like. Similarly, those lower alkanoyl groups havinc3 up to four carbons, and especially acetyl, are preferred.
The four cor,lmon haloyens are included by the term "halo" but chlorine and bromine are preferred. The substi-tuted phenyl groups preferably bear the substituent in the 4-position of the ring. The hydroxy-lower alkyl groups have a hydroxy group on an alkyl chain like those described above, preferably on the terminal carbon, e.g., hydroxymethyl, 2-hydroxyethyl, etc. The amino~carboxy)lower alkyl groups have one amino and one carbo~y on a lower alkyl group such as those described above, preferably both on one carbon, e.g., on the terminal carbon as in the preferred 2-amino-2-carboxyethyl group.
The products of formula I and the preferred subgroups can be produced by various methods of synthesis.
In general, the products of this invention are produced by acylating a compound of the formula (III) 2C (C~l)m HN Cll - COR
with an acid of the formula (IV) R~ II
1~2 5 (Cl~ C()OII
_ rj_ . . . _ ilA13Sb or its chemical equivalent.
Thus, the final product can be produced not only by direct acylation with an acid of formula IV hut also by~ intermediates such as ta) l~-haloalkanoic acids of the formula (V) 1 11 X - - (CH) -CH - COOH
wherein X is bromo, chloro or iodo, or (b) a tosyloxyalkanoic acid, i,e., X in formula V is tosyloxy~ (CH3 ~ -S2-) (c) a substituted acrylic acid of the formula 14 IRl (VI) CH - C COOH
The product of this acylation is then subjected to displacement or addition with the anion of a thiol or thioacid of the formula (VII) R2 SH
Acylation can also be effected with a thiolactone of the formula (VIII) (fH)n CH
S =O
wherein n is 1 or 2, or a mercaptoalkanoic acid of the formula IR4 IRl (IX) Y S -(CH)n CH COR
, ~IA~35i~
wherein Y is R2 or, in adclition, i~ d product of forml~la I
whereill R2 is hydro~en is desired, then Y can also be a protecting group such as ~a) C~i30~ C~l2-, (b) ~ , (c) CH3CONHC~l2, (cl) ~-O-C-C~I-(C~I) -S- or other sulfur protecting group. "Depro-tection" can be effected by conventional means such as treatment with hot trifluoroacetic acid, cold trifluoromethanesulfonic acid, mercuric acetate, sodium in liquid ammonia, zinc and hydrochloric acid or the like. For a review oE these methods see Methoden der Organischen Chemie (Houben-Weyl), Vol. XV, part I, page 736 et seq. (197~) When the acid oE forlllula IV is used as the acylatinc agent, th~ acylation can be effected in the presence of a coupling agent like dicyclohexycarbodiimide or the like, or the acid ean be activated by formation of its mixed annydride, sy~netrical anhydride, acid chloride, acid ester or use of Woodward reagent ~, N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline or the like. For a review of the methods for acylation, see Methoden der Orc~anischen Chemie 2Q (Houben-Weyl), Vol. XV, part II, page 1 et seq. (1974).
Compounds of formula III include, for example, proline, hydroxyproline, 4-methylproline, pipecolic acid, 5-hydroxypipecolic acid, azetidine-2-carboxylic acid, their lower alkyl esters and the like. The acylation of SUC}l eompounds is described in greater detail below.
Aeeording to a preferred method for producinc3 compounds of formula I, espeeially wherein R2 is R5-CO-, an acid or ester of formula III is coupled with a haloalkanoic acid of the formula _ .
-IIA135b (V) i~ L
X-(Cil)n- Ci~ COOii wherein X is a halocJen, prefera~ly chlorine or bromine. This can be effected by one of the kno~-~n procedures in which the acid IV is activated, prior to reaction with the acid III, involving formation of a mixed anhydride, symmetrical anhydride, acid chloride, active estcr, or use of ~ooclward reagent K, EEDQ
(N-ethoxycarbonyl-2-ethoxy-1,2-dihydroxyquinoline) or the like.
The product of this reaction is a compound of the formula (X) 13 14 1l 21 (Cil)m .
X - ~C~ -C~l- CO - N- CII-COR
The product of formula X is subjectcd to a ciis~laccmc.~nt reaction with the anion of a thioacid oE the formula (VII) R2-S~]
yielding a product of the formula (XI) 14 1l 112C- (Cll)m R2- S - (Cll)n- CII-CO - N - CII-COR
l~hen R2 is R5CO, this product can thcll bc convcrteci to the product (XII) R3 ~ 2C (Cii) liS - (Cll) - C'li - CO - N CII-COR
-------by ammonolysis. When R2 is a protecting ~roup, then the compound o formula XIl can be obtained by "deprotection"
as described above. When R is an ester group (i.e., R is lower alkoxy), the ester group can be removed, e.g., when R is tert. butoxy or tert. amyloxy, by treatment oE the ester of formula XI or XII with trifluoroacetic acid and anisole to give the corresponding free acid. When other alkoxy groups are present, alkaline hydrolysis will yield the corresponding acid.
A variation of this procedure involves the use of an acrylic acid of the formula (VI) IR4 Rll CH=C-COOH
as starting material. This acrylic acid is first conver-ted to the acid halide form then made to react with a compound of formula III to obtain a compound of the for-mula (XIII) 13 IR4 IRl H2C (CH)m CH=C - CO - N CH-COR
and this intermediate is subjected to the addi-tion reac-tion with the thiol or thioacid VII as described above.
A tosyloxyalkanoic acid of the formula (XIV) R14 IRl CH3- ~ _ S020-(CH)n CH - COOH
can also be used as the agent to acylate the acid of formula III, then the acylation product is subjected to the displacement reaction, etc., as described above.
The acrylic acid of formula VI can alternatively be first made to react with the thioacid of formula VII to '!
-obtain a product of the formula (XV) R4 Rl R -S-C~-CH-COOH
which is converted to its acid halide, e.g., with thionyl chloride, then coupled to the compound of formula III and the same sequence as above then fo:Llowed.
The acid or ester of formula :[II can also be acylated with a "proteeted" form of a ~-mercaptoalkanoic aeid of the formula (XVI) R4 Rl R8~S~(CH)n~CH~CH
whe ein R8 is the "protecting" group. Such "protecting"
groups can take the form deseribed above.
Following the aeylation, the produet ean be "depro-teeted" by one of the known methods referred to above.
Still another aeylating agent ean take the form of a thiolaetone, e.g., ~-propiothiolaetone, ~-methyl-~-pro-piothiolaetone or the like.
Additional details of preferred modes of produeing eompounds of this invention ean be found in the following and in the speeifie examples.
Aecording to a partieularly preferred modifieation, the aeid or ester of formula III is acylated with a halo-alkanoyl halide of the formula (XVII) R4 Rl X-(CH)n CH-COX
wherein eaeh X is independently a halogen, preferably ehlorine or bromine, Rl is hydrogen, lower alkyl or phenyl-lower alkyl and n is 0, 1 or 2. ~his reaction is effected in an alka-line medium, e.g., dilute alkali metal hyclroxide solution, alkali metal dicarbonate or alkali metal carbonate solution at a reduced temperature, e.g., about 0 to 15C. The re-action product is subjected to displacement with the anion of the thiol or thio acid of the formula ~II above, also in alkaline medium, preferably alkali metal carbonate so-lution, and then worked up in conventional manner. The product of this reaction, wherein R2 of formula I is R5-CO, is converted to the product wherein R2 is hydrogen by ammonolysis, e.g., alcoholic ammonia or concentrated ammonium hydroxide solution, or alkaline hydrolysis, e.g., with aqueous metal hydroxide. When an acid of formula III is used as starting material, the final product ob-tained as the free carboxylic acid can then be converted to its ester, for example by esterification with a dia-zoalkane, like diazomethane, l-alkyl-3-p-tolyl-triazene, like l-n-butyl-3-p-tolyltriazene or the like. l`reatment of an ester, preferably the methyl ester, with an alco-holic ammonia solution, converts the free acid to theamide, i.e., R is NH2.
According to another variation, an ester, prefer-ably the t-butyl ester, of formula III, in an anhydrous medium such as dichloromethane, tetrahydrofuran, dioxane or the like, is treated with a thioalkanoic acid of the formula (XVIII) 1l R2-S-(CH2)n - CH ~ COOH
in the presence of dicyclohexylcarbodiimide, N,N'-car-bonyl-bisimidazo]e, ethoxyacetylene, diphenylphosphoryl azide or similar coupling agents at a temperature in the range of about 0 to 10C. The ester group (R) can then be removed, for ., .~, --11--.
.3 ~
e.~ample, by treatment with trifluoroacetic acid and ani-sole at about room temperature.
When an ester of formula III (e.g., R is lower al-koxy, especially, t-butoxy) is acylated with a thiolactone, e.g., ~-propiothiolactone, ~-me-thyl-~-propiothiolactone or the like, the reaction can be eEfected in an anhydrous solvent like tetrahydrofuran, dioxane, methylene chloride or the like at about 0 C. to about room temperature. The ester group can be removed with anisole and trifluorace-tic acid as described above. M
In similar manner, when R2 is R5-M-C-, products of formula I having this substituent are formed by reacting a compound of formula XII with the halogenated compound (XIX) M
or alternatively reacting a compound of formula X with an alkali metal salt or alkaline earth metal salt of the formula (XX~ M
R5-M-C-S-Me wherein Me represents the alkali metal or alkaline earth metal. M
When R2 is R5-NH-C-, products of formula I having - this substituent are produced by reacting a compound of formula XII with the appropriately substituted isocyanate or isothiocyanate of the formula (XXI) R5-N=C=M
Alternatively, the same products can be produced by coup-ling an acid of the formula ~ -12-:i~l ~5~
HA135b (XXII) 1'1 R~ IRl R5-N}I-C-S-(CII)-- CH-COOH
with an amino acid of formula III.
Compounds oE formula I, wherein R2 is lower alkyl, phenyl, substituted phenyl, phenyl-lower alkyl, -triphenyl-lower alkyl, lower alkylthiomethyl or phenyl-lower alkylthiome-thyl are produced by reacting a compound or formula XII with the corresponding halide R2X or by reacting a compound of formula X
with the corresponding thiol R2SH in the same manner as described above.
When R2 is lower alkanoylamidomethyl, -the product of Eormula I is produced by condensing a compound of formula XII
with the corresponding hydroxymethyl-lower alkanoylamide of the formula ~XXIII) lower alkyl-CO-NI~CH2OH
in the presence of an acid catalyst like trifluoroacetic acid.
Products of formula I wherein R2 is R6-S can be prepared by any of the known methods for the synthesis of mi~ed disulfides, e.g., by the reaction of a compound of formula(XII)with a thiosulfinate(xxIv), thiosulfonate (XXV), sulfenyl halide (XXVI), thiosulfa-te (XXVII) or sulfenyl thiocyanate (XXVIII) O
tXXIV) R6-S-S-R6 , tXXV) R6-S-S-R6 , (XXVI) R6S-X, O i, (XXVII) R6-S-SO3~1, (XXVIII) R6-S-SCN
In the particular case wherein R7 is l4 IRl CH2 - (IcH)n -S(O)--- (CH) -- CH _ CO - N--- CH-COR, .
~ }IA135b R, Rl, R3 and R4 are t~e same as -the corresponding substituents in formula I and p is O, the symmetrical disulfides can be obtained by direct oxidation of a compound of formula XII
with iodine. When p is 1 or 2, such products are obtained by the stepwise oxidation of the corresponding compound wherein p is O. Mixed disulfides are obtained by the modification shown in the examples.
Products of formula I have one or more asymmetric carbons.
When Rl, R3 or R~ is other than hydrogen the carbon to which it is attached is asymmetric. These carbon atoms are indicated by an asterisk in formula I. The compounds accordingly exist in stereoisomeric forms or in racemic mixtures thereof. All of these are within the scope of the invention. The above described synthesis can utilize the racemate or one of the enantiomers as starting material.
When the racemic starting material is used in the synthetic procedure, the stereoisomers obtained in the product can be separated by conventional chromatographic or fractional crystallization methods. In general, the L-isomer with respect to the carbon of the amino acid constitutes the preferred isomeric form. Also the D-isomer with respect to the a-carbon in the acyl side chain (i.e., the carbon bearing Rl) is preferred.
The compounds of this invention form basic salts with various inorganic and organis bases which are also within the scope af the invention. Such salts include ammonium salts, alkali metal salts like sodium and potassium salts ~which are preferred), alkaline earth metal salts like the calcium and magnesium salts, salts with organic bases, e.g., dicyclohe~ylamine salt, benzathine, N-methyl-D-glucamine, ;
~A135b hydrabamine salts, salts with amino acids like arginine, I
lysine and the like. The non-toxic, physiologically acceptable salts are preferred, although other salts are also useful, e.g., in isolating or purifyinc~ the product, as illustrated in the examples in the case of tihe dicyclohexylamine salt.
The salts are formed in conventional manner by reacting the free acid form of the product with one or more equivalents of the appropriate base providing the desired cation in a solvent or medium in which the salt is insoluble, or in water and removing the water by freeze drying. By neutralizing the salt with an insoluble acid like a cation exchange resin in the hydrogen form (e.g., polystyrene sulfonic acid resin like Dowex 50) or with an aqueous acid and extraction with an organic solvent, e.g., ethyl acetate, dichloromethane or the like, the free acid form can be obtained, and, if desired, another salt formed.
. -- --Additional experimelltal details are found in the examples which are preferred embodiments and also serve as models for the preparation of other members of the ~roup.
The compounds of this invention inhibit the ~onver-sion of the decapeptide angiotensin I to angiotensin II
and therefore are useful in reducing or relieving angio-tensin related hypertension. The action of the enzyme re-nin on angiotensinogen, a pseudoglobulin in blood plasma, produces angiotensin I. Angiotensin I is converted by angiotensin converting enzyme (ACE) to angiotensin II.
The latter is an active pressor substance which has been implicated as the causative agent in various forms of hypertension in various mammalian species, e.g., rats and dogs. The compounds of this invention intervene in the angiotensin(renin)~angiotensin I ~angiotensin II sequence by inhibiting angiotensin converting enzyme and reducing or eliminating the formation of the pressor substance angiotensin II. Thus by the administration of a composi-tion containing one or a combination of compounds of for-mula I or physiologically acceptable salt thereof, angio-tensin dependent hypertension in the species of mammal suffering therefrom is alleviated. A single dose, or pre-~ferably two to four divided daily doses, provided on a ,~
basis of about 0.1 to 100 mg. per kilogram per day, pre-ferably about 1 to 50 mg. per kilogram per day is appro-priate to reduced blood pressure as indicated in the ani-mal model experiments described by S.L. Engel, T.R.
Schaeffer, M.H. Waugh and B. Rubin, Proc. Soc. Exp. Biol.
Med. 143, 483 (1973). The substance is preferably admin-istered orally, but parenteral routes such as subcutaneous,intramuscular, intravenous or -16- -~
intraperitoneal can also be employed.
~ ne cc~pounds of this invention can be utilized to aehieve the reduction of blood pressure by formulating in cc~positions sueh as tablets, eapsules or elixirs for oral acbministration or in s-terile solutions or suspensions for parenteral administration. About 10 to 500 mg. of a compound or mixture of compounds of formula I or physio-logieally acceptable salt is compounded with a physiologieally accep-table vehiele, carrier, excipient, binder, preservative, stabilizer, flavor, ete., in a unit dosage form as ealled for by accepted phar-maceutieal praetiee. m e amount of active substance in these cc~positions or preparations is sueh that a suitable dosage in the range indicated is obtained.
Illustrative of the adjuvants whieh may be ineorporated in tablets, eapsules and the like are the follc~ing: a binder such as gum tragaeanth, aeaeia, eorn stareh or gelatin; an exeipient such as diealeium phosphate; a disintegrating agent sueh as eorn stareh, potato stareh, alginie aeid and the like; a lubrieant sueh as magnesium stearate; a sweetening agent such as sucrose, laetose or ~ !
saeeharin; a flavoring agent such as peppermint, oil of winter-green or eherry. When the dosage unit form is a eapsule, it may eontain in addition to materials of the above type a liquid earrier sueh as a fatty oil. Various other materials may be present as eoatings or to otherwise modify the physieal form of the dosage unit. For instanee, tablets may be eoated with shellae, sugar or both. A syrup or elixir may eontain the aetive compound, suerose as a sweetening agent, methyl and propyl parabens as preservatives, a dye and a flavoring such as eherry or orange flavor.
Sterile eompositions for injeetion ean be formulated according to conventional pharmaceutical practice by dis-solving or suspending the active substance in a vehicle such as water for injection, a naturally occurring vege-table oil like sesame oil, coconut oil, peanut oil, cotton-seed oil, etc., or a synthetic Eatty vehicle like e-thyl oleate or the like. Buffers, preservatives, antioxidants and the like can be incorporated as required.
The following examples are illustrative of the in-ven~ion and cons-titute especially preferred embodiments t All temperatures are in degrees celsius.
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E~ample 1 _ 1-(2-Benzoylthioacetyl)-L-Proline L-Proline (5.75 g.) is dissolved in N sodium hydr-oxide (50 ml.) and the solution is chilled in an ice-water bath. Sodium hydroxide 2N (26 ml.~ and chloroacetyl chlo-ride (5.65 g.) are added and the mixture is stirred vigo-rously at room temperature for three hours. A suspension of thiobenzoic acid (7.5 g.) and potassium carbonate (4.8 g.) in water (50 ml.) is added. After 18 hours stirring at room temperature, the reaction mixture is acidified and extracted with ethyl acetate. The ethyl acetate layer is washed with water, dried over magnesium sulfate and con-centrated to dryness in vacuo. The residue (14.6 g.) is dissolved in ethyl acetate (150 ml.) and dicyclohexylamine (11 ml.) is added. The crystals are filtered and recry-stallized from ethyl acetate, yield 5.7 g. m.p. 151-152.
To convert the salt to the acid, the crystals are dissol-ved in a mixture of 5~ aqueous potassium bisulfate (100 ml.) and ethyl acetate (300 ml.). The organic phase is washed onee with water, dried over magnesium sulfate and eoncentrated to dryness in vacuo, yield 3.45 g.
Example 2 1-(2-Mercaptoacetyl)-L-Proline 1-(2-Benzoylthioacetyl)-L-proline (3O4 g.) is dis-solved in a mixture of water (10.5 ml.) and concentrated ammonia (6.4 ml.~. After one hour, the reaction mixture is diluted with water and filtered. The filtrate is ex-traeted with ethyl aeetate and then aeidified with eon-eentrated hydrochloric acid, saturated with sodium chlo-ride and extracted twice with ethyl acetate. The ethylacetate extraets are washed with saturated sodium ehlo-ride and eoneentrated to dryness, yield 1.5 g.
,~
The product, 1-(2-mercaptoacetyl)-L-proline is crystallized from ethyl acetate (m.p. 133-135) .
Exam~le 3 1-(2-Benzoylthioacetyl)-L-Proline Methyl Ester 1-(2-Benzoylthioacetyl)-L-proline o~tained in Example 1, is dissolved in methanol and an ethereal solution of diazomethane is added until there is a persistent yellow color. After 15 minutes, a few drops of acetic acid are added and the solvent is removed in vacuo to obtain 1-(2-benzoylthioacetyl)-L-proline methyl ester.
Example 4 1-(2-Mercaptoacetyl)-L-Proline Amide The product of Example 3 is dissolved in 10~ metha-nolic ammonia and the solution is stored at room tempera-ture in a pressure bottle. When thin layer chromatogra-phic analysis indicates that the two ester functions have been ammonolyzed, the reaction mixture is concentrated to dryness to obtain l-(2-mercaptoacetyl)-L-proline amide.
Example 5 1-(2-Benzoylthioacetyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example 1, 1-(2-benzoylthioacetyl~-L-hydroxyproline is obtained.
Example 6 1-(2-Mercaptoacetyl~-L-Hydroxyproline By treating the product of Example 5 with ammonia as in Example 2, 1-t2-mercaptoacetyl)-L-hydroxyproline is obtained.
Example 7 1-(2-Benzoylthioacetyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid for the L-proline in the procedure of Example 1, 1-(2-benzoylthio-acetyl)-L-azetidine-2-carboxylic acid is obtained.
Example 8 1-(2-Mercaptoace-tyl)-L-Aze idine-2-Carboxylic ~cid By treating the product of ~xample 7 with ammonia as in Example 2, 1-(2-mercaptoacetyl)-L-azetidine-2-car-boxylic acid is obtained.
Example 9 1-(2-Benzoylthioacetyl)-L-pipecolic Acid , .
By substituting L-pipecolic acid for the L-proline in the procedure of Example 1, 1-(2-benzoylthioacetyl)-L-pipecolic acid is obtained.
Example 10 1-(2-Mercaptoacetyl)-L-Pipecolic Acid By treating the product of Example 9 with ammonia as in Example 2, 1-(2-mercaptoacetyl)-L-pipecolic acid is obtained.
Example 11 l-t2-Benzoylthiopropanoyl)-L-Proline L-Proline (5.75 g.) is dissolved in aqueous N so-dium hydroxide (50 ml.) and the solution is chilled in an ice bath with stirring. 2N sodium hydroxide (25 ml.) and 2-bromopropionyl chloride (8.57 g.) are added in that order and the mixture is removed from the ice bath and stirred at room temperature for one hour. A mixture of thiobenzoic acid (7.5 g.) and potassium carbonate (4.8 g.) in water (50 ml.) is added and the mixture is stirred over-night at room temperature. After acidification with con-centrated hydrochloric acid, the aqueous solution is ex-tracted with ethy:l acetate and the organic phase is washed with water, dried and concentrated to dryness. The resid~le (14.7 g.) is chroma-to~raphed on a column of 440 g. o~ silica gel with a mix-ture of benzene-acetic acid (7:1). The fractions contain-ing the desired material are pooled, concentrated to dry-ness, and the residue is precipitated -twice with ether-hexane and converted to a dicyclohexylamine salt in ether-hexane, yield 9.4 g. m.p., (142) 148-156. The dicyclo-hexylamine salt is converted back to the acid as in Exam-ple 1, yield 5.7 g.
Example 12 1-(2-Mercaptopropanoyl)-L-Proline 1-(2-Benzoylthiopropanoyl)-L-proline (5.7 g.) is dissolved in a mixture of water (12 ml.) and concentrated ammonium hydroxide (9 ml.) with stirring. After one hour, the mixture is diluted with water (10 ml.) and filtered.
The filtrate is extracted twice with ethyl acetate, con-centrated to one-third of the original volume, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with satura-ted sodium chloride, dried and concentrated to dryness invacuo. The residue, 1-(2-mercaptopropanoyl)-L-proline, is crystallized from ethyl acetate-hexane, yield 3 g., m.p.
(105) 116-120.
Example 13 1-(3-Benzoylthiopropanoyl)-L-Proline L-Proline (5.75 g.) is dissolved in normal sodium hydroxide (50 ml.) and the solution is chilled in an ice bath. 3-Bromopropionyl chloride (8.5 g.) and 2N sodium hydroxide (27 ml.) are added and the mixture is stirred for 10 minutes in the ice bath and three hours at room temperature. A suspension of thiobenzoic acid (7.5 g.) and potassium ca:rbonate (4 5 g.) in water (50 ml.) is added and the mixture is stirred .
.
v~7 for 18 hours at room tempera-ture. After acidification with concentrated hydrochloric acid, the aqueous phase is ex-tracted twice with ethyl acetate. The organic layers are dried over magnesium sulEate and concentrated to dryness in vacuo to obtain l-(3-benzoylthiopropanoyl)-L-proline, yield 7.1 g., m.p. 101-102 (ethyl acetate-hexane).
Example 14 L-Proline tert.-butyl ester L-Proline (230 g.) is dissolved in a mixture of water (1 1.) and 5 N sodium hydroxide (400 ml.). The so-lution is chilled in an ice bath, and under vigorous stir-ring, 5 N sodium hydroxide (460 ml.) and benzyloxycarbonyl chloride (340 ml.) are added in five equal aliquots during a half hour period. After one hour stirring at room tem-perature, the mixture is extracted twice with ether and acidified with concentrated hydrochloric -acid. The preci-pitate is filtered and dried. Yield 442 g., m.p. 78-80.
The benzyloxycarbonyl-L-proline thus obtained (180 g.~ is dissolved in a mixture of dichloromethane (300 ml.), liquid isobutylene (800 ml.) and concentrated sulfuric acid (7.2 ml.). The solution is shaken in a pressure bottle for 72 hours. The pressure is released, the isobutylene is allowed to evaporate and the solution is washed with 5 sodium carbonate, water, dried over magnesium sulfate and concentrated to dryness in vacuo, to obtain benzyloxycar-bonyl-L-proline tert. butyl ester, yield 205 g.
Benzoyloxycarbonyl-L-proline tert. butyl ester (205 g.) is dissolved in absolute ethanol (1.2 1) and hy-drogenated at normal pressure with 10~ Pd on carbon (10 g.) until only a trace of carbon dioxide is observed in the hydrogen exit gas ~4 hours). The catalyst is filtered off and the Eiltrate is c~ncentrated in vacuo at 30 mm. H~. The residue is distilled in vacuo, to obtain L-proline tert.butyl ester, b.p. 50-51.
lmm Example 15 1-(3-~cetylthiopropanoyl)-L-Proline tert-butyl Ester L-Proline tert-butyl ester (5.13 g.) is dissolved in dichloromethane (40 ml.) and the solution is chilled in an ice-water bath. A solution of dicyclohexylcarbodiimide (6.18 g.) in dichloromethane (20 ml.) is added followed immediately by 3-acetylthiopropionic acid (4.45 g.).
After 15 minutes stirring in the ice-water bath and 16 hours at room temperature, the precipitate is filtered off and the filtrate is concentrated to dryness in vacuo.
The residue is dissolved in ethyl acetate and washed neu-tral. The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo to obtain 9.8 g. of 1-(3-acetylthiopropanoyl)-L-proline tert-butyl ester.
Example 16 1-(3-Acetylthiopropanoyl)-L-Proline 1-(3-Acetylthiopropanoyl)-L-proline-t-butyl ester (4.7 g.) is dissolved in a mixture of anisole (34 ml.) and trifluoroacetic acid (68 ml.) and the mixture is kept at room temperature for one hour. The solvents are removed in vacuo and the residue is precipitated from ether-hexane several times. The residue (3.5 g.) is dis-solved in acetonitrile (25 ml.) and dicyclohexylamine (2.8 ml.) is added. The crystalline salt is filtered and recrystallized from isopropanol. Yield 3.8 g.. m.p.
176-177. The salt is reconverted to 1-(3-acetylthio-propanoyl)-L-proline as in --?4--~ r~ ~
E.~ample 1, yield 1 25 g., m.p. 89-90 (ethyl acetate-he~ane).
Example 17 1-(3-Mercaptopropanoyl)-L-proline tert-butyl Ester To a solution of L-proline tert-butyl ester (3.42 g.) in dry tetrahydrofuran (10 ml.) chilled in an ice bath, propiothiolactone (1.76 g.) is added. After 5 minutes storage in the ice bath and three hours at room -temperature, the reaction mixture is diluted with ethyl acetate (200 ml.) and washed with 5~ potassium bisulfate, and water.
The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo. The residue 1-(3-mer-captopropanoyl)-L-proline tert-butyl ester is crystallized from ether-hexane, yield 3.7 g., m.p. 57-58.
Example 18 1-(3-~lercaptopropanoyl)-L-Proline Procedure A
1-(3-Benzoylthiopropanoyl)-L-proline (4.9 g.) is dissolved in a mixture of water (8 ml.) and concentrated ammonium hydroxide (5.6 ml.) and the solution is stored with stirring under argon for one hour. The reaction mix-ture is diluted with water, filtered, and the filtrate is extracted with ethyl acetate. The aqueous phase is acid-ified with concentrated hydrochloric acid, saturated with sodium chloride and extracted with ethyl acetate. The organic layers are washed with saturated sodium chloride, dried over magnesium sulfate, and concen-trated to dryness in vacuo. The residue, 1-(3-mercaptopropanoyl)-L-proline, is crystallized from ethyl acetate hexane, yield 2.5 g., m.p. 68-70.
Procedure B
.
l-t3-Acetylthiopropanoyl)-L-proline (0.8 g.) is dis-solved :: `
in 5.5 N methanolic ammonia (5 ml.) and the solution kept under ar~on at room temperature. After 2 hours the solvent is removed in vacuo, the residue is dissolved in water and applied to an ion exchange column on the H cycle [Dowex 50 (Analytical grade)] and eluted with water. The fractions that give thiol positive reaction are pooled and concentra-ted to dryness, yield 0.6 g. This product is crystallized from ethyl acetate-hexane as in Procedure A to obtain 1-(3-mercaptopropanoyl)-L-proline.
Procedure C
1-(3-Mercaptopropanoyl)-L-proline t-butyl ester (2.3 g.) is dissolved in a mixture of anisole (20 ml.) and tri-fluoroacetic acid (45 ml.). After one hour storage at room temperature under argon, the reaction mixture is con-centrated to dryness in vacuo and the residue precipitated from ethyl acetate-hexane several times. The residue (1.9 g.) is dissolved in ethyl acetate t30 ml.) and di-cyclohexylamine (1.85 ml.) is added. The crystalline salt is filtered and recrystallized from isopropanol, yield 2 g., m.p. 187-188.
The salt is converted to the acid as in Example 1, yield 1.3 g. The product is crystallized from ethyl ace-tate hexane as in Procedure A.
Salts sOaium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.) is dis-solved in a mixture of water (2.5 ml.) and N sodium hy-droxide (2.5 ml.). The solution is freeze dried to ob-tain the sodium salt.
Magnesium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.), mag-nesium oxide (49.5 mg.), and water ~10 ml.) are stirred ~ -26-with slight heating until complete solution is obtained.
Then the solvent is removed by freeze drying to obtain the magnesium salt.
Calcium 1-(3-Mercaptopropanoyl)-2-proline (S00 mg.) is dissolved in a mixture of calcium hydroxide (91 mg.) and water (10 ml.), and the solu-tion is freeze dried to obtain the calcium salt.
Potassium 1-(3-Mercaptopropanoyl)-L-proline (500 mg.) is dissolved in a mixture of potassium bicarbonate (246 mg.) and water (10 ml.) and freeze dried to obtain the potassium salt.
N-Methyl-D-Glucamine 1-(3-Mercaptopropanoyl)-L-proline(500 mg.) and N-methyl-D-glucamine (480 mg.) are dissolved in water (10 ml.) - and freeze dried to obtain the N-methyl-D-glucamine salt.
Example 19 1-(3-Mercaptopropanoyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example 11 and then treating the pro-duct by Procedure A of Example 18, 1-(3-benzoylthiopropan-oyl)-L-hydroxyproline and 1-(3-mercaptopropanoyl)-L-hydr-, oxyproline, respectively, are obtained.
Example 20 1-(3-Mercaptopropanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid tert-butyl ester (prepaxed by substituting L-azetidine-2-carboxylic acid for the proline in Example 14) for the L-proline tert-butyl ester in the procedure of Example 15, ~AI
treating the product as in Example 16 and the 1-(3-acetyl-thiopropanoyl)-L-azetidine-2-carboxylic acid thus obtained by Procedure B of Example 18, 1-(3-acetylthiopropanoyl)-L-azetidine-2-carboxylic acid tert-butyl ester and 1-(3-mercaptopropanoyl)-L-azetidine-2-carboxylic acid, respec-tively, are obtained.
Example 21 1-(3-Mercaptopropanoyl)-L-Pipecolic Acid By substituting L-pipecolic acid tert-bu-tyl ester (prepared by substituting L-pipecolic acid for the L-pro-line in Example 14) for the L-proline tert-butyl ester in the procedure of Example 15 and treating the product by Procedure C of Example 18, 1-(3-mercaptopropanoyl)-L-pipecolic acid tert-butyl ester and l-(3-mercaptopropan-oyl)-L-pipecolic acid, respectively, are obtained.
Example 22 1-(3-Mercaptopropanoyl)-4-Methyl-L-Proline By substituting 4-methyl-L-proline for L-proline in the procedure of Example 13 and then treating the product by Procedure A of Example 18, 1-(3-benzoylthiopropanoyl)-4-methyl-L-proline and 1-(3-mercaptopropanoyl)-4-methyl-L-proline, are obtained.
Example 23 1-(3-Mercaptopropanoyl)-5-Hydroxy-L-Pipecolic Acid By substituting 5-hydroxy-L-pipecolic acid for L-proline in the procedure of Example 13 and then treating the product by the Procedure A of Example 18, 1-(3-ben-zoylthiopropanoyl)-5-hydroxy-L-pipecolic, and 1-(3-mercaptopropanoyl)-5-hydroxy-L-pipecolic acid are obtained.
Example 24 1-(3-Mercaptopropano~l)-D-Proline By substituting D-proline for L-proline in the pro-cedure of E~ample 13 and then treating the product by Pro-cedure A of Example 18, 1-(3-benzoylthiopropanoyl)-D-proline and l-(3-mercaptopropanoyl)-D-proline, m.p. 68-70, are ob-tained.
Example 25
3-Acetylthio-2-Methylpropanoic Acid A mixture of thioacetic acid (50 g.) and methacry-lie acid (40.7 g.) is heated on the steam bath for one hour and then stored at room temperature for 18 hours. After eonrirming by nmr spectroscopy that complete reaction of the methacrylic acid has been achieved, the reaction mix-ture is distilled in vacuo and the desired 3-acetylthio-2-methylpropanoic acid is separated in the fraction with boiling point 128.5-131 (2.6 mmHg.), yield 64 g.
Example 26 3-Benzoylthio-2-Methylpropanoic Acid By substituting thiobenzoic acid for the thioace-tie aeid in the proeedure of Example 25, 3-benzoylthio-2-methylpropanoie acid is obtained.
Example 27 , 3-Phenylacetylthio-2-Methylpropanoie Aeid By substituting thiophenylacetic aeid for the thio-acetie aeid in the procedure of Example 25, 3-phenylacetyl thio-2-methylpropanoic aeid is obtained.
Example 28 1-(3-Aeetylthio-2-methylpropanoyl)-L-Proline tert-butyl Ester L-Proline tert-butyl ester (5.1 g.) is dissolved in diehloromethane (40 ml.) and the solution stirred and ehilled ~ -29-~t3,r~
in an ice bath. Dicyclohexylcarbodiimide t6-2 g.) dissolved in dichloromethane (15 ml.) is added followed immediately by a solution of 3-acetylthio-2-methylpropanoic acid (4.9 g.) in dichloromethane (5 ml.). After 15 minutes stirring in the ice bath and 16 hours at room t:emperature, the precipi-tate is filtered off and the filtrate is concentrated to dryness in vacuo. The residue is clissolved in ethyl acet-ate and washed neutral. The organic phase is dried over magnesium sulfate and concentrated to dryness in vacuo.
The residue 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester is purified by column chromatography (silica gel-chloroform), yield 7.9 g.
Example 29 ~-(3-Acetylthio-2-methylpropanoyl)-L-Proline Procedure A
The 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester of Example 28 (7.8 g.) is dissolved in a mixture of anisole (55 ml.) and trifluoroacetic acid (110 ml.). After one hour storage at room temperature the sol-vent is removed in vacuo and the residue is precipitated . ~several times from ether-hexane. The residue (6.8 g.) is dissolved in acetonitrile (40 ml.) and dicyclohexylamine (4.5 ml.) is added. The crystalline salt is boiled with fresh acetonitrile (100 ml.), chilled to room temperature and filtered, yield 3.8 g., m.p. (165) 187-188. This ma-terial is recrystallized from isopropanol [~]D-67 (C 1.4, EtOH). The crystalline dicyclohexylamine salt is suspen-ded in a mixture of 5~ aqueous potassium bisulfate and ethyl acetate. The organic phase is washed with water and concentrated to dryness. The residue is crystallized from ethyl acetate-hexane to yield the l-(3-acetyl-~
thio-2-D-methylpropanoyl-L-proline, m.p. 83-85[~]D5-162 (c,1.7,EtOH).
Procedure B
3-Acetylthio-2-methylpropanoic acid (8.1 g.) and thio-nyl chloride (7 g.) are mixed and the suspension is stirred for 16 hours at room temperature. The reaction mixture is concentrated to dryness and distilled in vacuo (b.p. 80).
This 3-acetylthio-2-methylpropanoic acid chloride (5.4 g.) and 2N sodium hydroxide (15 ml.) are added to a solution of L-proline (3.45 g.) in normal sodium hydroxide (30 ml.) chilled in an ice water bath. After 3 hours stirring at room temperature, the mixture is extracted with ether, the ~-aqueous pnase is acidified and extracted with ethyl acetate.
The organic phase is dried over magnesium sulfate and con-centrated to dryness to obtain l-(3-acetylthio-2-DL-methyl-propanoyl-L-proline).
Procedure C
Methacryloyl chloride (4.16 g.) is added to a solu-tion of L-proline (3.45 g.) in a mixture of water (100 ml.) and sodium bicarbonate (12 g.j chilled in an ice water bath, with vigorous stirring. When the addition is com-pleted, the mixture is stirred at room temperature for two hours, and then extracted with ether. The aqueous phase is acidified with N hydrochloric acid and extracted with ethyl acetate. The organic phase is concentrated to dry-ness in vacuo, the residue is mixed with thiolacetic acid (3.5 g.), a few crystals of azobisisobutyronitrile are added and the mixture is heated on the steam bath for two hours. The reaction mixture is dissolved in benzene-ace-tic acid (75:25), and applied to a column of silica gel.Elution with the same solvent mixture yields the 1-(3-ace-tylthio-2-DL-methylpropanoyl)-L-proline.
, 1~
E~.lmple 30 1-(3-Benzoylthio-2-methylpropanoyl)-L-proline tert-butyl Ester By substituting 3-benzoylthio-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the procedure of Example 28, 1-(3-benzoylthio-2-rnethylpropanoyl)-L-proline tert.butyl ester is obtained.
Example 31 1-(3-Phenylacetylthio-2-methylpropanoyl)-L-Proline tert-butyl Ester By substituting 3-phenylacetylthio-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the procedure of Example 28, 1-(3-phenylacetylthio-2-methyl-propanoyl)-L-proline tert butyl ester is obtained.
Example 32 1-~3-Benzoylthio-2-methylpropanoyl)-L-proline By substituting 1-(3-benzoylthio-2-methylpropanoyl)-L-proline tert-butyl ester for the 1-(3-acetylthio)-2-methylpropanoyl-l-proline tert-butyl ester in Procedure A
of Example 29, 1-(3-benzoylthio-2-methylpropanoyl)-L-pro-line is obtained.
Example 33 1-(3-Phenylacetylthio-2-methylpropanoyl)-L-proline By substituting 1-(3-phenylacetylthio-2-methylpro-panoyl)-L-proline tert-butyl ester for 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester in Procedure A of Example 29, 1-(3-phenylacetylthio-2-methylpropanoyl)-L-proline is obtained.
Example 34 1-(3-Mercapto-2-D-methylpropanoyl)-L-Proline 1-(3-Mercapto-2-methylpropanoyl)-L-proline is ob-tained by treating the product of each of Examples 29, 32 and 33 as follows:
~i ., ,., ,,, .. , -- ~ .
The thioester t0.85 g.) is dissolved in 5.5 N metha-nolic ammonia and the solution is kept at room temperature for 2 hours. The solvent is removed in v cuo and -the re-sidue is dissolved in water, applied to a ion exchange co- -lumn on the H cycle (Dowex 50, analytical grade) and eluted with water. The fractions that give positive thiol reaction are pooled and freeze dried. The residue is cry-stallized from ethyl acetate-hexane, yield 0.3 g. The 1-(3-mercapto-2-D-methylpropanoyl-L-proline has m.p. 103-104, ~']D -1,1 (C,2,EtOH).
Example 35 1-~3-Acetylthio-2-methylpropanoyl)-L-Proline Methyl Ester 1-(3-Acetylthio-2-methylpropanoyl)-L-proline is re-acted with an ethereal solution of diazomethane according to the procedure described in Example 3 to obtain 1-(3-acetylthio-2-methylpropanoyl)-L-proline methyl ester.
Example 36 1-(3-Mercapto-2-methylpropanoyl)-L-Proline amide By substituting 1-(3-acetylthio-2-methylpropanoyl)-L-proline methyl ester in the procedure of Example 4, 1-(3-mercapto-2-methylpropanoyl)-L-proline amide is obtained.
Example 37 3-Acetylthio-2-Benzylpropanoic Acid By substituting 2-benzylacrylic acid for the metha-crylic acid in the procedure of Example 25, 3-acetylthio-2-benzylpropanoic acid is obtained.
Example 38 1-(3-Acetylthio-2-benzylpropanoyl)-L-Proline tert-butyl Ester By substituting 3-acetylthio-2-benzylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the proce-dure of ,j . :~
l3~l35b E,~:3mple 2~ (3-acetylthio-2-benzylpropanoyl)-L-proline tert-butyl ester is obtained.
_xamp e 39 anoyl?-L-Proline The product of Example 38 is substituted for the 1-(3-acetylthio-2-methylpropanoyl-L-proline tert-butyl ester in the Procedure A of Example 29 to obtain 1-(3-acetylthio-2-benzylpropanoyl)-L-proline.
Example 40_ 1-(3-Mercapto-2-benzylpropanoyl)-L-Proline 1-(3-Acetylthio-2-benzylpropanoyl)-L-pxoline is treated with methanolic ammonia according to the procedure of Example 34 to obtain 1-(3-mercapto-2-benzylpropanoyl)-L-proline as an oil, Rf = 0.47 (silica gel, benzene-acetic acid (75:25).
Example 41 1-(3-Mercapto-2-methylpropanoyl)-L-Hydroxy Proline By substituting L-hydroxy proline tert-butyl ester in the procedure of Example 28, treating the product according to Procedure A of Example 29 and then continuing as in Example 34, 1-(3-acetylthio-2-methylpropanoyl)-L-hydroxyproline tert-butyl ester, l-(3-acetylthio-2-methylpropanoyl)-L-hydroxyproline and l-(3-mercapto-2-methylpropanoyl)-L-hydroxyproline, respectively, are obtained.
Example 42 1-(3-Mercapto-2-methylpropanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid tert-butyl ester in the procedure of Example 28, treating the product according to the Procedure A of Example 29 and then continuing as ;'5.~
in Example 34, 1-(3-acetylthio-2-methylpropanoyl)-L-azetidine-2-carboxylic acid tert-butyl ester, 1,3-acetylthio-2-methyl-propanoyl)-L-azetidine-2-carboxylic acid and 1-(3-mercapto-2-methylpropanoyl-L-azetidine-2-carboxylic acid are obtained.
Example 43 1-(3-Mercapto-2-methylpropanoly)-L-Pipecollc Acid By substituting L-pipecolic acid in the procedure of Example 28, treating the product according to Procedure A of Example 29 and then continuing as in Example 34, 1-(3-acetyl-thio-2-methylpropanoyl)-L-pipecolic acid tert-butyl ester, 1-(3-acetylthio-2-methylpropanoyl)-L-pipecolic acid and 1-(3-mercapto-2-methylpropanoyl)-L-pipecolic acid, respectively, are obtained.
Example 44 1_(4-senzoylthiobutanoyl)-L-Proline To a solution of L-proline (2.88 g.) in normal sodium hydroxide (25 ml.) chilled in an ice bath, 2N sodium hydroxide (12.5 ml.) and 4-chlorobutyryl chloride (3.5 g.) are added.
The reaction mixture is stirred at room temperature for 3.5 hours and a suspension of thiobenzoic acid (3.75 g.) and potassium carbonate (2.4 g.) in water (25 ml.) is added. After overnight stirring at room temperature, the reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried over magnesium sul-fate and concentrated to dryness in vacuo. The residue is chromatographed on a column of silica gel with benzene-acetic acid (7:1). The fractions containing the desired material are pooled and concentrated to dryness, yield 1.35 g. A small aliquot of this material is dissolved in ethyl acetate and dicyclohexylamine is added until pH 8-10 (on a wet pH paper).
~i .
The dicyclohe~ylamine salt crystalli~es out, immediately, m.p.
159-161~.
E~ample 45 1-_(4-~ercaptobu-~anoyl)-L-Proline 1-(4-Benzylthiobutanoyl)-L-proline (1.08 g.) is dissolved in a mix-ture of water (4 ml.) and concentrated ammonia (2.7 ml.). After one hour stirring at room tempera-ture, the mixture is diluted with water, filtered, extracted with ethyl acetate, and the aqueous phase was concentrated in vacuo.
This ammonium salt of 1-(4-mercaptobu-tanoyl)-L-proline is puri ied by ion exchange chromatography on a column of diethyl-aminoethyl-Sephadex (cross linked dextran) with a gradient of ammonium bicarbonate, yield 0.7 g. The ammonium salt is dissolved in water ~2 ml.) and applied to a column of Dowex 50 sul~onic acid resin analytical grade in the hydrogen form, and the free acid is eluted with water. The fractions containing the desired material (sulfhydryl reagent and carboxyl reagent positive) are pooled and freeze dried to obtain l-(4-mercaptobutanoyl)-L-proline. The dicyclohexyl ammonium salt is produced by the procedure of Example 44, m.p. 157-158.
Example 46
Example 26 3-Benzoylthio-2-Methylpropanoic Acid By substituting thiobenzoic acid for the thioace-tie aeid in the proeedure of Example 25, 3-benzoylthio-2-methylpropanoie acid is obtained.
Example 27 , 3-Phenylacetylthio-2-Methylpropanoie Aeid By substituting thiophenylacetic aeid for the thio-acetie aeid in the procedure of Example 25, 3-phenylacetyl thio-2-methylpropanoic aeid is obtained.
Example 28 1-(3-Aeetylthio-2-methylpropanoyl)-L-Proline tert-butyl Ester L-Proline tert-butyl ester (5.1 g.) is dissolved in diehloromethane (40 ml.) and the solution stirred and ehilled ~ -29-~t3,r~
in an ice bath. Dicyclohexylcarbodiimide t6-2 g.) dissolved in dichloromethane (15 ml.) is added followed immediately by a solution of 3-acetylthio-2-methylpropanoic acid (4.9 g.) in dichloromethane (5 ml.). After 15 minutes stirring in the ice bath and 16 hours at room t:emperature, the precipi-tate is filtered off and the filtrate is concentrated to dryness in vacuo. The residue is clissolved in ethyl acet-ate and washed neutral. The organic phase is dried over magnesium sulfate and concentrated to dryness in vacuo.
The residue 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester is purified by column chromatography (silica gel-chloroform), yield 7.9 g.
Example 29 ~-(3-Acetylthio-2-methylpropanoyl)-L-Proline Procedure A
The 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester of Example 28 (7.8 g.) is dissolved in a mixture of anisole (55 ml.) and trifluoroacetic acid (110 ml.). After one hour storage at room temperature the sol-vent is removed in vacuo and the residue is precipitated . ~several times from ether-hexane. The residue (6.8 g.) is dissolved in acetonitrile (40 ml.) and dicyclohexylamine (4.5 ml.) is added. The crystalline salt is boiled with fresh acetonitrile (100 ml.), chilled to room temperature and filtered, yield 3.8 g., m.p. (165) 187-188. This ma-terial is recrystallized from isopropanol [~]D-67 (C 1.4, EtOH). The crystalline dicyclohexylamine salt is suspen-ded in a mixture of 5~ aqueous potassium bisulfate and ethyl acetate. The organic phase is washed with water and concentrated to dryness. The residue is crystallized from ethyl acetate-hexane to yield the l-(3-acetyl-~
thio-2-D-methylpropanoyl-L-proline, m.p. 83-85[~]D5-162 (c,1.7,EtOH).
Procedure B
3-Acetylthio-2-methylpropanoic acid (8.1 g.) and thio-nyl chloride (7 g.) are mixed and the suspension is stirred for 16 hours at room temperature. The reaction mixture is concentrated to dryness and distilled in vacuo (b.p. 80).
This 3-acetylthio-2-methylpropanoic acid chloride (5.4 g.) and 2N sodium hydroxide (15 ml.) are added to a solution of L-proline (3.45 g.) in normal sodium hydroxide (30 ml.) chilled in an ice water bath. After 3 hours stirring at room temperature, the mixture is extracted with ether, the ~-aqueous pnase is acidified and extracted with ethyl acetate.
The organic phase is dried over magnesium sulfate and con-centrated to dryness to obtain l-(3-acetylthio-2-DL-methyl-propanoyl-L-proline).
Procedure C
Methacryloyl chloride (4.16 g.) is added to a solu-tion of L-proline (3.45 g.) in a mixture of water (100 ml.) and sodium bicarbonate (12 g.j chilled in an ice water bath, with vigorous stirring. When the addition is com-pleted, the mixture is stirred at room temperature for two hours, and then extracted with ether. The aqueous phase is acidified with N hydrochloric acid and extracted with ethyl acetate. The organic phase is concentrated to dry-ness in vacuo, the residue is mixed with thiolacetic acid (3.5 g.), a few crystals of azobisisobutyronitrile are added and the mixture is heated on the steam bath for two hours. The reaction mixture is dissolved in benzene-ace-tic acid (75:25), and applied to a column of silica gel.Elution with the same solvent mixture yields the 1-(3-ace-tylthio-2-DL-methylpropanoyl)-L-proline.
, 1~
E~.lmple 30 1-(3-Benzoylthio-2-methylpropanoyl)-L-proline tert-butyl Ester By substituting 3-benzoylthio-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the procedure of Example 28, 1-(3-benzoylthio-2-rnethylpropanoyl)-L-proline tert.butyl ester is obtained.
Example 31 1-(3-Phenylacetylthio-2-methylpropanoyl)-L-Proline tert-butyl Ester By substituting 3-phenylacetylthio-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the procedure of Example 28, 1-(3-phenylacetylthio-2-methyl-propanoyl)-L-proline tert butyl ester is obtained.
Example 32 1-~3-Benzoylthio-2-methylpropanoyl)-L-proline By substituting 1-(3-benzoylthio-2-methylpropanoyl)-L-proline tert-butyl ester for the 1-(3-acetylthio)-2-methylpropanoyl-l-proline tert-butyl ester in Procedure A
of Example 29, 1-(3-benzoylthio-2-methylpropanoyl)-L-pro-line is obtained.
Example 33 1-(3-Phenylacetylthio-2-methylpropanoyl)-L-proline By substituting 1-(3-phenylacetylthio-2-methylpro-panoyl)-L-proline tert-butyl ester for 1-(3-acetylthio-2-methylpropanoyl)-L-proline tert-butyl ester in Procedure A of Example 29, 1-(3-phenylacetylthio-2-methylpropanoyl)-L-proline is obtained.
Example 34 1-(3-Mercapto-2-D-methylpropanoyl)-L-Proline 1-(3-Mercapto-2-methylpropanoyl)-L-proline is ob-tained by treating the product of each of Examples 29, 32 and 33 as follows:
~i ., ,., ,,, .. , -- ~ .
The thioester t0.85 g.) is dissolved in 5.5 N metha-nolic ammonia and the solution is kept at room temperature for 2 hours. The solvent is removed in v cuo and -the re-sidue is dissolved in water, applied to a ion exchange co- -lumn on the H cycle (Dowex 50, analytical grade) and eluted with water. The fractions that give positive thiol reaction are pooled and freeze dried. The residue is cry-stallized from ethyl acetate-hexane, yield 0.3 g. The 1-(3-mercapto-2-D-methylpropanoyl-L-proline has m.p. 103-104, ~']D -1,1 (C,2,EtOH).
Example 35 1-~3-Acetylthio-2-methylpropanoyl)-L-Proline Methyl Ester 1-(3-Acetylthio-2-methylpropanoyl)-L-proline is re-acted with an ethereal solution of diazomethane according to the procedure described in Example 3 to obtain 1-(3-acetylthio-2-methylpropanoyl)-L-proline methyl ester.
Example 36 1-(3-Mercapto-2-methylpropanoyl)-L-Proline amide By substituting 1-(3-acetylthio-2-methylpropanoyl)-L-proline methyl ester in the procedure of Example 4, 1-(3-mercapto-2-methylpropanoyl)-L-proline amide is obtained.
Example 37 3-Acetylthio-2-Benzylpropanoic Acid By substituting 2-benzylacrylic acid for the metha-crylic acid in the procedure of Example 25, 3-acetylthio-2-benzylpropanoic acid is obtained.
Example 38 1-(3-Acetylthio-2-benzylpropanoyl)-L-Proline tert-butyl Ester By substituting 3-acetylthio-2-benzylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the proce-dure of ,j . :~
l3~l35b E,~:3mple 2~ (3-acetylthio-2-benzylpropanoyl)-L-proline tert-butyl ester is obtained.
_xamp e 39 anoyl?-L-Proline The product of Example 38 is substituted for the 1-(3-acetylthio-2-methylpropanoyl-L-proline tert-butyl ester in the Procedure A of Example 29 to obtain 1-(3-acetylthio-2-benzylpropanoyl)-L-proline.
Example 40_ 1-(3-Mercapto-2-benzylpropanoyl)-L-Proline 1-(3-Acetylthio-2-benzylpropanoyl)-L-pxoline is treated with methanolic ammonia according to the procedure of Example 34 to obtain 1-(3-mercapto-2-benzylpropanoyl)-L-proline as an oil, Rf = 0.47 (silica gel, benzene-acetic acid (75:25).
Example 41 1-(3-Mercapto-2-methylpropanoyl)-L-Hydroxy Proline By substituting L-hydroxy proline tert-butyl ester in the procedure of Example 28, treating the product according to Procedure A of Example 29 and then continuing as in Example 34, 1-(3-acetylthio-2-methylpropanoyl)-L-hydroxyproline tert-butyl ester, l-(3-acetylthio-2-methylpropanoyl)-L-hydroxyproline and l-(3-mercapto-2-methylpropanoyl)-L-hydroxyproline, respectively, are obtained.
Example 42 1-(3-Mercapto-2-methylpropanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid tert-butyl ester in the procedure of Example 28, treating the product according to the Procedure A of Example 29 and then continuing as ;'5.~
in Example 34, 1-(3-acetylthio-2-methylpropanoyl)-L-azetidine-2-carboxylic acid tert-butyl ester, 1,3-acetylthio-2-methyl-propanoyl)-L-azetidine-2-carboxylic acid and 1-(3-mercapto-2-methylpropanoyl-L-azetidine-2-carboxylic acid are obtained.
Example 43 1-(3-Mercapto-2-methylpropanoly)-L-Pipecollc Acid By substituting L-pipecolic acid in the procedure of Example 28, treating the product according to Procedure A of Example 29 and then continuing as in Example 34, 1-(3-acetyl-thio-2-methylpropanoyl)-L-pipecolic acid tert-butyl ester, 1-(3-acetylthio-2-methylpropanoyl)-L-pipecolic acid and 1-(3-mercapto-2-methylpropanoyl)-L-pipecolic acid, respectively, are obtained.
Example 44 1_(4-senzoylthiobutanoyl)-L-Proline To a solution of L-proline (2.88 g.) in normal sodium hydroxide (25 ml.) chilled in an ice bath, 2N sodium hydroxide (12.5 ml.) and 4-chlorobutyryl chloride (3.5 g.) are added.
The reaction mixture is stirred at room temperature for 3.5 hours and a suspension of thiobenzoic acid (3.75 g.) and potassium carbonate (2.4 g.) in water (25 ml.) is added. After overnight stirring at room temperature, the reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried over magnesium sul-fate and concentrated to dryness in vacuo. The residue is chromatographed on a column of silica gel with benzene-acetic acid (7:1). The fractions containing the desired material are pooled and concentrated to dryness, yield 1.35 g. A small aliquot of this material is dissolved in ethyl acetate and dicyclohexylamine is added until pH 8-10 (on a wet pH paper).
~i .
The dicyclohe~ylamine salt crystalli~es out, immediately, m.p.
159-161~.
E~ample 45 1-_(4-~ercaptobu-~anoyl)-L-Proline 1-(4-Benzylthiobutanoyl)-L-proline (1.08 g.) is dissolved in a mix-ture of water (4 ml.) and concentrated ammonia (2.7 ml.). After one hour stirring at room tempera-ture, the mixture is diluted with water, filtered, extracted with ethyl acetate, and the aqueous phase was concentrated in vacuo.
This ammonium salt of 1-(4-mercaptobu-tanoyl)-L-proline is puri ied by ion exchange chromatography on a column of diethyl-aminoethyl-Sephadex (cross linked dextran) with a gradient of ammonium bicarbonate, yield 0.7 g. The ammonium salt is dissolved in water ~2 ml.) and applied to a column of Dowex 50 sul~onic acid resin analytical grade in the hydrogen form, and the free acid is eluted with water. The fractions containing the desired material (sulfhydryl reagent and carboxyl reagent positive) are pooled and freeze dried to obtain l-(4-mercaptobutanoyl)-L-proline. The dicyclohexyl ammonium salt is produced by the procedure of Example 44, m.p. 157-158.
Example 46
4-Bromo-2-Methylbutanoic Acid Ethyl-4-bromo-2-methylbutanoate [G. Jones and J. Wood, Tetrahedron, 21, 2961 (1965)] (1.04 g.) is dissolved in dichloromethane (50 ml.) and cooled to -10. A 1 M solution of boron tribromide in dichloromethane (50 ml.) is added dropwise with stirring and the stirring is continued for 1 hour at -10 and 2 hours at 25. The reaction is terminated b~ the cclreful addition of water. The layers are separated and the oryanic phase is washed with water, dried and concen-trated to dryness to obtain 4-bromo-2-methylbutanoic acid.
Example 47 _-(4-Benzoylthio-2-methylbutanoyl?-L-Proline a) 4-sromo-2-methylbutanoic acid (8 g.) and thionyl chloride (7 g.) are mixed and the mixture is stirred for 16 hours at room temperature. The reaction mixture is concentrated to dryness and distilled in vacuo.
b) To a solution of L-proline (2.88 g.) in normal sodium hydroxide (25 ml.) chilled in an ice bath, 2N sodium hydroxide (12.5 ml.) and the 4-bromo-2-methylbutanoic acid chloride obtained in part (a) (3.9 g.) are added. The reaction mixture is stirred at room temperature for 3.5 hours and a suspension of thiobenzoic acid (3.75 g.) and potassium carbonate (2.4 g.) in water (25 ml.) is added. After overnight stirring at room temperature, the reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo. The residue is chromatographed on a column of silica gel with benzene-acetic acid (7:1). The fractions containing the desired product, 1-(4-ben~oylthio-2-methylbutanoyl)-L-proline are pooled and concentrated to dryness in vacuo.
Example 48 l-(4-Mercapto-2-methylbutanoyl)-L-Pro]ine By substituting 1-(4-benzoylthio-2-methylbutanoyl)-L-proline for the 1-(4-benzoylthiobutanoyl)-L-proline in the procedure of Example 45, l-(mercapto-2-methylbutanoyl)-L-proline in obtained.
~,i
Example 47 _-(4-Benzoylthio-2-methylbutanoyl?-L-Proline a) 4-sromo-2-methylbutanoic acid (8 g.) and thionyl chloride (7 g.) are mixed and the mixture is stirred for 16 hours at room temperature. The reaction mixture is concentrated to dryness and distilled in vacuo.
b) To a solution of L-proline (2.88 g.) in normal sodium hydroxide (25 ml.) chilled in an ice bath, 2N sodium hydroxide (12.5 ml.) and the 4-bromo-2-methylbutanoic acid chloride obtained in part (a) (3.9 g.) are added. The reaction mixture is stirred at room temperature for 3.5 hours and a suspension of thiobenzoic acid (3.75 g.) and potassium carbonate (2.4 g.) in water (25 ml.) is added. After overnight stirring at room temperature, the reaction mixture is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is dried over magnesium sulfate and concentrated to dryness in vacuo. The residue is chromatographed on a column of silica gel with benzene-acetic acid (7:1). The fractions containing the desired product, 1-(4-ben~oylthio-2-methylbutanoyl)-L-proline are pooled and concentrated to dryness in vacuo.
Example 48 l-(4-Mercapto-2-methylbutanoyl)-L-Pro]ine By substituting 1-(4-benzoylthio-2-methylbutanoyl)-L-proline for the 1-(4-benzoylthiobutanoyl)-L-proline in the procedure of Example 45, l-(mercapto-2-methylbutanoyl)-L-proline in obtained.
~,i
5~
xample 49 4-Bromo-2-benzylbutanoic acid By subs-tituting ethyl-4-bromo-2-benzylbu-tanoate [prepared by the procedure of G. Jones and J. Wood [Te-trahedron, 21, 2961 (1965) starting with diethylbenzylmalonate]] for the ethyl-4-bromo-2-methylbutanoate in the procedure of Example 46, 4-bromo-2-benzylbu~anoic acid is obtained.
Example 50 1-(4-Benzoylthio-2-benzylbutanoyl)--L-Proline By substituting 4-bromo-2-benzylbutanoic acid for the 4-bromo-2-methylbutanoic acid in the procedure o~ -Example 47, 1-(4-benzoylthio-2-benzylbutanoyl)-L-proline is obtained.
Example 51 1-(4-Mercapto-2-benzylbutanoyl)-L-Proline By substituting 1-(4-benzoylthio-2-benzylbutanoyl)-L-pr~line for the l-(4-benzoylthiobutanoyl)-L-proline in the procedure of Example 45, 1-(mercapto-2-benzylbutanoyl)-L-proline is obtained.
Example 52 1-~4-Mercaptobutanoyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example ~ and subjecting the product to ammonolysis as in Example 45, 1-(4-benzoylthiobutanoyl)-L-hydroxy-proline and l-(4-mercaptobutanoyl)-L-hydroxyproline, respectively, are obtained.
Example 53 1- (4-Mercaptobutanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid for the L-proline in the procedure of Example 44 and subjecting the ~ 2 3~
product to ammonolysis as in Example 45, 1-(4-benzoylthio-butanoyl)-L-azetidine-2-carboxylic acid and 1-(4-mercapto-butanoyl)-L-azetidine-2-carboxylic acid, respectivel~, are obtained.
Example 5~
- ( d -Mercaptobutanoyl)-L-Pipecolic i~cid _ . . .. . _ ..
By substituting L-pipecolic acid for the L-proline in the procedure of Example 44 and subjecting the product to ammonolysis as in Example 45, 1-(4-benzoylthiobutanoyl)-L-pipecolic acid and 1-(4-mercaptobutanoyl)-L-pipecolic acid, respectively, are obtained.
Example 55 1-(3-Acetylthiobutanoyl)-L-Proline tert-butyl Ester Dicyclohexylcarbodiimide (6.2 g.) and 3--acetylthio-butyric acid (4.86 g.) are added to a solution of L-proline tert-butyl ester (5.1 g.) in dichloromethane (60 ml.) stirred in an ice bath. After 15 minutes the ice bath is removed and the mixture is stirred at room temperature for 16 hours.
The precipitate is filtered, the filtrate is concentrated to dryness and the residue is chromatographed on a column of silica gel with chloroform to obtain l-(3-acetylthiobutanoyl)-L-proline tert-butyl ester, yield 5.2 g.
Example 56 1-(3-Acetylthiobutanoyl)-L-Proline The 1-(3-acetylthiobutanoyl)-L-proline tert-butyl ester of Example 55 (5.2 g.) is dissolved in a mixture of trifluoro-acetic acid 160 ml.) and anisole (30 ml.) and the solution is kept at room temperature for one hour. The solvents are removed in vacuo and the residual l-(3-acetylthiobutanoyl)-L-proline is reprecipitated from ether-hexane several times, yield ~ 3..'a7 4 g.. The dicyclohexylamine salt is made by the procedure or Example 44, m.p. 175-176.
Example 57 ._ -(3-~lercaptobutanoyl)-L-Proline The 1-(3-acetylthio~utanoyl~-L-proline of Example 56 (0.86 g.) is dissolved in 5.5 N. methanolic a~nonia (20 ml.) and the reaction rnixture is stored at room temperature for 2 hours. The solvent is removed in_vacuo and the residue chromatographed on an ion exchange column (Dowex 50) with water. The fractions containing the desired 1-(3-mercaptobutanoyl)-L-proline are pooled and lyophilized, yield 0.6 g. The dicyclohexylamine salt is produced by the procedure of Example 44, m.p. 183-184.
Example 58 l-E3-[[(Ethoxy)carbonyl]thio]propanoyl]-L-proline Ethyl chloro~ormate (1.2 g.) is added to a solution of 3-mercaptopropanoyl-L-proline (2.03 g.) in normal sodium bicar-bonate (30 ml.) and the mixture is stirred vigorously at 5 for one hour, and for two hours at room temperature. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic phase is washed with water, dried over magnesium sulfate, and concentra-ted to dryness to yield l-[3-[[(ethoxy)carbonyl]thio]propanoyl]-L-proline.
.5~
Example 59 1-[3-[[(Ethoxy)thiocarbollyl]thio]propanoyl]-L- roline Aqueous 2N sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added -to a solution of L-proline (5.75 g) in N sodium hydroxide (50 ml) chilled and stirred in an ice bath. After five minutes the ice bath is removed and the stirring is continued at room tempe;ra-ture. After three hours ethyl xantogenic acid potassium salt (9.6 g) is added and the mixture is stirred overnight at room temperature. The solution is acidified with concentrated hydrochloric acid and extracted with ethyl aceta-te. The organic layer is concentrated to dry-ness and the residue is chromatographed on a column of silica gel with a mixture of benzene-acetic acid (7:1) as solvent, to yield l-[3-[[tethoxy)thiocarbonyl]thio]propanoyl]-L-proline, m.p. 94-95.
Example 60 1-~3-1~(Benzylthio)carbonyl]thio]propanoyl]-L-proline A solution of benzylthiocarbonyl chloride (11 ml) in dio~ane (20 ml) is added in five portions to a solution of 1-(3-mercaptopropanoyl)-L-proline (1.6 g) in normal sodium bicarbonate (24 ml) chilled in an ice bath, over a period of 30 minutes. The ice bath is removed and the stirring is continued for 2.5 hours at room temperature. After acid-ification with concentrated hydrochloric acid, the aqueous phase is extracted with ethyl acetate. The organic phase is dried over magnesium sulfate and concentrated to dryness to yield l-[3-[[(benzylthio)carbonyl]thio]propanoyl]-L-proline.
--~1--i7 E~ample 61 1-[3-[[~Ethylthio)thiocarbonyl]thio]propanoyl]-L-proline Aqueous 2N sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N sodium hydroxide (50 ml) chilled and stirred in an ice bath. After five minu-tes, the ice bath is removed and the stirring is continued at room temperature. After three hours, etnyl trithiocarbonate potassium salt (10.5 g) is added and the mixture is stirred at room -temperature overnight. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic layer is dried over magnesium sulfate and concentrated to dryness to yield 1-[3-[[(ethylthio]t~iocarbonyl]thio]propanoyl]-L-proline.
Example 62 3-~[(~lethylamino)thiocarbonyl]thio]propionic acid Methylisothiocyanate (4 g) is added to a solution of 3-mercaptopropionic acid (5.3 g) in a mixture of pyridine (250 ml) and 0.5 N sodium hydroxide (100 ml). The solution is kept at 40 for two hours and concentrated to dryness in vacuo. The residue is dissolved in water (100 ml.), acidified with concentrated hydrochloric acid and extracted with ether. The organic phase is concentrated to dryness to yield 3-[[(methylamino)thiocarbonyl]thio]propionic acid, m.p. 86-87.
~ J~7 E~ample 63 1-[3-[[(Methylamino)thiocarbonyl]thio]propanoyl]-L-proline tert-butyl ester To a solution of L-proline t:ert-butyl ester (1.71 g) and hydroxybenzotriazole (1.35 g) in dichloromethane (10 ml) chilled and stirred in an ice bath, dicyclohexylcarbodiimide (2.06 g) and 3-methylaminothiocarbonyl-thiopropionic acid (1.79 g) are added. After 15 minutes, -the bath is removed and the stirring is continued overnight. The precipitate is filtered off and the filtrate is diluted with ethyl acetate and washed neutral.
The organic phase is concentrated to dryness to yield 1-~3-[[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline tert-butyl ester, m.p. 129-130.
Example 64 1-~3-[~(Methylamino)thiocarbonyl]thio]propanoyl]-L-proline A) l-(Methylaminothiocarbonylthiopropanoyl)-L-proline tert-butyl ester (0.98 g) is dissolved in a mixture of anisole (3.6 ml) and trifluoroacetic acia (7.5 ml). After one hour at room temperature the mixture is concentrated to dryness in vacuo and the residue precipitated from ether-hexane three times. This material is chromatographed on a column of silica gel with a solvent mixture of benzene-acetic acid (75:25) to yield 1-[3-~[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline, Rf = 0.4 [silica gel-benzene:acetic acid (75:25)]. The dicyclohexylammonium salt has m.p. 127-129.
~ '`
B) l~ethylisothiocyanate (9 g) is added to a solution of 3-mercaptopropa~oyl-L-proline (10.1 g) in a mixture oE pyridine (250 ml) and 0.5 N sodium hydroxide (100 ml). The solu-tion is ~ept at 40 for two hours and concentrated to dryness in vacuo.
The residue is dissolved in water (100 ml), acidified with con-centrated hydrochloric acid and extracted with ethyl acetate.
The organic phase is concentrated to dryness to yield 1-[3-[[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline.
Example 65 1-[3-[[(Ethylamino)carbonyl]thio]propanoyl]-L-proline Ethylisocyanate (0.45 ml) is added to a solution of 1-(3-mercaptopropanoyl)-L-proline (1 g) in a mixture of N
sodium hydroxide (5 ml) and pyridine (5 ml). The solution is heated at 40 for four hours and concentrated ln vacuo.
The residue is distributed between 0.1 N hydrochloric acid and ethyl acetate. The organic layer is washed with water, dried over magnesium sulfate and concentrated to dryness to yield l-[3-[~(ethylamino)carbonyl]thio]propanoyl]-L-proline.
The dicyclohexylammonium salt is prepared by adding dicyclo-hexylamine to a solution of the free acid in ethyl acetate,m.p. 150-152.
Example 66 1-[3-[1~Ethoxy)carbonyl]thio]-2-methylpropanoyl]-L-proline By substituting 1-(3-mercapto-2-methylpropanoyl]-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[ r (ethoxy)carbonyl]thio]-2-methylpropanoyl]-L-proline is obtained.
~ ,i ~3~7 Example 67 1-[3-~[(Ethoxy)carbonyl]thio]butanoyl]-L-pro_ine By substituting 1-(3-mercaptobutanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[[(e-thoxy)carbonyl]thio]butanoyl]-L-pro-line is obtained.
Example 6~3 1-13-[[(Ethoxy)thiocarbonyl]thio]propanoyl]-L-azetidine-2-carboxylic acid .
By substituting L-azetidine-2-carboxylic acid for L-proline in the procedure oE Example 59, 1-[3-[[(ethoxy)-thiocarbonyl]thio]propanoyl]-L-azetidine-2-carboxylic acid is obtained.
Example 69 1-[3-[[(Ethoxy)thiocarbonyl]thio]propanoyl]-L-pipecolic acid By substituting L-pipecolic acid for L-proline in the procedure of Example 59, 1-[3-[[(ethoxy)thiocarbonyl]-thio]propanoyl]-L-pipecolic acid is obtained.
Example 70 1-[4-[[(Benzylthio)carbonyl]thio]butanoyl]-L-proline By substituting 4-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 60, 1-[4-[[(benzylthio)carbonyl]thio]butanoyl]-L-proline is obtained.
Example 71 1-[2-[[(Benzylthio)carbonyl]thio]propanoyl]-L-proline By substituting 2-mercaptopropanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 60, 1-[2-[[(benzylthio)carbonyl]-thio]propanoyl]-L-proline is obta:ined.
' Example 72 1-[3-[[(Ethylthio)thiocarbonyl]thio]propanoyl]-L-proline methyl ester A solution of 1-[3-[[(ethylthio)thiocarbonyl]-thio]-propanoyl]-L-proline in ethyl acetate is trea-ted wi-th an ethereal solution of diazomethane until persis-tent yellow color. AEter discharging the yellow color with a few drops of acetic acid, the solvents are removed in vacuo to yield l-[3-[[(ethylthio)-thiocarbonyl]thio]propanoyl]-L-proline methyl ester.
Example 73 1-[3-[[(methylamino)thiocarbonyl]thio]propanoyl]-5-hydroxy-L-~i~ecolic acid .
sy substituting 1-(3-mercaptopropanoyl)-5-hydroxy-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the Procedure B of Example 64, 1-[3-[[(methylamino)thiocarbonyl]thio]-propanoyl]-5-hydroxy-L-pipecolic acid is obtained.
Example 74 1-[3-[[(Methylamino)thiocarbonyl]thio]-2-methylpropanoyl]-L-proline amide By substi-tuting 1-(3-mercapto-2-methylpropanoyl)-L-proline amide for the 3-mercaptopropanoyl-L-proline in the Procedure B of Example 64, 1-[3-[[(methylaminojthiocarbonyl]thio]-2-methyl-propanoyl]-L-proline amide is obtained.
Example 75 1-[3-[[(Phenoxy)carbonyl]thio]propanoyl]-L-proline By substituting phenylchloroformate for ethyl chloroformate in the procedure of Example 58, 1-[3-[[(phenoxy)carbonyl]thio]-propanoyl]-L-proline is obtained.
5~
Example 76 l-[3-[[(Phenoxy)carbonyl]thio]butanoyl]-L-proline By substituting phenylchloroformate for the ethyl chloroformate and 4-mercaptobu-tanoy:l-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[[(phenoxy)carbonyl]thio]butanoyl]-L-proline is obtained.
Example 77 l-[3-[[(Phenylamino)carbonyl]thio]propanoyl]-L-proline By substituting phenylisocyanate for the ethylisocyanate in the procedure of Example 65, 1-[3-[[(phenylamino)carbonyl]-thio]propanoyl]-L-proline is obtained.
Example 78 l-~3-[[(Phenethylamino)carbonyl]thio]propanoyl]-L-proline By substituting phenethylisocyanate for the ethylisocyanate in the procedure of Example 65, 1-[3-[[(phenethylamino)carbonyl]-thio]propanoyl]-L-proline is obtained.
Example 79 l-~3-[~(Ethylamino)carbonyl]thio]-2-benzylpropanoyl]-L-proline By substituting l-(3~mercapto-2-benzylpropanoyl)-L-proline for the l-(3-mercaptopropanoyl)-L-proline in the procedure of Example 65, 1-[3-[[(ethylamino)carbonyl]thio]-2-benzylpropanoyl]-L-proline is obtained.
Example 80 1-(3-Methylthiopropanoyl)-L-proline A) l~lethyl 3-methylthiopropionate (51 g) is saponified with a 10~ sodium hydro~ide solution (150 ml, 30 minutes at lOOD).
The cooled solution is extracted with ether and then acidified.
The crude acid thus obtained is distilled and converted to the acid chloride with thionyl chloride.
A solution of L-proline (11.5 g) in N sodium hydroxide (100 cc) is chilled in an ice bath and the 3-methylthiopropanoic acid chloride (6.9 g) is added dropwise with vigorous stirring over a ten minutes period. After five hours the reaction mixture is acidified and extracted with ethyl ether to yield 1-(3-methylthiopropanoyl)-L-proline. The dicyclohexylammonium salt is prepared by adding dicyclohexylamine to a solution of the free acid in ethyl acetate, m.p. 169-171.
B) ~lethyl iodide ~71 g) is added to a solution of 1-(3-mercaptopropanoyl)-L-proline ethyl ester (115 g) and sodium (11.5 g) in ethanol (400 ml). The reaction is allowed to proceed overnight, the ethanol is removed in vacuo and the residue is dissolved in a mixture of ethyl acetate and water.
Tne organic layer is dried and concentrated to dryness in vacuo.
The resulting 1-(3-methylthiopropanoyl)-L-proline ethyl ester (98 g) is suspended in a mixture of methanol (200 ml) and 5 N
sodium hydroxide (200 ml) and stirred at room temperature for five hours. The methanol is re~oved in vacuo, and the aqueous phase is extracted with ethyl acetate, acidified and reextracted with ethyl acetate. This last organic phase is washed with water, dried and concentrated to dryness to yield l-(3-methylthio-propanoyl)-L-proline.
~ -48-Example 81 __ 1-[3-(4-Chlorophenvlthio)propanoyl]-L-proline Aqueous 2 N sodium hydro~ide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N-sodium hydro~ide (50 ml) chilled and stirred in an ice bath. After five minu-tes, the ice bath is removed and the stirring is continued for three hours at room temperature. The reaction mixture is acidified with concentrated hydrochloric acid ~nd extracted with ethyl acetate. The organic layer is washed with water, dried and concentrated to dryness in vacuo. The residue is dissolved in a mixture of 4-chlorobenzenethiol (8 g), sodium hydroxide (4.2 g) and ethanol (300 ml). The solution is refluxed for 6 hours. The solvent is removed ln vacuo and the residue is dissolved in water, acidified with concentrated hydro-chloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried, and concentrated to dryness in vacuo to yield l-[3-(4-chlorophenylthio)propanoyl]-L-proline.
Example 82 1,[~(3-Benzylthiomethyl)thio]propanoyl]-L-proline 1-(3-Mercaptopropanoyl)-L-proline (8.1 g) is dissolved ,!
in boiling liquid ammonia (100 ml) and small pieces of sodium are added until permanent blue color is obtained which is then discharged with a small piece of ammonium chloride. Benzyl-thiomethyl chloride (6.9 g) is added and the ammonia is allowed to evaporate. The final traces of ammonia are removed in vacuo, the residue is dissolved in water and extracted with ethyl acetate. The aqueous phase is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried and concentrated to dryness to yield l-~[(3-benzylthiomethyl)-thio]propanoyl]-L,-proline.
Example 83 1-[[(3-Acetamidome-thyl)thio]pro _ oyl]-L-proline 1-(3-Mercaptopropanoyl)-L-proline (2 g) and N-hydroxy-methylacetamide (0.8~ g) are dissolved in trifluoroacetic acid (10 ml) and the solution is stored at room temperature for one hour. The excess tirfluoroacetic acid is removed in vacuo and the residue is precipitated several times from ether-hexane.
~inally, the residue is distributed between dilute hydrochloric acid and ethyl acetate. The or~anic layer is washed with water, dried and concentrated to dryness to yield l-[[3-acetamido-methyl)thio]propanoyl]-L-proline.
Example 84 l-(~ethylthioacetyl)-L-proline By substitutin~ methyl methylthioacetate for the methyl 3-methylthiopropionate in the Procedure A of Example 80, 1-(methylthioacetyl)-L-proline, m.p. 123-124, is obtained.
Example 85 l-(Benzylthioacetyl)-L-proline .
By substitutin~ benzylthioacetyl chloride for the,3-methylthiopropanoyl chloride in the Procedure A of Example 80, l-(benzylthioacetyl)-L-proline, m.p. 86-88, is obtained.
Example 86 1-[3-[(2-Phenylethyl)thio]propanoy.l]-L.-prol.ine By substituting phenethylbrom:ide for the methyl io-dide in the Procedure B of Example 80, 1-[3-[(2-phenyl-ethyl)thio]propanoyl]-L-proline is obtained.
Example 87 1-13-[(Triphenylmethyl)thio]propanoyl]-L-proline By substituting triphenylmethyl chloride for the methyl iodide in the Procedure B of Example 80, 1-~3-~(triphenylmethyl)thio]propanoyl]-L-proline is obtained.
Example 88 1-(3-Methylthio-2-methylpropanoyl)-L-proline amide By substituting 1-(3-mercapto-2-methylpropanoyl)-L-proline amide for the l-(3-mercaptopropanoyl)-L-proline ethyl ester in the Procedure B of Example 80 and elimina-ting the saponification step, l-(3-methy].thio-2-methylpro-panoyl)-L-proline amide is obtained.
Example 89 1-(3-Methylthiopropanoyl)-L-azetidine-2-carboxylic acid By substituting L-azetid.ine-2-carboxylic acid for the L-proline in the Procedure A of Example 80, 1-(3-methylthiopropanoyl)-L-azetidine-2-carboxylic acid is obtained.
;~ -~ D~br~i 3~
Example 90 1-['-(4-Methoxyphenylthio)propanoyl]-L-proline By substituting 4-methoxybenzenethiol Eor the 4-chloro-benzenethiol in the procedure of Example 81, 1-[3-(4-methoxy-phenylthio)propanoyl]-L-proline is obtained.
Example 91 1-(3-Methylthiopropanoyl)-L-pipecolic acid By substituting L-pipecolic acid for the L-proline in the Procedure A of Example 80, 1-(3-methylthiopropanoyl)-L-pipecolic acid is obtained.
Example 92l-E2-(4-Chlorophenylthio propanoyl]-L-proline sy substituting 2-bromopropionyl chloride for the -3-bromopropionyl chloride in the procedure of Example 81, 1-[2-(4-chlorophenylthio-propanoyl]-L-proline is obtained. ~`
Example 93 1-[3-~(Diphenylmethyl)thio]-2-benzylpropanoyl)]-L-proline Diphenylmethanol (0.92 g) and 1-(3-mercapto-2-benzyl-propanoyl)-L-proline (1.5 g) are dissolved in trifluoroacetic acid (10 ml) and the solution-is kept at room temperature for 30 minutes. The excess trifluoroacetic acid is removed in _acuo to yield 1-[3-[(diphenylmethyl)thio]-2-benzylpropanoyl]-L-proline.
.5"~ ..
- ,.. `=~
E~mple 94 1-[4~(4-Chlorophenylthio)butanoyl]-L-proline By substituting 4-bromoprop:ionyl chloride for the 3-bromopropionyl chloride in the procedure of Example 81, 1-[4-(4-chlorophenylthio)butanoyl]-L-proline is obtained.
Example 95 _ l-[3-[(Ben2ylthiomethyl)thio]butanoyl]-L-proline By substituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 82, 1-[3-[(benzylthiomethyl)thio]butanoyl]-L-proline is obtained.
Example 96 1-[[4-[(Acetamidomethyl)thio]-2-methylbutanoyl]-L-proline By substituting 1-(4-mercapto-2-methylbutanoyl)-L-proline for the 1-~3-mercaptopropanoyl)-L-proline in the procedure of Example 83, 1-[[4-(acetamidomethyl)thio]-2-methylbutanoyl]-L-proline is obtained.
Example 97 1-~3-(Ethyldithio)propanoyl]-L-proline A) 3-Mercaptopropanoyl-L-proline (10 g) is added to a solution of ethylthiosulfinate (8.4 g) in methanol (100 ml) and the reaction mixture is stirred vigorously at room temperature for four hours. The methanol is removed in vacuo to yield l-(3-ethyldithiopropanoyl)-L-proline.
B) A solution of ethylthiosulfinate (8.4 g) in ethanol (50 ml) is added to an aqueous solution of 3-mercapto-propanoyl-L-proline (10 g) maintained at pH 6-7 by careful addition of sodium hydroxide. The mixture is stirred vigorously ~ 53-at room temperature until negative thiol reaction. The mix-ture is diluted with water, adjusted to pH 8 and extrac-ted with ethyl acetate, the aqueous phase is acidified to pH 3 and extracted again wi-th ethyl acetate. This latter extract is washed with water, dried and concentrated to dryness to yield l-[3-(ethyldithio)propanoyl]-L-proline.
Example 98 .
1-[3-[(4-Methylphenyl)dithio]propanoyl]-L-proline A solution of 4-methylphenylsulfenyl chloride (1.76 g.) in ether (20 ml) is added to a solution of 3-mercaptopropanoyl-L-proline (2 g) ir 0.5 N sodium hydroxide (20 ml) chilled in an ice bath. The mixture is stirred vigorously for one hour, and the a~ueous phase is separated, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with water, dried and concentrated to dryness to yield 1-[[3-(4-methylphenyl)dithio]propanoyl]-L-proline.
Example 99 1-[3-(Phenyldithio-propanoyl]-L-proline By substituting phenylthiosulfinate [prepared from phenyl-disulfide according to U. Weber and P. Hartter, Z. Physiol. Chem., 351, 1384 (1970)] for the ethylthiosulfinate in the procedure of Example 97, 1-[3-(phenyldithio-propanoyl]-L-proline is obtained.
Example 100 1-[3-[(2-Phenylethyl)dithio]propanoyl]-L-proline By substituting 2-phenylethylthiosulfinate (prepared from phenethyldisulfide) for the ethylthiosuflinate in the procedure of Example 97, 1-[3-[(2-phenylethyl)dithio]propanoyl]-L-proline is obtained.
.~
E ample 101 1-[3-[(2-Hydroxyethyl)dithio]propanoyl]-L-proline To a solution oE 1,1'-[(sul~inylthio)-bis-(3-propanoyl~]-bis-L-proline (21 g) in methanol (100 ml), mercap-toethanol(4.2 g) is added and -the reaction mixture is stirred vigorously at room temperature for four hours. The methanol is removed in vacuo and the residue is pruified by chromatography on a silica gel column to yield l-[3-[(2-hydroxyethyl)dithio]propanoyl]-L-proline.
Example 102 _ .
1-[2-(Ethyldithio)propanoyl]-L-proline By substituting 2-mercaptopropanoyl-L-proline for 3-mercaptopropanoyl-L-proline in the procedure of Example 97, 1-[2-(ethyldithio)propanoyl]-L-proline is obtained.
Example 103 1-[3-~(4-Methylphenyl)dithio]butanoyl]-L-proline By sùbstituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 98, 1-[3-~4-methylphenyl)dithio]butanoyl]-L-proline is obtained.
Example 104 1-13-(Ethyldithio)-2-methylpropanoyl]-L-proline methyl ester By substituting 1-(3-mercapto-2-methylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 97 and then treat:ing the product with ethereal diazomethane as in the procedure of Example 72, 1-[3-(ethyldithio)-2-methylpro-panoyl~-L-proline methyl ester is obtained.
5~
Example 105 1-[3-(Ethyldithio)propanoyl]-L-azetidine-2-carboxylic acid By subs-tituting 3-mercaptopropanoyl-L-azetidine-2-carboxylic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 97, 1-[3-(ethyldi-thio)propanoyl]-L-aze-tidine-2-carboxylic acid is obtained.
Example 106 1-[3-[(4-Methylphenyl)dithio]-2-methylpropanoyl]-L-hydroxyproline By substituting 1-(3-mercapto-2-methylpropanoyl)-L-hydroxy proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 98, 1-[(3-[4-methylphenyl)dithio]-2-methylpropanoyl]-L-hydroxyproline is obtained.
Example 107 1-[4-~Ethyldithio)butanoyl]-L-pipecolic aeid By substituting 4-mercaptobutanoyl-L-pipecolie aeid for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 97, 1-[4-(ethyldithio-butanoyl]-L-pipeeolie aeid is obtained.
Example 108 1-13-(Ethyldithio)propanoyl]-5-hydroxy-L-pipeeolie aeid By substituting 1-(3-mereaptopropanoyl)-5-hydroxy-L-pipeeolie aeid for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 97, 1-[3-(ethyldithio)propanoyl]-5-hydroxy-L-pipecolic aeid is obtained.
.
Example 109 1-[3-[(2-Amino-2-car~o~yethyl)dithio]propanoyl]-L-proline A 0.5 M solution of thiocyano~en in glacial ace-tic acid is prepared by sha~ing Eor -ten minutes in a sealed flask 600 mg of dry lead thiocyanate with a solution of 75 1l1 of bromine in 3 ml of acetic acid. After removal of lead bromide and excess lead thiocyanate by centrifugation, 2.5 ml of this solution is mixed with 2.5 ml of a 0.41 M solution of cysteine hydrochloride previously neutralized with dilute sodium hydroxide. This mixture is immediately added to 0.75 ml of a 1.9 M solution of 3-mercaptopropanoyl-L-proline previously neutralized with dilute sodium hydroxide. After twenty minutes the mixture is titrated to incipient brown color with alcoholic iodine, and adjusted to pH 3. The precipitate is removed by filtration and the filtrate is applied to a column of cation exchange resin (Dowex 50). The column is washed with water until no more acidic material is removed and then eluted with pyridine-acetate buffer pH 6Ø
The fractions containing the disulfide of cysteine and 3-mercaptopropanoyl-L-proline are pooled and concentrated to dryness.
Example 110 1,1'-~Dithiobis)4-propanoyl)]-bis-L-proline 3-Mercaptopropanoyl-L-proline (0.95 g) is dissolved in water (20 ml) and the pH is adjusted to 6.5 with N-sodium hydroxide. An ethanolic solution of iodine is added drop-wise while maintaining the pH at 6.5 with careful addition of N sodium hydroxide. When a permanent yellow color is obtained the addition of iodine is stopped and the color , .~
:' ~ ',. .. .
~.~Ll, 31~d57 is discharc~ed with a small amount of sodium thiosulfate.
The reaction mixture is acidified with concentra-t~d hydro-chloric acid and extracted with ethyl aceta-te. The organie phase is washed with wa-ter, dried and concentrated to dryness to yield l,l'-~dithiobis(3-prop~noy~l)]-bis-L-proline. The di-cyclohexylammonium salt is prepared by addition o~ dicyclo-nexylamine to a solution of the free acid in acetonitrile, m.p. 179-180.
Example 111 .
1,1'-[Dithiobis(2-D-methyl-3-propanoyl)]-bis-L-proline By substituting 3-mercapto-2-D-methylpropanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-D-methyl-3-propanoyl)]-bis-L-proline is obtained, m.p. 236-237.
Example 112 1,1'-[Dithiobis(2-propanoyl)-bis-L-proline By substituting 2-mercaptopropanoyl-L-proline for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 110, 1,1'-[dithiobis)2-propanoyl)]-bis-L-proline is obtained.
Example 113 l,l'-(Dithiobisaeetyl)-bis-L-hydroxy proline By substituting 1-(2-mereaptoacetyl)-L-hydroxyprolin~
for the 3-mercaptopropionyl-L-proline in the procedure of Example 110, l,l'-(dithiobisaeetyl)-bis-L-hydroxyproline is obtained.
Example 114 1,1'-(Dithiobisaeetyl)-bis-L-azetidine-2-earboxylie aeid By substituting 1-(2-mereaptoaeetyl)-L-azetidine-2-earboxylie aeid Eor the 3-mereaptopropanoyl-L-proline in the proeedure of Example 110, l,l'-(dithiobisaeetyl)-bis-L-azetidine-2-earboxylie aeid is obtained.
.
- Example 115 1,1'-[Dithiobis(3-propanoyl)]-bis-L-pipecolic acid sy substituting 3-mercaptopropanoyl-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-L-pipecolic acid is obtained.
Example 116 1,1'-[Dithiobis(3-propanoyl)]-bis-4-methyl-L-proline By substituting 1-(3-mercaptopropanoyl)-4-methyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-4-methyl-L-proline is obtained.
Example 117 1,1'-[Dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid By substituting 1-(3-mercaptopropanoyl)-S-hydroxy-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the pro-cedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid is obtained.
Example 118 1,1'-[Dithiobis(2-benzyl-3-propanoyl)]-bis-L-proline By substituting 1-(3-mercapto-2-benzylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-benzyl-3-propanoyl)]-bis-L-proline is obtained.
Example 119 1,1'-[Dithiobis)2-methyl-3-propanoyl)]-bis-L-pipecolic acid sy substit~lting 1-(3-mercap-to-2-methylpropanoyl)-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[di-thiobis(2-methyl-2-propanoyl)]-bis-L-pipecolic acid is obtained.
Example 120 1,1'-[Dithiobis)4-butanoyl)]-bis-L-proline By substituting 4-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(4-butanoyl)]-bis-L-proline is obtained.
Example 121 1,1'-[Dithiobis(2-benzyl-4-butanoyl)]-bis-L-proline By substituting 1-(4-mercapto-2-benzylbutanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-benzyl-4-butanoyl)]-bis-L-proline is obtained.
Example 122 1,1'-[Dithiobis(3-butanoyl)]-bis-L-proline By substituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-butanoyl)]-bis-L-proline is obtained.
Exam le 123 1,1'-[Dithiobis(3-propanoyl)]-bis-L-proline methyl es e A solution of 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in methanol is treated with ethereal diazomethane until persistent yellow color. After fifteen minutes a few drops of acetic acid are added and -the solvents are removed in vacuo to yield, l,l'-[dithiobis(3-propanoyl)]-bis-L-proline methyl ester.
Example 124 1,1'-[Dithiobis(3-propanoyl)]-bis-L-proline amide A solution of 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline methyl ester in methanol is saturated with am-monia while cooling in an ice-water bath. The reaction mixture is stored for 16 hours at room temperature in a pressure bottle, and then the solvents are removed in vacuo to yield l,l'-[dithiobis(3-propanoyl)]-bis-L-proline amide.
Example 125 1,1'-[Dithiobis(2-phenyl-3-propanoyl)]-bis-L-proline By substituting 1-(3-mercapto-2-phenylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the proceaure of Example 110, 1,1'-[dithiobis(2-phenyl-3-pro-panoyl)]-bis-L-proline is obtained.
2~;~
Example 126 1,1'-[(Sulfin_lthio)-bis-~3-propanoyl)]-bis-L-proline While cooling in an ice bath 0.12 mole of peracetic acid is added to a stirred solution of l,l'-[dithiobis(3-propanoyl)]-bis-L-proline (40 g) in glacial acetic acid (500 ml). The reac-tion mixture is allowed to stand over-night at room temperature and -the solvent is then removed _ vacuo to yield 1,1'-[(sulfinylthio)-bis-(3-propanoyl)]-bis-L-proline.
Example 127 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-L-proline A 30~ solution of hydrogen peroxide (2.0 ml) is added to a solution of l,l'-[dithiobis(3-propanoyl)~-bis-L-proline (4 g) in glacial acetic acid (80 ml) and the solution is stored for thirty hours at room temperature.
The solvent is removed in vacuo to yield l,l'-[(sulfonyl-thio)-bis-(3-propanoyl)]-bis-L-proline.
Example 128 1,1'-[(Sulfinylthio)-bis-(2-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-propanoyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-propanoyl)]-bis-L-proline is obtained.
:
'.~
i7 Exa ple 129 1,1'-[(S lfinylt_io)-bis-acetyl]-bis-L-azetidine-2-carboxy-lic acid By substituting l,l'-(dithiobisacetyl)-bis-L-azeti-dine carboxylic acid for the l,l'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-acetyl]-bis-L-azetidine-2-carboxylic acid is obtained.
Example 130 1,1'-[(Sulfinylthio)-bis-(3-propanoyl)]-bis-4-methyl-L-proline By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-4-methyl-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(3-propanoyl)]-bis-4-methyl~L-proline is obtained.
Example 131 1,1l-[(Sulfinylthio)-bis-(2-benzyl-3-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-benzyl-3-propa-noyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-benzyl-3-propanoyl)]-bis-L-proline is obtained.
Example 132 1,1'-[(Sulfinylthio)-bis-(4-butanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(4-butanoyl)]-bis-L-proline for the 1,1'-[dithlobis(3-propanoyl)]-bis-L-proline in -the procedure of Example 126, l,l'-[(sulfinyl-thio)-bis-(4-but.anoyl)]-bis-L-proline is obtained.
~ -63-3~
Example 133 1,1'-[(Sulfinylthio)-bis-(3-butanoyl)]-bis--L-proline By substituting 1,1'-[di-thiobis(3-butanoyl)]-bis-L-proline for the l,l'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sulfinylthio)-bis-(3-butanoyl)]-bis-L-proline is obtained.
Example 134 1,1'-[(Sulfinylthio)-bis-(2-methyl-3-propanoyl)-bis-L-pro-line By substituting 1,1'-[dithiobisl2-methyl-3-propan-oyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-proline is obtained.
Example 135 1,1'-[(Sulfinylthio)-bis-(2-phenyl-3-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-phenyl-3-propa-noyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,1'-[(sul-finylthio)-bis-(2-phenyl-3-propanoyl)]-bis-L-proline is obtained.
Example 136 1-[3-[[3-(2-Carboxy-l-pyrrolidinyl)-3-oxopropyl]-dithio]-2-methylpropanoyl]-L-proline By substituting 1,1'-[(sulfinylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-proline for the ethylthiosulfinate in the procedure of Example 97, 1-[3-[[3-(2-carboxy-1-pyrro-lidinyl)-3-oxopropyl]dithio]-2-methylpropanoyl]-L-proline is obtained.
.~ ~
Example 137 l,l'-[(Sulfon~lthio)-bis-acetyl)-bis-L-hydroxyproline By substituting l,l'-(dithiobisacetyl)-bis-L-hy-droxy proline for the l,l'-[di-thiobis(3-propanoyl)-bis-L-proline in the procedure of Example 127, l,l'-[(sulfonyl-thio)-bis-acetyl)-bis-L-hydroxypro:Line is obtained.
Example 138 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-L-pipecolic acid By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-L-pipecolic acid for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 127, l,l'-[(sul-fonylthio)-bis-(3-propanoyl)]-bis-L-pipecolic acid is ob-tained.
Example 139 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid for the 1,1'-[dithiobis-(3-propanoyl)]--bis-L-proline in the procedure of Example 127, 1,1'-~(sulfonylthio)-bis-(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid is obtained.
Example 140 1,1'-[(Sulfonylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-pipecolic acid By substituting 1,1'-[dithiobis(2-methyl-3-propa-noyl)]-bis-L-pipecolic acid for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 127, 1,1'-[(sulfonylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-pipecolic acid is obtained.
~ -65-E~ le 141 1,1'-[(Sulfonylthio)-bis-(2-be~ yl-~-butanoyl)]-bls-L-proline By substituting 1,1'-[dithiobis(2-benzyl-4-butan-onyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the proc~dure of Example 127, l,l'-[(sul-fonylthio)-bis-(2-benzyl-'-butanoyl)]-bis-L-proline is ob-tained.
Exall~ple 142 3-Acetylthio-2-phenylpro~ noic acid By substituting 2-phenylacrylic acid for the meth-acrylic acid in the procedure of Example 25, 3-acetylthio-2-phenylpropanoic acid is obtained.
1-(3-Acetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester By substituting 3-acetylthio-2-phenylpropanoic acid for the 3-acetylthio-2-me~hylpropanoic acid in the proce-dure of Example 28, 1-(3-~cetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester is obtained.
Exa~nple 143 1-(3-Mercapto-2-phenylprc~anoyl)-L-proline By substituting 1-(3-acetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester for the 1-(3-acetylthio-2-methyl-propanoyl-L-prolin~ tert-butyl ester in the pro-cedure of Example 29, and subjecting the product to am-monolysis as in Example 3~, 1-(3-acetylthio-2-phenylpro-panoyl)-L-proline and l-~,-mercapto-2-phenylpropanoyl)-L-proline are obtained.
xample 49 4-Bromo-2-benzylbutanoic acid By subs-tituting ethyl-4-bromo-2-benzylbu-tanoate [prepared by the procedure of G. Jones and J. Wood [Te-trahedron, 21, 2961 (1965) starting with diethylbenzylmalonate]] for the ethyl-4-bromo-2-methylbutanoate in the procedure of Example 46, 4-bromo-2-benzylbu~anoic acid is obtained.
Example 50 1-(4-Benzoylthio-2-benzylbutanoyl)--L-Proline By substituting 4-bromo-2-benzylbutanoic acid for the 4-bromo-2-methylbutanoic acid in the procedure o~ -Example 47, 1-(4-benzoylthio-2-benzylbutanoyl)-L-proline is obtained.
Example 51 1-(4-Mercapto-2-benzylbutanoyl)-L-Proline By substituting 1-(4-benzoylthio-2-benzylbutanoyl)-L-pr~line for the l-(4-benzoylthiobutanoyl)-L-proline in the procedure of Example 45, 1-(mercapto-2-benzylbutanoyl)-L-proline is obtained.
Example 52 1-~4-Mercaptobutanoyl)-L-Hydroxyproline By substituting L-hydroxyproline for the L-proline in the procedure of Example ~ and subjecting the product to ammonolysis as in Example 45, 1-(4-benzoylthiobutanoyl)-L-hydroxy-proline and l-(4-mercaptobutanoyl)-L-hydroxyproline, respectively, are obtained.
Example 53 1- (4-Mercaptobutanoyl)-L-Azetidine-2-Carboxylic Acid By substituting L-azetidine-2-carboxylic acid for the L-proline in the procedure of Example 44 and subjecting the ~ 2 3~
product to ammonolysis as in Example 45, 1-(4-benzoylthio-butanoyl)-L-azetidine-2-carboxylic acid and 1-(4-mercapto-butanoyl)-L-azetidine-2-carboxylic acid, respectivel~, are obtained.
Example 5~
- ( d -Mercaptobutanoyl)-L-Pipecolic i~cid _ . . .. . _ ..
By substituting L-pipecolic acid for the L-proline in the procedure of Example 44 and subjecting the product to ammonolysis as in Example 45, 1-(4-benzoylthiobutanoyl)-L-pipecolic acid and 1-(4-mercaptobutanoyl)-L-pipecolic acid, respectively, are obtained.
Example 55 1-(3-Acetylthiobutanoyl)-L-Proline tert-butyl Ester Dicyclohexylcarbodiimide (6.2 g.) and 3--acetylthio-butyric acid (4.86 g.) are added to a solution of L-proline tert-butyl ester (5.1 g.) in dichloromethane (60 ml.) stirred in an ice bath. After 15 minutes the ice bath is removed and the mixture is stirred at room temperature for 16 hours.
The precipitate is filtered, the filtrate is concentrated to dryness and the residue is chromatographed on a column of silica gel with chloroform to obtain l-(3-acetylthiobutanoyl)-L-proline tert-butyl ester, yield 5.2 g.
Example 56 1-(3-Acetylthiobutanoyl)-L-Proline The 1-(3-acetylthiobutanoyl)-L-proline tert-butyl ester of Example 55 (5.2 g.) is dissolved in a mixture of trifluoro-acetic acid 160 ml.) and anisole (30 ml.) and the solution is kept at room temperature for one hour. The solvents are removed in vacuo and the residual l-(3-acetylthiobutanoyl)-L-proline is reprecipitated from ether-hexane several times, yield ~ 3..'a7 4 g.. The dicyclohexylamine salt is made by the procedure or Example 44, m.p. 175-176.
Example 57 ._ -(3-~lercaptobutanoyl)-L-Proline The 1-(3-acetylthio~utanoyl~-L-proline of Example 56 (0.86 g.) is dissolved in 5.5 N. methanolic a~nonia (20 ml.) and the reaction rnixture is stored at room temperature for 2 hours. The solvent is removed in_vacuo and the residue chromatographed on an ion exchange column (Dowex 50) with water. The fractions containing the desired 1-(3-mercaptobutanoyl)-L-proline are pooled and lyophilized, yield 0.6 g. The dicyclohexylamine salt is produced by the procedure of Example 44, m.p. 183-184.
Example 58 l-E3-[[(Ethoxy)carbonyl]thio]propanoyl]-L-proline Ethyl chloro~ormate (1.2 g.) is added to a solution of 3-mercaptopropanoyl-L-proline (2.03 g.) in normal sodium bicar-bonate (30 ml.) and the mixture is stirred vigorously at 5 for one hour, and for two hours at room temperature. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic phase is washed with water, dried over magnesium sulfate, and concentra-ted to dryness to yield l-[3-[[(ethoxy)carbonyl]thio]propanoyl]-L-proline.
.5~
Example 59 1-[3-[[(Ethoxy)thiocarbollyl]thio]propanoyl]-L- roline Aqueous 2N sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added -to a solution of L-proline (5.75 g) in N sodium hydroxide (50 ml) chilled and stirred in an ice bath. After five minutes the ice bath is removed and the stirring is continued at room tempe;ra-ture. After three hours ethyl xantogenic acid potassium salt (9.6 g) is added and the mixture is stirred overnight at room temperature. The solution is acidified with concentrated hydrochloric acid and extracted with ethyl aceta-te. The organic layer is concentrated to dry-ness and the residue is chromatographed on a column of silica gel with a mixture of benzene-acetic acid (7:1) as solvent, to yield l-[3-[[tethoxy)thiocarbonyl]thio]propanoyl]-L-proline, m.p. 94-95.
Example 60 1-~3-1~(Benzylthio)carbonyl]thio]propanoyl]-L-proline A solution of benzylthiocarbonyl chloride (11 ml) in dio~ane (20 ml) is added in five portions to a solution of 1-(3-mercaptopropanoyl)-L-proline (1.6 g) in normal sodium bicarbonate (24 ml) chilled in an ice bath, over a period of 30 minutes. The ice bath is removed and the stirring is continued for 2.5 hours at room temperature. After acid-ification with concentrated hydrochloric acid, the aqueous phase is extracted with ethyl acetate. The organic phase is dried over magnesium sulfate and concentrated to dryness to yield l-[3-[[(benzylthio)carbonyl]thio]propanoyl]-L-proline.
--~1--i7 E~ample 61 1-[3-[[~Ethylthio)thiocarbonyl]thio]propanoyl]-L-proline Aqueous 2N sodium hydroxide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N sodium hydroxide (50 ml) chilled and stirred in an ice bath. After five minu-tes, the ice bath is removed and the stirring is continued at room temperature. After three hours, etnyl trithiocarbonate potassium salt (10.5 g) is added and the mixture is stirred at room -temperature overnight. After acidification with concentrated hydrochloric acid, the mixture is extracted with ethyl acetate. The organic layer is dried over magnesium sulfate and concentrated to dryness to yield 1-[3-[[(ethylthio]t~iocarbonyl]thio]propanoyl]-L-proline.
Example 62 3-~[(~lethylamino)thiocarbonyl]thio]propionic acid Methylisothiocyanate (4 g) is added to a solution of 3-mercaptopropionic acid (5.3 g) in a mixture of pyridine (250 ml) and 0.5 N sodium hydroxide (100 ml). The solution is kept at 40 for two hours and concentrated to dryness in vacuo. The residue is dissolved in water (100 ml.), acidified with concentrated hydrochloric acid and extracted with ether. The organic phase is concentrated to dryness to yield 3-[[(methylamino)thiocarbonyl]thio]propionic acid, m.p. 86-87.
~ J~7 E~ample 63 1-[3-[[(Methylamino)thiocarbonyl]thio]propanoyl]-L-proline tert-butyl ester To a solution of L-proline t:ert-butyl ester (1.71 g) and hydroxybenzotriazole (1.35 g) in dichloromethane (10 ml) chilled and stirred in an ice bath, dicyclohexylcarbodiimide (2.06 g) and 3-methylaminothiocarbonyl-thiopropionic acid (1.79 g) are added. After 15 minutes, -the bath is removed and the stirring is continued overnight. The precipitate is filtered off and the filtrate is diluted with ethyl acetate and washed neutral.
The organic phase is concentrated to dryness to yield 1-~3-[[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline tert-butyl ester, m.p. 129-130.
Example 64 1-~3-[~(Methylamino)thiocarbonyl]thio]propanoyl]-L-proline A) l-(Methylaminothiocarbonylthiopropanoyl)-L-proline tert-butyl ester (0.98 g) is dissolved in a mixture of anisole (3.6 ml) and trifluoroacetic acia (7.5 ml). After one hour at room temperature the mixture is concentrated to dryness in vacuo and the residue precipitated from ether-hexane three times. This material is chromatographed on a column of silica gel with a solvent mixture of benzene-acetic acid (75:25) to yield 1-[3-~[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline, Rf = 0.4 [silica gel-benzene:acetic acid (75:25)]. The dicyclohexylammonium salt has m.p. 127-129.
~ '`
B) l~ethylisothiocyanate (9 g) is added to a solution of 3-mercaptopropa~oyl-L-proline (10.1 g) in a mixture oE pyridine (250 ml) and 0.5 N sodium hydroxide (100 ml). The solu-tion is ~ept at 40 for two hours and concentrated to dryness in vacuo.
The residue is dissolved in water (100 ml), acidified with con-centrated hydrochloric acid and extracted with ethyl acetate.
The organic phase is concentrated to dryness to yield 1-[3-[[(methylamino)thiocarbonyl]thio]propanoyl]-L-proline.
Example 65 1-[3-[[(Ethylamino)carbonyl]thio]propanoyl]-L-proline Ethylisocyanate (0.45 ml) is added to a solution of 1-(3-mercaptopropanoyl)-L-proline (1 g) in a mixture of N
sodium hydroxide (5 ml) and pyridine (5 ml). The solution is heated at 40 for four hours and concentrated ln vacuo.
The residue is distributed between 0.1 N hydrochloric acid and ethyl acetate. The organic layer is washed with water, dried over magnesium sulfate and concentrated to dryness to yield l-[3-[~(ethylamino)carbonyl]thio]propanoyl]-L-proline.
The dicyclohexylammonium salt is prepared by adding dicyclo-hexylamine to a solution of the free acid in ethyl acetate,m.p. 150-152.
Example 66 1-[3-[1~Ethoxy)carbonyl]thio]-2-methylpropanoyl]-L-proline By substituting 1-(3-mercapto-2-methylpropanoyl]-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[ r (ethoxy)carbonyl]thio]-2-methylpropanoyl]-L-proline is obtained.
~ ,i ~3~7 Example 67 1-[3-~[(Ethoxy)carbonyl]thio]butanoyl]-L-pro_ine By substituting 1-(3-mercaptobutanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[[(e-thoxy)carbonyl]thio]butanoyl]-L-pro-line is obtained.
Example 6~3 1-13-[[(Ethoxy)thiocarbonyl]thio]propanoyl]-L-azetidine-2-carboxylic acid .
By substituting L-azetidine-2-carboxylic acid for L-proline in the procedure oE Example 59, 1-[3-[[(ethoxy)-thiocarbonyl]thio]propanoyl]-L-azetidine-2-carboxylic acid is obtained.
Example 69 1-[3-[[(Ethoxy)thiocarbonyl]thio]propanoyl]-L-pipecolic acid By substituting L-pipecolic acid for L-proline in the procedure of Example 59, 1-[3-[[(ethoxy)thiocarbonyl]-thio]propanoyl]-L-pipecolic acid is obtained.
Example 70 1-[4-[[(Benzylthio)carbonyl]thio]butanoyl]-L-proline By substituting 4-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 60, 1-[4-[[(benzylthio)carbonyl]thio]butanoyl]-L-proline is obtained.
Example 71 1-[2-[[(Benzylthio)carbonyl]thio]propanoyl]-L-proline By substituting 2-mercaptopropanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 60, 1-[2-[[(benzylthio)carbonyl]-thio]propanoyl]-L-proline is obta:ined.
' Example 72 1-[3-[[(Ethylthio)thiocarbonyl]thio]propanoyl]-L-proline methyl ester A solution of 1-[3-[[(ethylthio)thiocarbonyl]-thio]-propanoyl]-L-proline in ethyl acetate is trea-ted wi-th an ethereal solution of diazomethane until persis-tent yellow color. AEter discharging the yellow color with a few drops of acetic acid, the solvents are removed in vacuo to yield l-[3-[[(ethylthio)-thiocarbonyl]thio]propanoyl]-L-proline methyl ester.
Example 73 1-[3-[[(methylamino)thiocarbonyl]thio]propanoyl]-5-hydroxy-L-~i~ecolic acid .
sy substituting 1-(3-mercaptopropanoyl)-5-hydroxy-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the Procedure B of Example 64, 1-[3-[[(methylamino)thiocarbonyl]thio]-propanoyl]-5-hydroxy-L-pipecolic acid is obtained.
Example 74 1-[3-[[(Methylamino)thiocarbonyl]thio]-2-methylpropanoyl]-L-proline amide By substi-tuting 1-(3-mercapto-2-methylpropanoyl)-L-proline amide for the 3-mercaptopropanoyl-L-proline in the Procedure B of Example 64, 1-[3-[[(methylaminojthiocarbonyl]thio]-2-methyl-propanoyl]-L-proline amide is obtained.
Example 75 1-[3-[[(Phenoxy)carbonyl]thio]propanoyl]-L-proline By substituting phenylchloroformate for ethyl chloroformate in the procedure of Example 58, 1-[3-[[(phenoxy)carbonyl]thio]-propanoyl]-L-proline is obtained.
5~
Example 76 l-[3-[[(Phenoxy)carbonyl]thio]butanoyl]-L-proline By substituting phenylchloroformate for the ethyl chloroformate and 4-mercaptobu-tanoy:l-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 58, 1-[3-[[(phenoxy)carbonyl]thio]butanoyl]-L-proline is obtained.
Example 77 l-[3-[[(Phenylamino)carbonyl]thio]propanoyl]-L-proline By substituting phenylisocyanate for the ethylisocyanate in the procedure of Example 65, 1-[3-[[(phenylamino)carbonyl]-thio]propanoyl]-L-proline is obtained.
Example 78 l-~3-[[(Phenethylamino)carbonyl]thio]propanoyl]-L-proline By substituting phenethylisocyanate for the ethylisocyanate in the procedure of Example 65, 1-[3-[[(phenethylamino)carbonyl]-thio]propanoyl]-L-proline is obtained.
Example 79 l-~3-[~(Ethylamino)carbonyl]thio]-2-benzylpropanoyl]-L-proline By substituting l-(3~mercapto-2-benzylpropanoyl)-L-proline for the l-(3-mercaptopropanoyl)-L-proline in the procedure of Example 65, 1-[3-[[(ethylamino)carbonyl]thio]-2-benzylpropanoyl]-L-proline is obtained.
Example 80 1-(3-Methylthiopropanoyl)-L-proline A) l~lethyl 3-methylthiopropionate (51 g) is saponified with a 10~ sodium hydro~ide solution (150 ml, 30 minutes at lOOD).
The cooled solution is extracted with ether and then acidified.
The crude acid thus obtained is distilled and converted to the acid chloride with thionyl chloride.
A solution of L-proline (11.5 g) in N sodium hydroxide (100 cc) is chilled in an ice bath and the 3-methylthiopropanoic acid chloride (6.9 g) is added dropwise with vigorous stirring over a ten minutes period. After five hours the reaction mixture is acidified and extracted with ethyl ether to yield 1-(3-methylthiopropanoyl)-L-proline. The dicyclohexylammonium salt is prepared by adding dicyclohexylamine to a solution of the free acid in ethyl acetate, m.p. 169-171.
B) ~lethyl iodide ~71 g) is added to a solution of 1-(3-mercaptopropanoyl)-L-proline ethyl ester (115 g) and sodium (11.5 g) in ethanol (400 ml). The reaction is allowed to proceed overnight, the ethanol is removed in vacuo and the residue is dissolved in a mixture of ethyl acetate and water.
Tne organic layer is dried and concentrated to dryness in vacuo.
The resulting 1-(3-methylthiopropanoyl)-L-proline ethyl ester (98 g) is suspended in a mixture of methanol (200 ml) and 5 N
sodium hydroxide (200 ml) and stirred at room temperature for five hours. The methanol is re~oved in vacuo, and the aqueous phase is extracted with ethyl acetate, acidified and reextracted with ethyl acetate. This last organic phase is washed with water, dried and concentrated to dryness to yield l-(3-methylthio-propanoyl)-L-proline.
~ -48-Example 81 __ 1-[3-(4-Chlorophenvlthio)propanoyl]-L-proline Aqueous 2 N sodium hydro~ide (25 ml) and 3-bromopropionyl chloride (8.5 g) are added to a solution of L-proline (5.75 g) in N-sodium hydro~ide (50 ml) chilled and stirred in an ice bath. After five minu-tes, the ice bath is removed and the stirring is continued for three hours at room temperature. The reaction mixture is acidified with concentrated hydrochloric acid ~nd extracted with ethyl acetate. The organic layer is washed with water, dried and concentrated to dryness in vacuo. The residue is dissolved in a mixture of 4-chlorobenzenethiol (8 g), sodium hydroxide (4.2 g) and ethanol (300 ml). The solution is refluxed for 6 hours. The solvent is removed ln vacuo and the residue is dissolved in water, acidified with concentrated hydro-chloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried, and concentrated to dryness in vacuo to yield l-[3-(4-chlorophenylthio)propanoyl]-L-proline.
Example 82 1,[~(3-Benzylthiomethyl)thio]propanoyl]-L-proline 1-(3-Mercaptopropanoyl)-L-proline (8.1 g) is dissolved ,!
in boiling liquid ammonia (100 ml) and small pieces of sodium are added until permanent blue color is obtained which is then discharged with a small piece of ammonium chloride. Benzyl-thiomethyl chloride (6.9 g) is added and the ammonia is allowed to evaporate. The final traces of ammonia are removed in vacuo, the residue is dissolved in water and extracted with ethyl acetate. The aqueous phase is acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer is washed with water, dried and concentrated to dryness to yield l-~[(3-benzylthiomethyl)-thio]propanoyl]-L,-proline.
Example 83 1-[[(3-Acetamidome-thyl)thio]pro _ oyl]-L-proline 1-(3-Mercaptopropanoyl)-L-proline (2 g) and N-hydroxy-methylacetamide (0.8~ g) are dissolved in trifluoroacetic acid (10 ml) and the solution is stored at room temperature for one hour. The excess tirfluoroacetic acid is removed in vacuo and the residue is precipitated several times from ether-hexane.
~inally, the residue is distributed between dilute hydrochloric acid and ethyl acetate. The or~anic layer is washed with water, dried and concentrated to dryness to yield l-[[3-acetamido-methyl)thio]propanoyl]-L-proline.
Example 84 l-(~ethylthioacetyl)-L-proline By substitutin~ methyl methylthioacetate for the methyl 3-methylthiopropionate in the Procedure A of Example 80, 1-(methylthioacetyl)-L-proline, m.p. 123-124, is obtained.
Example 85 l-(Benzylthioacetyl)-L-proline .
By substitutin~ benzylthioacetyl chloride for the,3-methylthiopropanoyl chloride in the Procedure A of Example 80, l-(benzylthioacetyl)-L-proline, m.p. 86-88, is obtained.
Example 86 1-[3-[(2-Phenylethyl)thio]propanoy.l]-L.-prol.ine By substituting phenethylbrom:ide for the methyl io-dide in the Procedure B of Example 80, 1-[3-[(2-phenyl-ethyl)thio]propanoyl]-L-proline is obtained.
Example 87 1-13-[(Triphenylmethyl)thio]propanoyl]-L-proline By substituting triphenylmethyl chloride for the methyl iodide in the Procedure B of Example 80, 1-~3-~(triphenylmethyl)thio]propanoyl]-L-proline is obtained.
Example 88 1-(3-Methylthio-2-methylpropanoyl)-L-proline amide By substituting 1-(3-mercapto-2-methylpropanoyl)-L-proline amide for the l-(3-mercaptopropanoyl)-L-proline ethyl ester in the Procedure B of Example 80 and elimina-ting the saponification step, l-(3-methy].thio-2-methylpro-panoyl)-L-proline amide is obtained.
Example 89 1-(3-Methylthiopropanoyl)-L-azetidine-2-carboxylic acid By substituting L-azetid.ine-2-carboxylic acid for the L-proline in the Procedure A of Example 80, 1-(3-methylthiopropanoyl)-L-azetidine-2-carboxylic acid is obtained.
;~ -~ D~br~i 3~
Example 90 1-['-(4-Methoxyphenylthio)propanoyl]-L-proline By substituting 4-methoxybenzenethiol Eor the 4-chloro-benzenethiol in the procedure of Example 81, 1-[3-(4-methoxy-phenylthio)propanoyl]-L-proline is obtained.
Example 91 1-(3-Methylthiopropanoyl)-L-pipecolic acid By substituting L-pipecolic acid for the L-proline in the Procedure A of Example 80, 1-(3-methylthiopropanoyl)-L-pipecolic acid is obtained.
Example 92l-E2-(4-Chlorophenylthio propanoyl]-L-proline sy substituting 2-bromopropionyl chloride for the -3-bromopropionyl chloride in the procedure of Example 81, 1-[2-(4-chlorophenylthio-propanoyl]-L-proline is obtained. ~`
Example 93 1-[3-~(Diphenylmethyl)thio]-2-benzylpropanoyl)]-L-proline Diphenylmethanol (0.92 g) and 1-(3-mercapto-2-benzyl-propanoyl)-L-proline (1.5 g) are dissolved in trifluoroacetic acid (10 ml) and the solution-is kept at room temperature for 30 minutes. The excess trifluoroacetic acid is removed in _acuo to yield 1-[3-[(diphenylmethyl)thio]-2-benzylpropanoyl]-L-proline.
.5"~ ..
- ,.. `=~
E~mple 94 1-[4~(4-Chlorophenylthio)butanoyl]-L-proline By substituting 4-bromoprop:ionyl chloride for the 3-bromopropionyl chloride in the procedure of Example 81, 1-[4-(4-chlorophenylthio)butanoyl]-L-proline is obtained.
Example 95 _ l-[3-[(Ben2ylthiomethyl)thio]butanoyl]-L-proline By substituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 82, 1-[3-[(benzylthiomethyl)thio]butanoyl]-L-proline is obtained.
Example 96 1-[[4-[(Acetamidomethyl)thio]-2-methylbutanoyl]-L-proline By substituting 1-(4-mercapto-2-methylbutanoyl)-L-proline for the 1-~3-mercaptopropanoyl)-L-proline in the procedure of Example 83, 1-[[4-(acetamidomethyl)thio]-2-methylbutanoyl]-L-proline is obtained.
Example 97 1-~3-(Ethyldithio)propanoyl]-L-proline A) 3-Mercaptopropanoyl-L-proline (10 g) is added to a solution of ethylthiosulfinate (8.4 g) in methanol (100 ml) and the reaction mixture is stirred vigorously at room temperature for four hours. The methanol is removed in vacuo to yield l-(3-ethyldithiopropanoyl)-L-proline.
B) A solution of ethylthiosulfinate (8.4 g) in ethanol (50 ml) is added to an aqueous solution of 3-mercapto-propanoyl-L-proline (10 g) maintained at pH 6-7 by careful addition of sodium hydroxide. The mixture is stirred vigorously ~ 53-at room temperature until negative thiol reaction. The mix-ture is diluted with water, adjusted to pH 8 and extrac-ted with ethyl acetate, the aqueous phase is acidified to pH 3 and extracted again wi-th ethyl acetate. This latter extract is washed with water, dried and concentrated to dryness to yield l-[3-(ethyldithio)propanoyl]-L-proline.
Example 98 .
1-[3-[(4-Methylphenyl)dithio]propanoyl]-L-proline A solution of 4-methylphenylsulfenyl chloride (1.76 g.) in ether (20 ml) is added to a solution of 3-mercaptopropanoyl-L-proline (2 g) ir 0.5 N sodium hydroxide (20 ml) chilled in an ice bath. The mixture is stirred vigorously for one hour, and the a~ueous phase is separated, acidified with concentrated hydrochloric acid and extracted with ethyl acetate. The organic phase is washed with water, dried and concentrated to dryness to yield 1-[[3-(4-methylphenyl)dithio]propanoyl]-L-proline.
Example 99 1-[3-(Phenyldithio-propanoyl]-L-proline By substituting phenylthiosulfinate [prepared from phenyl-disulfide according to U. Weber and P. Hartter, Z. Physiol. Chem., 351, 1384 (1970)] for the ethylthiosulfinate in the procedure of Example 97, 1-[3-(phenyldithio-propanoyl]-L-proline is obtained.
Example 100 1-[3-[(2-Phenylethyl)dithio]propanoyl]-L-proline By substituting 2-phenylethylthiosulfinate (prepared from phenethyldisulfide) for the ethylthiosuflinate in the procedure of Example 97, 1-[3-[(2-phenylethyl)dithio]propanoyl]-L-proline is obtained.
.~
E ample 101 1-[3-[(2-Hydroxyethyl)dithio]propanoyl]-L-proline To a solution oE 1,1'-[(sul~inylthio)-bis-(3-propanoyl~]-bis-L-proline (21 g) in methanol (100 ml), mercap-toethanol(4.2 g) is added and -the reaction mixture is stirred vigorously at room temperature for four hours. The methanol is removed in vacuo and the residue is pruified by chromatography on a silica gel column to yield l-[3-[(2-hydroxyethyl)dithio]propanoyl]-L-proline.
Example 102 _ .
1-[2-(Ethyldithio)propanoyl]-L-proline By substituting 2-mercaptopropanoyl-L-proline for 3-mercaptopropanoyl-L-proline in the procedure of Example 97, 1-[2-(ethyldithio)propanoyl]-L-proline is obtained.
Example 103 1-[3-~(4-Methylphenyl)dithio]butanoyl]-L-proline By sùbstituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 98, 1-[3-~4-methylphenyl)dithio]butanoyl]-L-proline is obtained.
Example 104 1-13-(Ethyldithio)-2-methylpropanoyl]-L-proline methyl ester By substituting 1-(3-mercapto-2-methylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 97 and then treat:ing the product with ethereal diazomethane as in the procedure of Example 72, 1-[3-(ethyldithio)-2-methylpro-panoyl~-L-proline methyl ester is obtained.
5~
Example 105 1-[3-(Ethyldithio)propanoyl]-L-azetidine-2-carboxylic acid By subs-tituting 3-mercaptopropanoyl-L-azetidine-2-carboxylic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 97, 1-[3-(ethyldi-thio)propanoyl]-L-aze-tidine-2-carboxylic acid is obtained.
Example 106 1-[3-[(4-Methylphenyl)dithio]-2-methylpropanoyl]-L-hydroxyproline By substituting 1-(3-mercapto-2-methylpropanoyl)-L-hydroxy proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 98, 1-[(3-[4-methylphenyl)dithio]-2-methylpropanoyl]-L-hydroxyproline is obtained.
Example 107 1-[4-~Ethyldithio)butanoyl]-L-pipecolic aeid By substituting 4-mercaptobutanoyl-L-pipecolie aeid for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 97, 1-[4-(ethyldithio-butanoyl]-L-pipeeolie aeid is obtained.
Example 108 1-13-(Ethyldithio)propanoyl]-5-hydroxy-L-pipeeolie aeid By substituting 1-(3-mereaptopropanoyl)-5-hydroxy-L-pipeeolie aeid for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 97, 1-[3-(ethyldithio)propanoyl]-5-hydroxy-L-pipecolic aeid is obtained.
.
Example 109 1-[3-[(2-Amino-2-car~o~yethyl)dithio]propanoyl]-L-proline A 0.5 M solution of thiocyano~en in glacial ace-tic acid is prepared by sha~ing Eor -ten minutes in a sealed flask 600 mg of dry lead thiocyanate with a solution of 75 1l1 of bromine in 3 ml of acetic acid. After removal of lead bromide and excess lead thiocyanate by centrifugation, 2.5 ml of this solution is mixed with 2.5 ml of a 0.41 M solution of cysteine hydrochloride previously neutralized with dilute sodium hydroxide. This mixture is immediately added to 0.75 ml of a 1.9 M solution of 3-mercaptopropanoyl-L-proline previously neutralized with dilute sodium hydroxide. After twenty minutes the mixture is titrated to incipient brown color with alcoholic iodine, and adjusted to pH 3. The precipitate is removed by filtration and the filtrate is applied to a column of cation exchange resin (Dowex 50). The column is washed with water until no more acidic material is removed and then eluted with pyridine-acetate buffer pH 6Ø
The fractions containing the disulfide of cysteine and 3-mercaptopropanoyl-L-proline are pooled and concentrated to dryness.
Example 110 1,1'-~Dithiobis)4-propanoyl)]-bis-L-proline 3-Mercaptopropanoyl-L-proline (0.95 g) is dissolved in water (20 ml) and the pH is adjusted to 6.5 with N-sodium hydroxide. An ethanolic solution of iodine is added drop-wise while maintaining the pH at 6.5 with careful addition of N sodium hydroxide. When a permanent yellow color is obtained the addition of iodine is stopped and the color , .~
:' ~ ',. .. .
~.~Ll, 31~d57 is discharc~ed with a small amount of sodium thiosulfate.
The reaction mixture is acidified with concentra-t~d hydro-chloric acid and extracted with ethyl aceta-te. The organie phase is washed with wa-ter, dried and concentrated to dryness to yield l,l'-~dithiobis(3-prop~noy~l)]-bis-L-proline. The di-cyclohexylammonium salt is prepared by addition o~ dicyclo-nexylamine to a solution of the free acid in acetonitrile, m.p. 179-180.
Example 111 .
1,1'-[Dithiobis(2-D-methyl-3-propanoyl)]-bis-L-proline By substituting 3-mercapto-2-D-methylpropanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-D-methyl-3-propanoyl)]-bis-L-proline is obtained, m.p. 236-237.
Example 112 1,1'-[Dithiobis(2-propanoyl)-bis-L-proline By substituting 2-mercaptopropanoyl-L-proline for the 3-mereaptopropanoyl-L-proline in the proeedure of Example 110, 1,1'-[dithiobis)2-propanoyl)]-bis-L-proline is obtained.
Example 113 l,l'-(Dithiobisaeetyl)-bis-L-hydroxy proline By substituting 1-(2-mereaptoacetyl)-L-hydroxyprolin~
for the 3-mercaptopropionyl-L-proline in the procedure of Example 110, l,l'-(dithiobisaeetyl)-bis-L-hydroxyproline is obtained.
Example 114 1,1'-(Dithiobisaeetyl)-bis-L-azetidine-2-earboxylie aeid By substituting 1-(2-mereaptoaeetyl)-L-azetidine-2-earboxylie aeid Eor the 3-mereaptopropanoyl-L-proline in the proeedure of Example 110, l,l'-(dithiobisaeetyl)-bis-L-azetidine-2-earboxylie aeid is obtained.
.
- Example 115 1,1'-[Dithiobis(3-propanoyl)]-bis-L-pipecolic acid sy substituting 3-mercaptopropanoyl-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-L-pipecolic acid is obtained.
Example 116 1,1'-[Dithiobis(3-propanoyl)]-bis-4-methyl-L-proline By substituting 1-(3-mercaptopropanoyl)-4-methyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-4-methyl-L-proline is obtained.
Example 117 1,1'-[Dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid By substituting 1-(3-mercaptopropanoyl)-S-hydroxy-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the pro-cedure of Example 110, 1,1'-[dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid is obtained.
Example 118 1,1'-[Dithiobis(2-benzyl-3-propanoyl)]-bis-L-proline By substituting 1-(3-mercapto-2-benzylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-benzyl-3-propanoyl)]-bis-L-proline is obtained.
Example 119 1,1'-[Dithiobis)2-methyl-3-propanoyl)]-bis-L-pipecolic acid sy substit~lting 1-(3-mercap-to-2-methylpropanoyl)-L-pipecolic acid for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[di-thiobis(2-methyl-2-propanoyl)]-bis-L-pipecolic acid is obtained.
Example 120 1,1'-[Dithiobis)4-butanoyl)]-bis-L-proline By substituting 4-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(4-butanoyl)]-bis-L-proline is obtained.
Example 121 1,1'-[Dithiobis(2-benzyl-4-butanoyl)]-bis-L-proline By substituting 1-(4-mercapto-2-benzylbutanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(2-benzyl-4-butanoyl)]-bis-L-proline is obtained.
Example 122 1,1'-[Dithiobis(3-butanoyl)]-bis-L-proline By substituting 3-mercaptobutanoyl-L-proline for the 3-mercaptopropanoyl-L-proline in the procedure of Example 110, 1,1'-[dithiobis(3-butanoyl)]-bis-L-proline is obtained.
Exam le 123 1,1'-[Dithiobis(3-propanoyl)]-bis-L-proline methyl es e A solution of 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in methanol is treated with ethereal diazomethane until persistent yellow color. After fifteen minutes a few drops of acetic acid are added and -the solvents are removed in vacuo to yield, l,l'-[dithiobis(3-propanoyl)]-bis-L-proline methyl ester.
Example 124 1,1'-[Dithiobis(3-propanoyl)]-bis-L-proline amide A solution of 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline methyl ester in methanol is saturated with am-monia while cooling in an ice-water bath. The reaction mixture is stored for 16 hours at room temperature in a pressure bottle, and then the solvents are removed in vacuo to yield l,l'-[dithiobis(3-propanoyl)]-bis-L-proline amide.
Example 125 1,1'-[Dithiobis(2-phenyl-3-propanoyl)]-bis-L-proline By substituting 1-(3-mercapto-2-phenylpropanoyl)-L-proline for the 3-mercaptopropanoyl-L-proline in the proceaure of Example 110, 1,1'-[dithiobis(2-phenyl-3-pro-panoyl)]-bis-L-proline is obtained.
2~;~
Example 126 1,1'-[(Sulfin_lthio)-bis-~3-propanoyl)]-bis-L-proline While cooling in an ice bath 0.12 mole of peracetic acid is added to a stirred solution of l,l'-[dithiobis(3-propanoyl)]-bis-L-proline (40 g) in glacial acetic acid (500 ml). The reac-tion mixture is allowed to stand over-night at room temperature and -the solvent is then removed _ vacuo to yield 1,1'-[(sulfinylthio)-bis-(3-propanoyl)]-bis-L-proline.
Example 127 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-L-proline A 30~ solution of hydrogen peroxide (2.0 ml) is added to a solution of l,l'-[dithiobis(3-propanoyl)~-bis-L-proline (4 g) in glacial acetic acid (80 ml) and the solution is stored for thirty hours at room temperature.
The solvent is removed in vacuo to yield l,l'-[(sulfonyl-thio)-bis-(3-propanoyl)]-bis-L-proline.
Example 128 1,1'-[(Sulfinylthio)-bis-(2-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-propanoyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-propanoyl)]-bis-L-proline is obtained.
:
'.~
i7 Exa ple 129 1,1'-[(S lfinylt_io)-bis-acetyl]-bis-L-azetidine-2-carboxy-lic acid By substituting l,l'-(dithiobisacetyl)-bis-L-azeti-dine carboxylic acid for the l,l'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-acetyl]-bis-L-azetidine-2-carboxylic acid is obtained.
Example 130 1,1'-[(Sulfinylthio)-bis-(3-propanoyl)]-bis-4-methyl-L-proline By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-4-methyl-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(3-propanoyl)]-bis-4-methyl~L-proline is obtained.
Example 131 1,1l-[(Sulfinylthio)-bis-(2-benzyl-3-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-benzyl-3-propa-noyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-benzyl-3-propanoyl)]-bis-L-proline is obtained.
Example 132 1,1'-[(Sulfinylthio)-bis-(4-butanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(4-butanoyl)]-bis-L-proline for the 1,1'-[dithlobis(3-propanoyl)]-bis-L-proline in -the procedure of Example 126, l,l'-[(sulfinyl-thio)-bis-(4-but.anoyl)]-bis-L-proline is obtained.
~ -63-3~
Example 133 1,1'-[(Sulfinylthio)-bis-(3-butanoyl)]-bis--L-proline By substituting 1,1'-[di-thiobis(3-butanoyl)]-bis-L-proline for the l,l'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sulfinylthio)-bis-(3-butanoyl)]-bis-L-proline is obtained.
Example 134 1,1'-[(Sulfinylthio)-bis-(2-methyl-3-propanoyl)-bis-L-pro-line By substituting 1,1'-[dithiobisl2-methyl-3-propan-oyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,l'-[(sul-finylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-proline is obtained.
Example 135 1,1'-[(Sulfinylthio)-bis-(2-phenyl-3-propanoyl)]-bis-L-proline By substituting 1,1'-[dithiobis(2-phenyl-3-propa-noyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 126, l,1'-[(sul-finylthio)-bis-(2-phenyl-3-propanoyl)]-bis-L-proline is obtained.
Example 136 1-[3-[[3-(2-Carboxy-l-pyrrolidinyl)-3-oxopropyl]-dithio]-2-methylpropanoyl]-L-proline By substituting 1,1'-[(sulfinylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-proline for the ethylthiosulfinate in the procedure of Example 97, 1-[3-[[3-(2-carboxy-1-pyrro-lidinyl)-3-oxopropyl]dithio]-2-methylpropanoyl]-L-proline is obtained.
.~ ~
Example 137 l,l'-[(Sulfon~lthio)-bis-acetyl)-bis-L-hydroxyproline By substituting l,l'-(dithiobisacetyl)-bis-L-hy-droxy proline for the l,l'-[di-thiobis(3-propanoyl)-bis-L-proline in the procedure of Example 127, l,l'-[(sulfonyl-thio)-bis-acetyl)-bis-L-hydroxypro:Line is obtained.
Example 138 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-L-pipecolic acid By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-L-pipecolic acid for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 127, l,l'-[(sul-fonylthio)-bis-(3-propanoyl)]-bis-L-pipecolic acid is ob-tained.
Example 139 1,1'-[(Sulfonylthio)-bis-(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid By substituting 1,1'-[dithiobis(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid for the 1,1'-[dithiobis-(3-propanoyl)]--bis-L-proline in the procedure of Example 127, 1,1'-~(sulfonylthio)-bis-(3-propanoyl)]-bis-5-hydroxy-L-pipecolic acid is obtained.
Example 140 1,1'-[(Sulfonylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-pipecolic acid By substituting 1,1'-[dithiobis(2-methyl-3-propa-noyl)]-bis-L-pipecolic acid for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the procedure of Example 127, 1,1'-[(sulfonylthio)-bis-(2-methyl-3-propanoyl)]-bis-L-pipecolic acid is obtained.
~ -65-E~ le 141 1,1'-[(Sulfonylthio)-bis-(2-be~ yl-~-butanoyl)]-bls-L-proline By substituting 1,1'-[dithiobis(2-benzyl-4-butan-onyl)]-bis-L-proline for the 1,1'-[dithiobis(3-propanoyl)]-bis-L-proline in the proc~dure of Example 127, l,l'-[(sul-fonylthio)-bis-(2-benzyl-'-butanoyl)]-bis-L-proline is ob-tained.
Exall~ple 142 3-Acetylthio-2-phenylpro~ noic acid By substituting 2-phenylacrylic acid for the meth-acrylic acid in the procedure of Example 25, 3-acetylthio-2-phenylpropanoic acid is obtained.
1-(3-Acetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester By substituting 3-acetylthio-2-phenylpropanoic acid for the 3-acetylthio-2-me~hylpropanoic acid in the proce-dure of Example 28, 1-(3-~cetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester is obtained.
Exa~nple 143 1-(3-Mercapto-2-phenylprc~anoyl)-L-proline By substituting 1-(3-acetylthio-2-phenylpropanoyl)-L-proline tert-butyl ester for the 1-(3-acetylthio-2-methyl-propanoyl-L-prolin~ tert-butyl ester in the pro-cedure of Example 29, and subjecting the product to am-monolysis as in Example 3~, 1-(3-acetylthio-2-phenylpro-panoyl)-L-proline and l-~,-mercapto-2-phenylpropanoyl)-L-proline are obtained.
-6 ~ am~ 4~
1-[3-(Acetylthio)-DL-pro~ yl]p pecolic acid _ Pipecolic acid (6.5 g.) is suspended in 200 ml. of dimetnylacetamid~. 3-acetylthiopropanoyl chloride (8.3 g.) is added dropwise at 23 to the suspension. A clear solu-tion forms and the temperature rises to 28. To this clear solution is added ~-methylmorpholine (10.1 g.). An imme-diate precipitate forms and the tempera-ture rises to 34.
The mixture is heated on a steam bath for 1 hour when a clear solution forms. On cooling, the precipitated solid is filtered to yield 5.1 g. of 1-[3-(acetylthio)-DL-pro-panoyl]pipecolic acid, m.p. 190-200. The solvent is re-moved and the viscous residue is triturated with isopropyl ether to yield 7.8 g. of product, m.p. 98-101. Recrystal-lization ~rom acetone-hexane yields a constant melting solid, m.p. 102-104; Rf 0.72 [silica gel, benzene, acetic acid (7:2)].
Example 145 DL-1-(3-Mercaptopropanoyl)pipecolic acid 12 ml. of concentrated ammonium hydroxide is stirred under nitrogen at 10 for about 15 minutes, then solid 1-[3-(acetylthio)-DL-propanoyl]-pipecolic acid (6.6 g.) is added at 5 to 10. A clear solution forms after 2-3 min-utes. The ice bath is removed and the solution is stirred at room temperature under nitrogen for 45 minutes. The solution is made strongly acid with 20% HCl (cooling) and the precipitated oil is extracted with 3 x 150 ml. of ethyl acetate. The ethy~l acetate extracts are dried over magne-sium sulfate and t:he solvent is removed to yield 6.0 g. of DL-1-(3-mercaptopropanoyl)pipecolic acid, Rf 0.77 [silica gel, benzene, acetic acid (7:1)].
~ %57 HAl3sb E:;ample 1~16 1-(3-,~lercaptopropanoyl)-L-pipecolic acid By s~stituting L-pipecolic acid ~or the DL-pipecolic acid in the proceclure of Examp:Le 144 and then submitting the product to the procedure of ~xample 145, 1-[3-(acetylthio)propanoyl~-L-pipecolic acid and 1-(3-mercaptopropanoyl)-L-pipecolic acid Rf 0.80 [silica gel, - benzene, acetic acid (7:1)], [~1 -51.5 (c, 1.0 abs.
ethanol), are obtained.
Example 1~7 1-[3-(Acetylthio)-2-methylpropanoyl-DL~pipecolic acid 6.5 g. (0.05 m.) of pipecolic acid are suspended in dimethylacetamide (200 mg.), 9.0 cJ. (0.05 m.) of 3-acetylthio-2-methylpropanoyl chloride is added dropwise. `
The temperature rises to 29 and a clear solution forms.
Then 10.1 g. of N-methylmorpholine is added all at once and the temperature rises to 3~1 . The mix-ture is heated on a steam bath for 1 hour when a clear solution forms.
This is allowed to stand at room temperature overnight and the solid which precipitates is filtered to yield 6.1 g., m.p. 203-204 . The solvent is removed and the viscous residue is triturated with water and 20~ HCl. The yellow oil is extracted with 3 x 150 ml. of ethyl acetate.
~he ethyl acetate extracts are dried over magnesium sulfate and removed to yield 14 g. of 1-[3-(ace-tylthio)-2-methyl-propanoyl-DI.--pipecolic acid as a viscous oil.
~xample 148 1-(3-Mercapto-2-meth~lpropanoyl)-DL-pipecolic acid Aqueous N~I~OII (30 ml. wa-ter and 20 ml. conc. NH40H) is stirred under nitrogen at lO for 15 minutes. This is -G~-~ IA13Sb added to 13.0 g. (0.05 m) of 1-[3-(acetylthio)-2-methylpropanoyl]-DL-pipecolic acid and the resulting solution is stirred for 10 minutes under nitrogen; then at room temperature for 50 miIlutes. It is then treated with water and 20~ HCl and the yellow oil extracted with 3 x 150 ml.
of ethyl acetate. The ethyl acetate extract is dried over magnesium sulfate and removed to yield 11.1 g. 1-(3-mercapto-2-methylpropanoyl)-DL-pipecolic acid as a viscous oil. Rf 0.62 [silica gel, benzene, acetic acid (7:2)]
Example 149 3-[(4-Methoxyphenyl)methylthio]-2-methylpropanoic acid p-Methoxy-~-toluene thiol (15.4 g., 0.1 mol.) is added to a solution of methacrylic acid (8.6 g., 0.1 mol.) in 50 ml. 2N sodium hydroxide. The mixture is heated on the steam bath for three hours, then refluxed for two hours and cooled. The mixture is extracted with ether, then the a~ueous layer is acidified with concentrated HCl and extracted with dichloromethane. The acidic extracts are washed with brine, dried (MgSO4) and evaporated in vacuo.
The resulting semi-solid is taken up in 50 ml. of dichloro-methane, diluted with 50 ml. hexane, and chilled. 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid is collected as a white crystalline solid, m.p. /4-82 (5.5 g.).
~xample 150 1-[3-(~l-Methoxyphenyl)methylthio]-2-methylpropanoyl-L
_ proline tert-butyl ester 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid ~3.6 g., 0.015 mol.), L-proline tert-butyl ester (2.6 g., 0.015 mol.), and dicyclohexylcarbodiimide (3.1 g., _~j9_ ~ 135b 0.015 mol.~ are dissolvecl in 50 ml. o~ dichloromethane and stirred thirty minutes at 0 . The cooling bath is removed and the mixture stirred overnight (sixteen hours).
The resulting suspension is filtered and the filtrate washed with 5~ potassium bisulfate, saturated sodium bicarbonate and brine, then dried ~ gS04 ) ancl evaporated in vacuo.
The resulting clear oil is applied to a 250 ml. silica gel column and chromatograpihed using 20~ ethyl acetate/
hexane as eluant. The main fraction (~f = 0.70, silica gel, ethyl acetate) is evaporated to 5.5 g. (93%) of 1-[3-(4-methoxyphenyl)methylthio)-2-methylpropanoyl-L-proline tert-butyl ester as a clear oil. Rf = 0.70 (silica gel, ethyl acetate); Rf = 0.60 (silica gel, ether).
Example 151 1-(3-Mercapto-2-meth~lpropanoyl)-L-prolinc The ester from Example 150 (1.2 g., 0.003 mol.), anisole t5 ml.) and -trifluoromethanesulfonic acid (0.5 ml.) are dissolved in 20 ml. of trifluoroacetic acid under nitrogen, and the resulting red solution let stand one hour at room temperature. The solution is evaporated in vacuo to a red residue which is taken up in ethyl acetate and washed with water, brine, then dried (MgSO4) and evaporated.
The residue is repeatedly triturated with hexane and the residual hexane evaporated; the oil residue amounts to 0.4 g. A portion (180 mg.) of this material is subjected to preparative thin-layer chromatography on 2 mm silica gel plates using benzene/acetic acid 75:25 as eluant. The main nitroprusside-positive band (Rf = 0.40) is recovered, affording 135 mg. of 1-(3-mercapto-2-methylpropanoyl)-L-proline as an oil. 'l`L.C using benzene/acetic acid 75:25 ~ . . _ .. . . .. _ _ .. _ IIA135b (Rf = 0.~0) and chloroLor~ lethanol/acetic acid 50:40:10 (Rf = 0.62).
Example 152 1-(3-1~lercapto-2-D-meth _~ropal_yl)-l,-prolltle IJnder a blanket of aryoll l-[3-(acetylthio)-2-~me~ylpropanoyl]-L-proline (10.0 g.) is slurried in water (150 ml.) at 10. To this mixture is added 5N
sodium hydroxide and the pH of the solution maintained at 13 for 1.5 hours. After this time, when the uptake of sodium hydroxide had ceased, the solution is acidified to a pH = 2.0 with concentrated sulfuric acid.
The aqueous solution is then extracted three times with methylene chloride (3 x 150 ml.) and the combined methylene chloride fractions concentrated to an oil. The concentrate is taken up in ethyl acetate, filtered and the filtrate diluted with hexane (30 ml.).
An additional amount of hexane is added after 1/2 hour and then the mixture cooled to 10 for 1 hour.
The crystals are filtered and washed with hexane (2 x 25 ml.) and dried to constant weight to give 1-(3-mercapto-2-D-methylpropanoyl)-L-proline as white crystals, 6.26 g., m.p. 100-iO2 .
Example 153 1-[3-Tosyloxy-2-metllylpropanoyl]-L-proline By substituting 3-tosyloxy-2-methylpropanoic acid chloride for the 3-acetylthio-2-methylpropanoic acid chloride in the procedure of Example 29b 1-[3-tosyloxy-2-methylpropanoyl]-L-prolille is obtained.
_ / I _ . .
~L ~ ; 11 A 1 3 5 b L:~;amp 1 e 1 5 ~1 1- [3-.~cetylthio-2-methylpropanoyl 1 -L-prc)line 1-[3-Tosyloxy-2-methylpropanoyl~-I,-proline (3.5 g.) is added to a solution of thiolacetic acid -`
(1.14 g.), and triethylamille (3. 5 ml . ) in ethyl acetate (20 ml.). The solution is maintained at 50 for three hours, cooled, diluted with ethyl acetate (100 ml.), and washed with dilute hydrochloric acid. The organic layer is dried and concentra-ted to dryness in vacuo. The residue is dissolved in acetonitrile and dicyclohexylamine is added. The crystalline precipitate is recrystallized from isopropanol to yield l-[3-acetylthio-2-D-methyl-pro~anoyl)-L-proline, dicyclohexylamine salt, m.p.
187-188 ~ [~]D -67 (c 1,~, EtO~I). This salt is converted to the free acid, m.p. 83-85 (an isomorphic form of m.p. 104-105 is obtained if the crystalli2ing solution is seeded with high melting material).
Example 155 1-(3-~lercaptopropanoyl)-L-proline, t-butyl ester To a stirred solution of 1.71 g. (10 mmoles) of proline t-butyl ester and 1.35 g. (10 mmoles) of 1-hydroxybenzotriazole hydrate~ in 20 ml. of N,N-dimethyl-formamide at 0-5 are added 2.06 g. (10 mmole) of N,N'-dicyclohexylcarbodiimide. The mixture is stirred for 10 minutes, followed by the addition of 1.06 g. (10 mmole) of 3-mercaptopropanoic acid in 2 ml. of N,N-dimethyl-formamide- The miY~ture is then stirred at 0-5 for 1 hour, and at room tem~erature overnight.
The precipitated N,N'-dicyclohcxylurea is filtercd o~E, and the ~iltrate concentrated in vacuo.
~ . . . . . _ _ _ . . ._ IIAl 3 Sb The residue is taken up in ethyl acetate, washed thoroughly with saturated aqueous sodi~ bicarbonate, dried, and concentrated in vacuo to 2.5 g. of oil.
The oil is taken up in 1:1 ethyl acetate-hexane and applied to a silica c3el column (100 g.). Elution wi~h 1:1 ethyl acetate-hexane affords 1.40 g. (54~) of 1-(3-mercaptopropanoyl)-L-proline, t-butyl ester as an oil, which crystallizes on standing. Recrystallization from ether-hexane yields 0.9 g. of colorless crystalline solid, m.p. 55-60 , identical to the compound of Example 17.
Example 156 1-(3-~lercaptopropanoyl)-L-proline A solution of 75 mg. (0.27 mmole) of 1-[3-[[(ethyl-amino)carbonyl]thio]propanoyl]-L-proline in 1 ml~ each of concentrated ammonium hydroxide and water is allowed to stand at room temperature for 18 hours under argon. The solution is diluted with a small amount of water and extracted with ether. The aqueous layer is acidified with cold concentrated hydrochloric acid and extracted with ethyl acetate. The combined extracts are dried and concentrated in vacuo to give a compound identical witll the product of Example 18. TEC (silica gel; benzene:acetic acid 7:3) Rf 0.4.
Example 157 Methacryloyl-L-Proline L-proline (23.0 g., 0.2 mol.) is dissolved in 100 ml. wate:r and stirred in an ice bath. Methacryloyl chloride (19.6 ml., 0.2 mol.) in 25 ml. of methyl isobutyl ketone is adclecl dropwise over three hours. Sodium hydroxide
1-[3-(Acetylthio)-DL-pro~ yl]p pecolic acid _ Pipecolic acid (6.5 g.) is suspended in 200 ml. of dimetnylacetamid~. 3-acetylthiopropanoyl chloride (8.3 g.) is added dropwise at 23 to the suspension. A clear solu-tion forms and the temperature rises to 28. To this clear solution is added ~-methylmorpholine (10.1 g.). An imme-diate precipitate forms and the tempera-ture rises to 34.
The mixture is heated on a steam bath for 1 hour when a clear solution forms. On cooling, the precipitated solid is filtered to yield 5.1 g. of 1-[3-(acetylthio)-DL-pro-panoyl]pipecolic acid, m.p. 190-200. The solvent is re-moved and the viscous residue is triturated with isopropyl ether to yield 7.8 g. of product, m.p. 98-101. Recrystal-lization ~rom acetone-hexane yields a constant melting solid, m.p. 102-104; Rf 0.72 [silica gel, benzene, acetic acid (7:2)].
Example 145 DL-1-(3-Mercaptopropanoyl)pipecolic acid 12 ml. of concentrated ammonium hydroxide is stirred under nitrogen at 10 for about 15 minutes, then solid 1-[3-(acetylthio)-DL-propanoyl]-pipecolic acid (6.6 g.) is added at 5 to 10. A clear solution forms after 2-3 min-utes. The ice bath is removed and the solution is stirred at room temperature under nitrogen for 45 minutes. The solution is made strongly acid with 20% HCl (cooling) and the precipitated oil is extracted with 3 x 150 ml. of ethyl acetate. The ethy~l acetate extracts are dried over magne-sium sulfate and t:he solvent is removed to yield 6.0 g. of DL-1-(3-mercaptopropanoyl)pipecolic acid, Rf 0.77 [silica gel, benzene, acetic acid (7:1)].
~ %57 HAl3sb E:;ample 1~16 1-(3-,~lercaptopropanoyl)-L-pipecolic acid By s~stituting L-pipecolic acid ~or the DL-pipecolic acid in the proceclure of Examp:Le 144 and then submitting the product to the procedure of ~xample 145, 1-[3-(acetylthio)propanoyl~-L-pipecolic acid and 1-(3-mercaptopropanoyl)-L-pipecolic acid Rf 0.80 [silica gel, - benzene, acetic acid (7:1)], [~1 -51.5 (c, 1.0 abs.
ethanol), are obtained.
Example 1~7 1-[3-(Acetylthio)-2-methylpropanoyl-DL~pipecolic acid 6.5 g. (0.05 m.) of pipecolic acid are suspended in dimethylacetamide (200 mg.), 9.0 cJ. (0.05 m.) of 3-acetylthio-2-methylpropanoyl chloride is added dropwise. `
The temperature rises to 29 and a clear solution forms.
Then 10.1 g. of N-methylmorpholine is added all at once and the temperature rises to 3~1 . The mix-ture is heated on a steam bath for 1 hour when a clear solution forms.
This is allowed to stand at room temperature overnight and the solid which precipitates is filtered to yield 6.1 g., m.p. 203-204 . The solvent is removed and the viscous residue is triturated with water and 20~ HCl. The yellow oil is extracted with 3 x 150 ml. of ethyl acetate.
~he ethyl acetate extracts are dried over magnesium sulfate and removed to yield 14 g. of 1-[3-(ace-tylthio)-2-methyl-propanoyl-DI.--pipecolic acid as a viscous oil.
~xample 148 1-(3-Mercapto-2-meth~lpropanoyl)-DL-pipecolic acid Aqueous N~I~OII (30 ml. wa-ter and 20 ml. conc. NH40H) is stirred under nitrogen at lO for 15 minutes. This is -G~-~ IA13Sb added to 13.0 g. (0.05 m) of 1-[3-(acetylthio)-2-methylpropanoyl]-DL-pipecolic acid and the resulting solution is stirred for 10 minutes under nitrogen; then at room temperature for 50 miIlutes. It is then treated with water and 20~ HCl and the yellow oil extracted with 3 x 150 ml.
of ethyl acetate. The ethyl acetate extract is dried over magnesium sulfate and removed to yield 11.1 g. 1-(3-mercapto-2-methylpropanoyl)-DL-pipecolic acid as a viscous oil. Rf 0.62 [silica gel, benzene, acetic acid (7:2)]
Example 149 3-[(4-Methoxyphenyl)methylthio]-2-methylpropanoic acid p-Methoxy-~-toluene thiol (15.4 g., 0.1 mol.) is added to a solution of methacrylic acid (8.6 g., 0.1 mol.) in 50 ml. 2N sodium hydroxide. The mixture is heated on the steam bath for three hours, then refluxed for two hours and cooled. The mixture is extracted with ether, then the a~ueous layer is acidified with concentrated HCl and extracted with dichloromethane. The acidic extracts are washed with brine, dried (MgSO4) and evaporated in vacuo.
The resulting semi-solid is taken up in 50 ml. of dichloro-methane, diluted with 50 ml. hexane, and chilled. 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid is collected as a white crystalline solid, m.p. /4-82 (5.5 g.).
~xample 150 1-[3-(~l-Methoxyphenyl)methylthio]-2-methylpropanoyl-L
_ proline tert-butyl ester 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid ~3.6 g., 0.015 mol.), L-proline tert-butyl ester (2.6 g., 0.015 mol.), and dicyclohexylcarbodiimide (3.1 g., _~j9_ ~ 135b 0.015 mol.~ are dissolvecl in 50 ml. o~ dichloromethane and stirred thirty minutes at 0 . The cooling bath is removed and the mixture stirred overnight (sixteen hours).
The resulting suspension is filtered and the filtrate washed with 5~ potassium bisulfate, saturated sodium bicarbonate and brine, then dried ~ gS04 ) ancl evaporated in vacuo.
The resulting clear oil is applied to a 250 ml. silica gel column and chromatograpihed using 20~ ethyl acetate/
hexane as eluant. The main fraction (~f = 0.70, silica gel, ethyl acetate) is evaporated to 5.5 g. (93%) of 1-[3-(4-methoxyphenyl)methylthio)-2-methylpropanoyl-L-proline tert-butyl ester as a clear oil. Rf = 0.70 (silica gel, ethyl acetate); Rf = 0.60 (silica gel, ether).
Example 151 1-(3-Mercapto-2-meth~lpropanoyl)-L-prolinc The ester from Example 150 (1.2 g., 0.003 mol.), anisole t5 ml.) and -trifluoromethanesulfonic acid (0.5 ml.) are dissolved in 20 ml. of trifluoroacetic acid under nitrogen, and the resulting red solution let stand one hour at room temperature. The solution is evaporated in vacuo to a red residue which is taken up in ethyl acetate and washed with water, brine, then dried (MgSO4) and evaporated.
The residue is repeatedly triturated with hexane and the residual hexane evaporated; the oil residue amounts to 0.4 g. A portion (180 mg.) of this material is subjected to preparative thin-layer chromatography on 2 mm silica gel plates using benzene/acetic acid 75:25 as eluant. The main nitroprusside-positive band (Rf = 0.40) is recovered, affording 135 mg. of 1-(3-mercapto-2-methylpropanoyl)-L-proline as an oil. 'l`L.C using benzene/acetic acid 75:25 ~ . . _ .. . . .. _ _ .. _ IIA135b (Rf = 0.~0) and chloroLor~ lethanol/acetic acid 50:40:10 (Rf = 0.62).
Example 152 1-(3-1~lercapto-2-D-meth _~ropal_yl)-l,-prolltle IJnder a blanket of aryoll l-[3-(acetylthio)-2-~me~ylpropanoyl]-L-proline (10.0 g.) is slurried in water (150 ml.) at 10. To this mixture is added 5N
sodium hydroxide and the pH of the solution maintained at 13 for 1.5 hours. After this time, when the uptake of sodium hydroxide had ceased, the solution is acidified to a pH = 2.0 with concentrated sulfuric acid.
The aqueous solution is then extracted three times with methylene chloride (3 x 150 ml.) and the combined methylene chloride fractions concentrated to an oil. The concentrate is taken up in ethyl acetate, filtered and the filtrate diluted with hexane (30 ml.).
An additional amount of hexane is added after 1/2 hour and then the mixture cooled to 10 for 1 hour.
The crystals are filtered and washed with hexane (2 x 25 ml.) and dried to constant weight to give 1-(3-mercapto-2-D-methylpropanoyl)-L-proline as white crystals, 6.26 g., m.p. 100-iO2 .
Example 153 1-[3-Tosyloxy-2-metllylpropanoyl]-L-proline By substituting 3-tosyloxy-2-methylpropanoic acid chloride for the 3-acetylthio-2-methylpropanoic acid chloride in the procedure of Example 29b 1-[3-tosyloxy-2-methylpropanoyl]-L-prolille is obtained.
_ / I _ . .
~L ~ ; 11 A 1 3 5 b L:~;amp 1 e 1 5 ~1 1- [3-.~cetylthio-2-methylpropanoyl 1 -L-prc)line 1-[3-Tosyloxy-2-methylpropanoyl~-I,-proline (3.5 g.) is added to a solution of thiolacetic acid -`
(1.14 g.), and triethylamille (3. 5 ml . ) in ethyl acetate (20 ml.). The solution is maintained at 50 for three hours, cooled, diluted with ethyl acetate (100 ml.), and washed with dilute hydrochloric acid. The organic layer is dried and concentra-ted to dryness in vacuo. The residue is dissolved in acetonitrile and dicyclohexylamine is added. The crystalline precipitate is recrystallized from isopropanol to yield l-[3-acetylthio-2-D-methyl-pro~anoyl)-L-proline, dicyclohexylamine salt, m.p.
187-188 ~ [~]D -67 (c 1,~, EtO~I). This salt is converted to the free acid, m.p. 83-85 (an isomorphic form of m.p. 104-105 is obtained if the crystalli2ing solution is seeded with high melting material).
Example 155 1-(3-~lercaptopropanoyl)-L-proline, t-butyl ester To a stirred solution of 1.71 g. (10 mmoles) of proline t-butyl ester and 1.35 g. (10 mmoles) of 1-hydroxybenzotriazole hydrate~ in 20 ml. of N,N-dimethyl-formamide at 0-5 are added 2.06 g. (10 mmole) of N,N'-dicyclohexylcarbodiimide. The mixture is stirred for 10 minutes, followed by the addition of 1.06 g. (10 mmole) of 3-mercaptopropanoic acid in 2 ml. of N,N-dimethyl-formamide- The miY~ture is then stirred at 0-5 for 1 hour, and at room tem~erature overnight.
The precipitated N,N'-dicyclohcxylurea is filtercd o~E, and the ~iltrate concentrated in vacuo.
~ . . . . . _ _ _ . . ._ IIAl 3 Sb The residue is taken up in ethyl acetate, washed thoroughly with saturated aqueous sodi~ bicarbonate, dried, and concentrated in vacuo to 2.5 g. of oil.
The oil is taken up in 1:1 ethyl acetate-hexane and applied to a silica c3el column (100 g.). Elution wi~h 1:1 ethyl acetate-hexane affords 1.40 g. (54~) of 1-(3-mercaptopropanoyl)-L-proline, t-butyl ester as an oil, which crystallizes on standing. Recrystallization from ether-hexane yields 0.9 g. of colorless crystalline solid, m.p. 55-60 , identical to the compound of Example 17.
Example 156 1-(3-~lercaptopropanoyl)-L-proline A solution of 75 mg. (0.27 mmole) of 1-[3-[[(ethyl-amino)carbonyl]thio]propanoyl]-L-proline in 1 ml~ each of concentrated ammonium hydroxide and water is allowed to stand at room temperature for 18 hours under argon. The solution is diluted with a small amount of water and extracted with ether. The aqueous layer is acidified with cold concentrated hydrochloric acid and extracted with ethyl acetate. The combined extracts are dried and concentrated in vacuo to give a compound identical witll the product of Example 18. TEC (silica gel; benzene:acetic acid 7:3) Rf 0.4.
Example 157 Methacryloyl-L-Proline L-proline (23.0 g., 0.2 mol.) is dissolved in 100 ml. wate:r and stirred in an ice bath. Methacryloyl chloride (19.6 ml., 0.2 mol.) in 25 ml. of methyl isobutyl ketone is adclecl dropwise over three hours. Sodium hydroxide
-7~-' i7 50` z.ion (2N) is ~n~ed simult~neo~sl~, maintaining the pH
of the ~eaction mixture at 7Ø Addition of base is con-tinued for four hours after addition of acid chloride has bee~ completed. The reaction mixture is adju~ted to pH 5 wi~ concentrated HCl and extracted with ethyl acetate.
The aqueous layer is then acidified to pH 2 A 5 and extracted tho-oughly with ethyl acetate. The acidic extracts are washed with brine ana dried (MgSO4). The ethyl acetate solution is treated with dicyclohexylamine (40 ml.) and chilled overnight. The resulting white precipitate is fil.ered and dried, ~ielding 29 g. (39~) of white solid, m p. 202-210~. The solid is crystallized from 1.5 liters 3:1 acetonitrile/iso?ropanol to yield 19.7 g. of metha-crtrloyl-L-proline, cicyclohexylamine salt as fine white nee-les, m.p. 202-210D.
The salt is dissolved in water/ethyl acetate and the mixture acidified with concentrated HCl. The result-ins suspension is filtered to remove a fine white preci-pit_te which is washed well with etnyl acetate. The fil-tra.e is saturated with sodium chloride and extractedtho=oughly with ethyl acetate. The extracts are washed with brine, dried (~5gS04) and evaporated to a clear oil which solidifies. Crystallization rom ethyl acetate/hex-ane yields 7.5 g. (83~) of methacryloyl-L-proline as a whi.e crystalline solid, m.p. 89-93. An analytical sam?le is obtained by recrystallization, m.p. 95-98.
Example 158 1-(3-Acetylthio-2-D-methylpropanoyl)-L-proline Methacryloyl-L-proline (183 mg., 0.0001 mol.) is dissolved in thiolacetic acid (0.5 ml.) and allowed }3A135b to stand at room tempera~ul-e for sixteen hours. The solution is evaporated in vacuo to a yellow residue.
Preparative thin layer chromat:ocjraphy (silica gel, di-chloromethane/methanol/acetic acid 90:5:5) allows isolation of a clear oil (240 m~.) as the main fraction. TLC (dichloro-methane/methanol/acetic acid ~0:5:5) shows this material to be l-(3-acetylthio-2-DL-methylpropanoyl)-L-proline corresponding to the product of Example 29B. Rf = 0.35;
(benzene/acetic acid 75:25~ Rf = 0.38.
The oil is dissolved in 3 ml. acetonitrile, treated with dicyclohexylamine until the solution is basic, and chilled. ~ white crystalline solid (106 mg.) m.p. 175-181 , is collected. Crystallization from isopropanol gives 1-(3-acetylthio-2-D-methylpropanoyl)-L-proline, dicyclohexylamine salt, m.p. 187-188 , identical with this product in Example 29A.
Example 159 l-[Dithiobis-(2-methyl-3-propanoyl)]-bis-L-proline By substituting 3,3'-dithiobis-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the pro-cedure of Example 29B, l-[dithiobis-(2-methyl-3-propanoyl)l-bis-L-proline is obtained.
Exarnple 160 1-(3-Mercapto-2 - methylpropanoyl)-L-proline Zinc dust (10.0 g.) is added to a slurry of the product of Example 15~ (5.0 g.) in 100 ml. of l.O N
sulfuric acid and the mixture is stirred at 18 for four hours under a hlanket of nitrogen. The solution is then filtered, the Zinc washed with wat~r (~0 ml.) and the combined fi:Ltrates are ~xtracted with methylene chloride '. ': ' - ", ': ' '' , ' ~ -J~
HA135b (3 x 75 ml.). The methyl~lle chloride washes are back extracted with ~ater (25 ml.) and then the or~anic solution concentrated to an oil. This oil is taken up in ethyl acetate (20 ml.) and filtered. Ilexane (15 ml.) is added to the filtrate and the mixture is stirred for 15 minutes. After this time, an additional volume of hexane (30 ml.) is added and the solution cooled to 5 for 1 hour. The mixture is then filtered, and the product is washed with hexane (2 x 10 ml.) and dried to give 4.17 g.
of white crystals of the product, 1-(3-mercapto-2-methylpropanoyl)-L-proline. TLC, Rf = 0.60 (Solvent system: benzene/acetic acid 75:25).
Example 161 3-Benzylthio-2-methylpropanoic acid By substituting a-toluenethiol for p-methoxy-~-toluenethiol in the procedure of Example 149, 3-benzylthio-2-methylpropanoic acid is obtained.
Example 162 1-[3-tBenzylthio)-2-methylpropanoyl]-L-proline tert.
butyl ester By sub~tituting 3-benzylthio-2-methylpropanoic acid for -the 3-[(4-methoxyphenyl)methylthio]-2-methyl-propanoic acid in the procedure of Example 150, 1-~3-~benzylthio)-2-methylpropanoyl]-L- proline tert. butyl ester is obtained.
~xample 163 1-[3-(Benzylthio)-2-methylpropanoyl]-L-proline 1-[3-(benzylthio),-2-methylpropanoyl]-L-proline tert. butyl ester (7.8 g.) is dissolved in a mixture of anisole (55~ml.) and triEluoroacetic acid (110 ml.). After ~lA135b one hour storage at room temperature, the solvent is removed in vacuo and the residue is dissolved in ether, washed several times t~ith saturated sodium chloride, dried over magnesium sulfate and evaporated to clryness in vacuo to yield 1-[3-(benzylthio)-2-methylpropanoyl]~L-proline.
Rf 0.5 (Silica gel, Benzenetacetic acid 3:1) RE 0.5.
(Silica gel, Methyl-ethylketone~acetic acid/pyridine/water 14:1:2:1).
Example 164 1-(3-Mercapto-2-methylpropanoyl)-L-proline 1-[3-(benzylthio)-2-methylpropanoyl]-L-proline (0.1 g.) is suspended in boiling liquid ammonia (10 ml.) and small pieces of sodium are added with stirring until persistent blue color. The color is discharged with a few crystals of ammonium sulfate and the ammonia is allowed to evaporate under a current of nitrogen. The residue is dissolved in a mixture of dilute hydrochloric acid and ethyl acetate. The organic layer is dried and concentrated to dryness in vacuo to yield l-(3-mercapto-2-methylpropanoyl)-L-proline. Rf: 0.35 (Silica gel; Benzene/acetic acid 3:1), Rf 0.5 (Silica gel; Methyl-ethylketone/acetic acid/
pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 165 3-Triphenylmethyl-thio-2-methylpropanoic acid A solu-tion of 3-mercapto-2-methylpropanoic acid (1.2 g.) and tritylchlori~e (2.~ g.) in methylene chloride (50 ml.) is kept at room temperature for 2 hours.
The mixture is warmed in a stcam bath Eor 20 minu-tes and then evaporated t:o dryness in vacuo and the residue is dissolved LJ~ `ir~ 3r~ ~7 HAl35b in saturated aqueous sodi~l bicarbonate and the solution is washed with ethyl acetate. The aqueous phase is acidified to pH 3 and e~-tracted with ethyl acetate. The organic layer is dried and conc~ntrated -to dryness to give 3-triphcnylmethylthio-2-methylpropanoic acid. Rf 0.8 (Silica gel, Benzene/ace-tic acid 3:1).
Example 166 l-[3-(Triphenylmethylthio)-2-methylpro~noyl]-L-proline tert.butyl ester By substituting 3-triphenylmethylthio-2-methyl-propanoic acid ~or the 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid in the procedure oE Example 150, 1-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline tert.butyl ester is obtained.
Example 167 1-[3-(Triphenylme-thylthio)-2-methylpropanoyl]-L-proline 3-Triphenylmethylthio-2-methylpropanoic acid (1.8 g.) and N,N'-carbonyldiimidazole (0.8 g.) are dissolved in tetrahydrofuran (lO ml.) with stirring at room temperature. After -twenty minutes, the solution is added to a mixture of L-proline (0.6 g.) and N-methyl-morpholine (l g.) in dimethylacetamide (20 ml.). The resulting mixture is stirred overnight at room temperature, çoncentra-ted to dryness and the residue dissolved in a mixture of ethyl acetate and 10% aqueous potassium bisulfate.
The organic layer is separated and dried and concentrated tc, dryness in vacuo to obtain l-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline. Rf: = 0.4 (Silica gel, Benzene/acetic acid 3:1), Rf l.0 (Silica gel, Methyl-ethyl-ketone/acetic acid/pyridine/wa-ter 14:1:2:1).
~IAl35b Example 168 1-[3-Mercapto-2-methylpropanoyl)-L-proline l-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline tert.butyl ester (5 g.) is dissolved in a mixture of anisole t55 ml.) and -trifluoroacetic acid (llO ml.).
After one hour storac3e at room temperature, the solvents are removed in vacuo and the residue is applied to a column of silica gel equilibrated with benzene:acetic acid (75:25) and eluted with the same solvent. The fractions corresponding to the component with Rf 0.40 (TLC silica gel wi-th same system) are pooled and concentrated to dryness to yield l-[3-mercapto-2-methylpropanoyl)-L-proline. Rf 0.62 (silica gel, chloroform/methanol:
acetic acid:water 50:40:lO), identical to the compound of Example 34.
Example l63 3-(Tetrahydropy-ran-2-ylthio)-2-methylpropanoic acid To a solution of 3-mercapto-2-methylpropanoic acid (2.4 g.) and freshly distillled 2,3-dihydro-4H-pyrane (l.9 g.) in benzene (60 ml.), boron trifluoride etherate (2.8 g.) is added. After two hours, potassium carbonate (4 g.) is added, the mixture is stirred and filtered. The filtrate is concentrated to dryness to yield 3-(tetrahydropyran-2-ylthio)-2-methylpropanoic acid.
Example l70 l-[3-(Tetrahydropyran-2-ylthio)-2-methylpropanoyl]-L-proline By substituting 3-(tetrahydropyran-2-ylthio)-2-methylpropanoic acid for the 3-triphenylmethylthio-2-methylpropanoic acid in the procedure of Example 167, l-[3-(tetrahydropyran-2-yl-thio)-2-methylpropanoyl]-L-proline ;f~ ~IA135b is obtained. R~: 0.8 (~ilica gel, Benzene/acetic acid 3:1; Rf: 0.75 (Silica gel, Methyl-ethylketone/Acetic acid/
pyridine/water; 14:1:2:1).
Example 17l 1-(3-~Iercapto-2-methylpropanoyl)-L-proline A solution of 1-[3-(tetrahydropyran-2-ylthio)-2-methylpropanoyl)-L-proline (1 g.) in a mixture of methanol (25 ml.) and concentrated hydrochloric acid (25 ml.) is stored at room temperature for 30 minutes. The solvents are removed in vacuo to yield 1-(3-mercapto-2-methyl-propanoyl)-L-proline. Rf: 0.35 (silica gel, Benzene/
acetic acid, 3:1), Rf 0.5 ~silica gel, Methyl-ethylketone/
acetic acid/pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 17 ?
3-Acetamidomethylthio-2-methylpropanoic acid 3-Mercapto-2-methylpropanoic acid (2.4 g.) and N-hydroxymethylacetamide (1.8 g.) are dissolved in trifluoroace-tic acid and the solution is stored at room temperature for one hour. The trifluoroacetic acid is removed in vacuo and the residue is dried in vacuo over potassium hydroxide to yield 3-acetamidomethylthio-2-- methylpropanoic acid.
Example 173 1-[3-(Acetamidomethylthio)-2-methylpropanoyl]-L-proline By substituting 3-acetamidomethyl-thio-2-methyl-propanoic acid ~or the 3-(tetrahydropyran-2-yl thio)-2-methylpropanoic acid in -the procedure of Example 170 1-[3-(acetamidomethylthio)-2---~0--.
' J~
IIA135b methylpropanoyl]-L-proline is obtained. RE 0.2 (Silica gel, Benzene/aeetic aeid 3:1) Rf 0.3 (Silica ~el, Methyl-ethylketone/acetic acid/pyridine/water 14:1:2:1).
Example 17'~
1-(3-~lercapto-2-methylpropanoyl)-I.-proline 1-[3-(acetamidomethylthio)-2-methylpropanoyl]-L-proline (1.4 g.) and mercuric aeetate (1.93 g.) are dissolved in a mixture of aeetie acid (25 ml.) and water (25 ml.).
After one hour stirring on the s-team bath, hydrogen sulfide is bubbled through until no more precipitation of mercuric sulfide is observed. The mixture is-filterd, the precipitate is washed with ethanol, and the filtrate is concentrated to dryness in vacuo to yield l-(3-mercapto-2-methylpropanoyl)-L-proline. Rf: 0.35 (Siliea gel, Ben~ene/Aeetie aeid 3:1);
Rf : 0.5 (Siliea gel, Methyl-ethylketone/Acetic acid/
pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 175 1-(3-Mercapto-2-methylpropanoyl)-L-proline tert. butyl ester 2Q To the cold (5 ) solution of 1.2 g. (10 mMol.) of 3_ mercapto-2-methylpropanoic acid and 1.7 g. (10 mMol.) of L-proline tert. butyl ester in 25 ml. dichloromethane - 2.26 g. of dieyelohexylcarbodiimide in 5 ml. dichloromethane is added in portions. After 2 hours at room temperature, 5 drops of acetie aeid are added, the mixture is filtered and the filtrate evaporated to an oily residue. This residue is t:aken up in 20 ml. of pe-troleum ether-ethyl aeetate (3:].) and applied to a 150 ml. silica gel column prepared in petroleum ether. The fraction eluted with petroleum ether-ethyl acetate (1:1) contains the product, ~IA135b 1-(3-mercapto-2-methylpropanoyl)-L-prolirle tert.butyl ester.
This fraction ~0.6 g.) is dried over P2O5 in vacuo for 12 hours Rf 0.6 (Silica gel, Benzene/Acetic acid 3:1), Rf 0.8 (Silica gel, Methyl-ethylketone/acetic acid/pyridine/
water 14:1:2:1).
Example 176 1-(3-Mercapto-2-methylpropanoyl)-L-proline By substitu-ting 1-(3-rr~ercapto-2-methylpropanoyl)-L-proline tert. butyl es-ter for the l-(3-mercaptopropanoyl-L-proline tert.butyl ester in the procedure of Example 18C, 1-(3-mercapto-2-methylpropanoyl)-L-proline is obtained.
Rf 0.35 (Silica gel, Benzene/acetic acid 3:1), Rf 0.5 (Silica gel, Methyl-ethylketone/Acetic acid/Pyridine/Water 14:1:2:1), identical to the compound of Example 34.
-~2-I-IAl35b Tlle racc~mic ~orm OL ~nc final procluct in any of the ,oregoing examples is produced by utilizing tlle DL-form of the starting arnino aeid instead of the I,-form.
Similarly, the D-form o~ thc? final produets in any of the ~oregoing e~a~l"les is i~roduced by utili~irlcJ t}le D-form of the starting ami.lo acid insteacl of the I.-form.
E.~ample 177 1000 tablets each containillc3 100 mc~. of 1-(2-mereaptopropanoyl)-L-proline are produced from the follo~ing 10 insredients:
1-(2-l~1ereaptopropanoyl)-L-proline 100 cJ.
Corn stareh 50 CJ, Gelatin 7 5 ~J
Avieel (mieroerystalline eellulose) 25 c;.
~lagnesium stearate 2.5 cJ.
The 1-(2-Mereaptopropanoyl)-L-proline and eorn stareh are admixed with an aqueous solution oE the gelatin.
The mixture is dried and c3round to a fine powcler. The Avicel and then the magnesium stearate are admi~ed ~ith the granulation. This is then eompressed in a tablet to ~orm 1000 tablets eaeh eontaining 100 mg. of aetive inc;reciient.
E.~ample 178 - By substituting 100 g. of 1-(3-rnereap~o-2-D-methylpropanoyl)-L-proline for the 1-(2-mereaptopropanoyl)-L-proline in Example 177, 1000 tablets eaeh eontaining 100 mg.
of the 1-(3-mereapto-2-D-methylpropanoyl-L-proline are produced.
-~3-HA 135b 1000 tablets each containincj 200 rng. of 1-(2-mercaptoacetyl)-L-proline ~re produced from ~he following ingredients:
1-(2-Mercaptoacetyl)-L-proline200 c;.
Lactose 100 g.
~vicel 150 g.
Corn starch 50 g.
Magnesium stearate 5 cJ.
l`he 1-t2-mercaptoacetyl)-I.-proline, lactose and Avicel are admixed, then blended with the corn starch.
~lagnesium stearate is addecl. The dry mixture is compressed in a tablet press to form 1000 505 mg. tablets each containiny 200 mg. of active ingredient. The tablets are coatcd with a solution of Methocel E 15 (methyl cellulose) includiny as a color a lake containing yellow ~.
Example 180 Two piece ~1 gelatin capsules each containinc; 250 mc~.
of 1-(2-mercaptopropanoyl)-L-proline are filled with a mixturc of the following ingredien-ts:
1-(2-Mercaptopropanoyl)-L-proline 250 mg.
Magnesium stearate 7 my.
USP lactose 193 my.
Example 181 An injectable solution is produced as follows:
1-(2-Mercaptopropanoyl)-L-proline 500 mg.
Methyl paraben 5 g.
Propyl paraben 1 g.
Sodium chloride 25 g.
Watcr for injccc~ 5 1.
of the ~eaction mixture at 7Ø Addition of base is con-tinued for four hours after addition of acid chloride has bee~ completed. The reaction mixture is adju~ted to pH 5 wi~ concentrated HCl and extracted with ethyl acetate.
The aqueous layer is then acidified to pH 2 A 5 and extracted tho-oughly with ethyl acetate. The acidic extracts are washed with brine ana dried (MgSO4). The ethyl acetate solution is treated with dicyclohexylamine (40 ml.) and chilled overnight. The resulting white precipitate is fil.ered and dried, ~ielding 29 g. (39~) of white solid, m p. 202-210~. The solid is crystallized from 1.5 liters 3:1 acetonitrile/iso?ropanol to yield 19.7 g. of metha-crtrloyl-L-proline, cicyclohexylamine salt as fine white nee-les, m.p. 202-210D.
The salt is dissolved in water/ethyl acetate and the mixture acidified with concentrated HCl. The result-ins suspension is filtered to remove a fine white preci-pit_te which is washed well with etnyl acetate. The fil-tra.e is saturated with sodium chloride and extractedtho=oughly with ethyl acetate. The extracts are washed with brine, dried (~5gS04) and evaporated to a clear oil which solidifies. Crystallization rom ethyl acetate/hex-ane yields 7.5 g. (83~) of methacryloyl-L-proline as a whi.e crystalline solid, m.p. 89-93. An analytical sam?le is obtained by recrystallization, m.p. 95-98.
Example 158 1-(3-Acetylthio-2-D-methylpropanoyl)-L-proline Methacryloyl-L-proline (183 mg., 0.0001 mol.) is dissolved in thiolacetic acid (0.5 ml.) and allowed }3A135b to stand at room tempera~ul-e for sixteen hours. The solution is evaporated in vacuo to a yellow residue.
Preparative thin layer chromat:ocjraphy (silica gel, di-chloromethane/methanol/acetic acid 90:5:5) allows isolation of a clear oil (240 m~.) as the main fraction. TLC (dichloro-methane/methanol/acetic acid ~0:5:5) shows this material to be l-(3-acetylthio-2-DL-methylpropanoyl)-L-proline corresponding to the product of Example 29B. Rf = 0.35;
(benzene/acetic acid 75:25~ Rf = 0.38.
The oil is dissolved in 3 ml. acetonitrile, treated with dicyclohexylamine until the solution is basic, and chilled. ~ white crystalline solid (106 mg.) m.p. 175-181 , is collected. Crystallization from isopropanol gives 1-(3-acetylthio-2-D-methylpropanoyl)-L-proline, dicyclohexylamine salt, m.p. 187-188 , identical with this product in Example 29A.
Example 159 l-[Dithiobis-(2-methyl-3-propanoyl)]-bis-L-proline By substituting 3,3'-dithiobis-2-methylpropanoic acid for the 3-acetylthio-2-methylpropanoic acid in the pro-cedure of Example 29B, l-[dithiobis-(2-methyl-3-propanoyl)l-bis-L-proline is obtained.
Exarnple 160 1-(3-Mercapto-2 - methylpropanoyl)-L-proline Zinc dust (10.0 g.) is added to a slurry of the product of Example 15~ (5.0 g.) in 100 ml. of l.O N
sulfuric acid and the mixture is stirred at 18 for four hours under a hlanket of nitrogen. The solution is then filtered, the Zinc washed with wat~r (~0 ml.) and the combined fi:Ltrates are ~xtracted with methylene chloride '. ': ' - ", ': ' '' , ' ~ -J~
HA135b (3 x 75 ml.). The methyl~lle chloride washes are back extracted with ~ater (25 ml.) and then the or~anic solution concentrated to an oil. This oil is taken up in ethyl acetate (20 ml.) and filtered. Ilexane (15 ml.) is added to the filtrate and the mixture is stirred for 15 minutes. After this time, an additional volume of hexane (30 ml.) is added and the solution cooled to 5 for 1 hour. The mixture is then filtered, and the product is washed with hexane (2 x 10 ml.) and dried to give 4.17 g.
of white crystals of the product, 1-(3-mercapto-2-methylpropanoyl)-L-proline. TLC, Rf = 0.60 (Solvent system: benzene/acetic acid 75:25).
Example 161 3-Benzylthio-2-methylpropanoic acid By substituting a-toluenethiol for p-methoxy-~-toluenethiol in the procedure of Example 149, 3-benzylthio-2-methylpropanoic acid is obtained.
Example 162 1-[3-tBenzylthio)-2-methylpropanoyl]-L-proline tert.
butyl ester By sub~tituting 3-benzylthio-2-methylpropanoic acid for -the 3-[(4-methoxyphenyl)methylthio]-2-methyl-propanoic acid in the procedure of Example 150, 1-~3-~benzylthio)-2-methylpropanoyl]-L- proline tert. butyl ester is obtained.
~xample 163 1-[3-(Benzylthio)-2-methylpropanoyl]-L-proline 1-[3-(benzylthio),-2-methylpropanoyl]-L-proline tert. butyl ester (7.8 g.) is dissolved in a mixture of anisole (55~ml.) and triEluoroacetic acid (110 ml.). After ~lA135b one hour storage at room temperature, the solvent is removed in vacuo and the residue is dissolved in ether, washed several times t~ith saturated sodium chloride, dried over magnesium sulfate and evaporated to clryness in vacuo to yield 1-[3-(benzylthio)-2-methylpropanoyl]~L-proline.
Rf 0.5 (Silica gel, Benzenetacetic acid 3:1) RE 0.5.
(Silica gel, Methyl-ethylketone~acetic acid/pyridine/water 14:1:2:1).
Example 164 1-(3-Mercapto-2-methylpropanoyl)-L-proline 1-[3-(benzylthio)-2-methylpropanoyl]-L-proline (0.1 g.) is suspended in boiling liquid ammonia (10 ml.) and small pieces of sodium are added with stirring until persistent blue color. The color is discharged with a few crystals of ammonium sulfate and the ammonia is allowed to evaporate under a current of nitrogen. The residue is dissolved in a mixture of dilute hydrochloric acid and ethyl acetate. The organic layer is dried and concentrated to dryness in vacuo to yield l-(3-mercapto-2-methylpropanoyl)-L-proline. Rf: 0.35 (Silica gel; Benzene/acetic acid 3:1), Rf 0.5 (Silica gel; Methyl-ethylketone/acetic acid/
pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 165 3-Triphenylmethyl-thio-2-methylpropanoic acid A solu-tion of 3-mercapto-2-methylpropanoic acid (1.2 g.) and tritylchlori~e (2.~ g.) in methylene chloride (50 ml.) is kept at room temperature for 2 hours.
The mixture is warmed in a stcam bath Eor 20 minu-tes and then evaporated t:o dryness in vacuo and the residue is dissolved LJ~ `ir~ 3r~ ~7 HAl35b in saturated aqueous sodi~l bicarbonate and the solution is washed with ethyl acetate. The aqueous phase is acidified to pH 3 and e~-tracted with ethyl acetate. The organic layer is dried and conc~ntrated -to dryness to give 3-triphcnylmethylthio-2-methylpropanoic acid. Rf 0.8 (Silica gel, Benzene/ace-tic acid 3:1).
Example 166 l-[3-(Triphenylmethylthio)-2-methylpro~noyl]-L-proline tert.butyl ester By substituting 3-triphenylmethylthio-2-methyl-propanoic acid ~or the 3-[(4-methoxyphenyl)methylthio]-2-methylpropanoic acid in the procedure oE Example 150, 1-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline tert.butyl ester is obtained.
Example 167 1-[3-(Triphenylme-thylthio)-2-methylpropanoyl]-L-proline 3-Triphenylmethylthio-2-methylpropanoic acid (1.8 g.) and N,N'-carbonyldiimidazole (0.8 g.) are dissolved in tetrahydrofuran (lO ml.) with stirring at room temperature. After -twenty minutes, the solution is added to a mixture of L-proline (0.6 g.) and N-methyl-morpholine (l g.) in dimethylacetamide (20 ml.). The resulting mixture is stirred overnight at room temperature, çoncentra-ted to dryness and the residue dissolved in a mixture of ethyl acetate and 10% aqueous potassium bisulfate.
The organic layer is separated and dried and concentrated tc, dryness in vacuo to obtain l-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline. Rf: = 0.4 (Silica gel, Benzene/acetic acid 3:1), Rf l.0 (Silica gel, Methyl-ethyl-ketone/acetic acid/pyridine/wa-ter 14:1:2:1).
~IAl35b Example 168 1-[3-Mercapto-2-methylpropanoyl)-L-proline l-[3-(triphenylmethylthio)-2-methylpropanoyl]-L-proline tert.butyl ester (5 g.) is dissolved in a mixture of anisole t55 ml.) and -trifluoroacetic acid (llO ml.).
After one hour storac3e at room temperature, the solvents are removed in vacuo and the residue is applied to a column of silica gel equilibrated with benzene:acetic acid (75:25) and eluted with the same solvent. The fractions corresponding to the component with Rf 0.40 (TLC silica gel wi-th same system) are pooled and concentrated to dryness to yield l-[3-mercapto-2-methylpropanoyl)-L-proline. Rf 0.62 (silica gel, chloroform/methanol:
acetic acid:water 50:40:lO), identical to the compound of Example 34.
Example l63 3-(Tetrahydropy-ran-2-ylthio)-2-methylpropanoic acid To a solution of 3-mercapto-2-methylpropanoic acid (2.4 g.) and freshly distillled 2,3-dihydro-4H-pyrane (l.9 g.) in benzene (60 ml.), boron trifluoride etherate (2.8 g.) is added. After two hours, potassium carbonate (4 g.) is added, the mixture is stirred and filtered. The filtrate is concentrated to dryness to yield 3-(tetrahydropyran-2-ylthio)-2-methylpropanoic acid.
Example l70 l-[3-(Tetrahydropyran-2-ylthio)-2-methylpropanoyl]-L-proline By substituting 3-(tetrahydropyran-2-ylthio)-2-methylpropanoic acid for the 3-triphenylmethylthio-2-methylpropanoic acid in the procedure of Example 167, l-[3-(tetrahydropyran-2-yl-thio)-2-methylpropanoyl]-L-proline ;f~ ~IA135b is obtained. R~: 0.8 (~ilica gel, Benzene/acetic acid 3:1; Rf: 0.75 (Silica gel, Methyl-ethylketone/Acetic acid/
pyridine/water; 14:1:2:1).
Example 17l 1-(3-~Iercapto-2-methylpropanoyl)-L-proline A solution of 1-[3-(tetrahydropyran-2-ylthio)-2-methylpropanoyl)-L-proline (1 g.) in a mixture of methanol (25 ml.) and concentrated hydrochloric acid (25 ml.) is stored at room temperature for 30 minutes. The solvents are removed in vacuo to yield 1-(3-mercapto-2-methyl-propanoyl)-L-proline. Rf: 0.35 (silica gel, Benzene/
acetic acid, 3:1), Rf 0.5 ~silica gel, Methyl-ethylketone/
acetic acid/pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 17 ?
3-Acetamidomethylthio-2-methylpropanoic acid 3-Mercapto-2-methylpropanoic acid (2.4 g.) and N-hydroxymethylacetamide (1.8 g.) are dissolved in trifluoroace-tic acid and the solution is stored at room temperature for one hour. The trifluoroacetic acid is removed in vacuo and the residue is dried in vacuo over potassium hydroxide to yield 3-acetamidomethylthio-2-- methylpropanoic acid.
Example 173 1-[3-(Acetamidomethylthio)-2-methylpropanoyl]-L-proline By substituting 3-acetamidomethyl-thio-2-methyl-propanoic acid ~or the 3-(tetrahydropyran-2-yl thio)-2-methylpropanoic acid in -the procedure of Example 170 1-[3-(acetamidomethylthio)-2---~0--.
' J~
IIA135b methylpropanoyl]-L-proline is obtained. RE 0.2 (Silica gel, Benzene/aeetic aeid 3:1) Rf 0.3 (Silica ~el, Methyl-ethylketone/acetic acid/pyridine/water 14:1:2:1).
Example 17'~
1-(3-~lercapto-2-methylpropanoyl)-I.-proline 1-[3-(acetamidomethylthio)-2-methylpropanoyl]-L-proline (1.4 g.) and mercuric aeetate (1.93 g.) are dissolved in a mixture of aeetie acid (25 ml.) and water (25 ml.).
After one hour stirring on the s-team bath, hydrogen sulfide is bubbled through until no more precipitation of mercuric sulfide is observed. The mixture is-filterd, the precipitate is washed with ethanol, and the filtrate is concentrated to dryness in vacuo to yield l-(3-mercapto-2-methylpropanoyl)-L-proline. Rf: 0.35 (Siliea gel, Ben~ene/Aeetie aeid 3:1);
Rf : 0.5 (Siliea gel, Methyl-ethylketone/Acetic acid/
pyridine/water 14:1:2:1) identical to the compound of Example 34.
Example 175 1-(3-Mercapto-2-methylpropanoyl)-L-proline tert. butyl ester 2Q To the cold (5 ) solution of 1.2 g. (10 mMol.) of 3_ mercapto-2-methylpropanoic acid and 1.7 g. (10 mMol.) of L-proline tert. butyl ester in 25 ml. dichloromethane - 2.26 g. of dieyelohexylcarbodiimide in 5 ml. dichloromethane is added in portions. After 2 hours at room temperature, 5 drops of acetie aeid are added, the mixture is filtered and the filtrate evaporated to an oily residue. This residue is t:aken up in 20 ml. of pe-troleum ether-ethyl aeetate (3:].) and applied to a 150 ml. silica gel column prepared in petroleum ether. The fraction eluted with petroleum ether-ethyl acetate (1:1) contains the product, ~IA135b 1-(3-mercapto-2-methylpropanoyl)-L-prolirle tert.butyl ester.
This fraction ~0.6 g.) is dried over P2O5 in vacuo for 12 hours Rf 0.6 (Silica gel, Benzene/Acetic acid 3:1), Rf 0.8 (Silica gel, Methyl-ethylketone/acetic acid/pyridine/
water 14:1:2:1).
Example 176 1-(3-Mercapto-2-methylpropanoyl)-L-proline By substitu-ting 1-(3-rr~ercapto-2-methylpropanoyl)-L-proline tert. butyl es-ter for the l-(3-mercaptopropanoyl-L-proline tert.butyl ester in the procedure of Example 18C, 1-(3-mercapto-2-methylpropanoyl)-L-proline is obtained.
Rf 0.35 (Silica gel, Benzene/acetic acid 3:1), Rf 0.5 (Silica gel, Methyl-ethylketone/Acetic acid/Pyridine/Water 14:1:2:1), identical to the compound of Example 34.
-~2-I-IAl35b Tlle racc~mic ~orm OL ~nc final procluct in any of the ,oregoing examples is produced by utilizing tlle DL-form of the starting arnino aeid instead of the I,-form.
Similarly, the D-form o~ thc? final produets in any of the ~oregoing e~a~l"les is i~roduced by utili~irlcJ t}le D-form of the starting ami.lo acid insteacl of the I.-form.
E.~ample 177 1000 tablets each containillc3 100 mc~. of 1-(2-mereaptopropanoyl)-L-proline are produced from the follo~ing 10 insredients:
1-(2-l~1ereaptopropanoyl)-L-proline 100 cJ.
Corn stareh 50 CJ, Gelatin 7 5 ~J
Avieel (mieroerystalline eellulose) 25 c;.
~lagnesium stearate 2.5 cJ.
The 1-(2-Mereaptopropanoyl)-L-proline and eorn stareh are admixed with an aqueous solution oE the gelatin.
The mixture is dried and c3round to a fine powcler. The Avicel and then the magnesium stearate are admi~ed ~ith the granulation. This is then eompressed in a tablet to ~orm 1000 tablets eaeh eontaining 100 mg. of aetive inc;reciient.
E.~ample 178 - By substituting 100 g. of 1-(3-rnereap~o-2-D-methylpropanoyl)-L-proline for the 1-(2-mereaptopropanoyl)-L-proline in Example 177, 1000 tablets eaeh eontaining 100 mg.
of the 1-(3-mereapto-2-D-methylpropanoyl-L-proline are produced.
-~3-HA 135b 1000 tablets each containincj 200 rng. of 1-(2-mercaptoacetyl)-L-proline ~re produced from ~he following ingredients:
1-(2-Mercaptoacetyl)-L-proline200 c;.
Lactose 100 g.
~vicel 150 g.
Corn starch 50 g.
Magnesium stearate 5 cJ.
l`he 1-t2-mercaptoacetyl)-I.-proline, lactose and Avicel are admixed, then blended with the corn starch.
~lagnesium stearate is addecl. The dry mixture is compressed in a tablet press to form 1000 505 mg. tablets each containiny 200 mg. of active ingredient. The tablets are coatcd with a solution of Methocel E 15 (methyl cellulose) includiny as a color a lake containing yellow ~.
Example 180 Two piece ~1 gelatin capsules each containinc; 250 mc~.
of 1-(2-mercaptopropanoyl)-L-proline are filled with a mixturc of the following ingredien-ts:
1-(2-Mercaptopropanoyl)-L-proline 250 mg.
Magnesium stearate 7 my.
USP lactose 193 my.
Example 181 An injectable solution is produced as follows:
1-(2-Mercaptopropanoyl)-L-proline 500 mg.
Methyl paraben 5 g.
Propyl paraben 1 g.
Sodium chloride 25 g.
Watcr for injccc~ 5 1.
-8~-IIAl 35b The active substance, pr~servatives arl~ so~ium chloride are dissolved in 3 liters of water ~or injection and then the volume is hrou~ht up to 5 liters. The solution is filtered throu~h a sterile rilter and aseptically filled into presterilized vials ~hicll are then closed with pre-sterilized rubber closures. Each vial contains 5 ml. of solution in a concentration of 100 mg. of active ingrcdient per ml. of solution for injection.
Example 182 By substituting 100 g. or 1,1'-~dithiobis~2-D-methyl-3-propanoyl)]-bis-L-proline for the 1-(2-mercaptopropanoyl)-L-proline in E~ample 177, 1000 tablets each containing 100 mg.
of the l,l'-[dithiobis[2-D-methyl-3-propanoyl)]-bis-L-proline are produced.
Each of the products of the examples can be similarly formulated by substituting it for the active ingredient in Examples 177, 179, 180 or 181.
Example 182 By substituting 100 g. or 1,1'-~dithiobis~2-D-methyl-3-propanoyl)]-bis-L-proline for the 1-(2-mercaptopropanoyl)-L-proline in E~ample 177, 1000 tablets each containing 100 mg.
of the l,l'-[dithiobis[2-D-methyl-3-propanoyl)]-bis-L-proline are produced.
Each of the products of the examples can be similarly formulated by substituting it for the active ingredient in Examples 177, 179, 180 or 181.
Claims (14)
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:
1. A process for preparing a compound of the formula wherein R is hydroxy, NH2 or lower alkoxy; R1 and R4 each is hydrogen, lower alkyl, phenyl or phenyl-lower alkyl; R2 is hydrogen, lower alkyl, phenyl, substituted phenyl wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, phenyl-lower alkyl, diphenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl, phenyl-lower alkylthiomethyl, lower alkanoylamidomethyl, , , or R6-S-;
R3 is hydrogen, hydroxy or lower alkyl; R5 is lower alkyl, phenyl or phenyl-lower alkyl; R6 is lower alkyl, phenyl, substituted phenyl, wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, hydroxy-lower alkyl or amino-(carboxy)lower alkyl; M is O or S; m is 1, 2 or 3; n is 1 or 2 and p is 0, 1 or 2; and basic salts thereof, characterized by acylating a compound of the formula with a compound of the formula ,
R3 is hydrogen, hydroxy or lower alkyl; R5 is lower alkyl, phenyl or phenyl-lower alkyl; R6 is lower alkyl, phenyl, substituted phenyl, wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, hydroxy-lower alkyl or amino-(carboxy)lower alkyl; M is O or S; m is 1, 2 or 3; n is 1 or 2 and p is 0, 1 or 2; and basic salts thereof, characterized by acylating a compound of the formula with a compound of the formula ,
2. A process according to claim 1 wherein a compound of the formula is acylated with a compound of the formula to form a product of the formula
3. A process according to claim 1 wherein a compound of the formula in the L-form is acylated with a compound of the formula to form a product of the formula wherein the proline moiety is in the L-form.
4. A process according to claim 1 wherein a compound of the formula in the L-form is acylated with a compound of the formula to form a product of the formula wherein the proline moiety is in the L-form.
5. A process according to claim 1 wherein a compound of the formula in the L-form is acylated with a compound of the formula wherein R1 is C1-C4 alkyl to form a product of the formula wherein R1 is C1-C4 alkyl and the proline moiety is in the L-form.
6. A process according to claim 1 wherein a compound of the formula in the L-form is acylated with a compound of the formula to form a product of the formula wherein the proline moiety is in the L-form.
7. A process according to claim 1 wherein L-proline-t-butyl ester is reacted with propiothiolactone to form 1-(3-mercaptopropanoyl)-L-proline-t-butyl ester.
8. A compound of the formula wherein R is hydroxy, NH2 or lower alkoxy; R1 and R4 each is hydrogen, lower alkyl, phenyl or phenyl-lower alkyl; R2 is hydrogen, lower alkyl, phenyl, substituted phenyl wherein the phenyl substituent is halo, lower alkyl or lower alkoxy;
phenyl-lower alkyl, diphenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl, phenyl-lower alkylthiomethyl, lower alkanoylamidomethyl, , , or R6-S-;
R3 is hydrogen, hydroxy or lower alkyl; R5 is lower alkyl, phenyl or phenyl-lower alkyl; R6 is lower alkyl, phenyl, substituted phenyl, wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, hydroxy-lower alkyl or amino-(carboxy)lower alkyl; M is O or S; m is 1, 2 or 3; n and p each is 0, 1 or 2; and basic salts thereof, whenever prepared by the process of claim 1.
phenyl-lower alkyl, diphenyl-lower alkyl, triphenyl-lower alkyl, lower alkylthiomethyl, phenyl-lower alkylthiomethyl, lower alkanoylamidomethyl, , , or R6-S-;
R3 is hydrogen, hydroxy or lower alkyl; R5 is lower alkyl, phenyl or phenyl-lower alkyl; R6 is lower alkyl, phenyl, substituted phenyl, wherein the phenyl substituent is halo, lower alkyl or lower alkoxy, hydroxy-lower alkyl or amino-(carboxy)lower alkyl; M is O or S; m is 1, 2 or 3; n and p each is 0, 1 or 2; and basic salts thereof, whenever prepared by the process of claim 1.
9. A compound as in claim 8 having the formula whenever prepared by the process of claim 2.
10. A compound as in claim 8 having the formula wherein the proline moiety is in the L-form, whenever prepared by the process of claim 3.
11. A compound as in claim 8 having the formula wherein the proline moiety is in the L-form, whenever prepared by the process of claim 4.
12. A compound as in claim 8 having the formula wherein R1 is C1-C4 alkyl and the proline moiety is in the L-form, whenever prepared by the process of claim 5.
13. A compound as in claim 8 having the formula whenever prepared by the process of claim 6.
14. A compound as in claim 8 having the name 1-(3-mercaptopropanoyl)-L-proline-t-butyl ester, whenever prepared by the process of claim 7.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA349,476A CA1103257A (en) | 1976-02-13 | 1980-04-09 | Proline derivatives and related compounds |
Applications Claiming Priority (8)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US05/657,792 US4046889A (en) | 1976-02-13 | 1976-02-13 | Azetidine-2-carboxylic acid derivatives |
| US657,792 | 1976-02-13 | ||
| US69843276A | 1976-06-21 | 1976-06-21 | |
| US698,432 | 1976-06-21 | ||
| US05/751,851 US4105776A (en) | 1976-06-21 | 1976-12-22 | Proline derivatives and related compounds |
| US751,851 | 1976-12-22 | ||
| CA270,184A CA1101864A (en) | 1976-02-13 | 1977-01-21 | Proline derivatives and related compounds |
| CA349,476A CA1103257A (en) | 1976-02-13 | 1980-04-09 | Proline derivatives and related compounds |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1103257A true CA1103257A (en) | 1981-06-16 |
Family
ID=27508082
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA349,476A Expired CA1103257A (en) | 1976-02-13 | 1980-04-09 | Proline derivatives and related compounds |
Country Status (1)
| Country | Link |
|---|---|
| CA (1) | CA1103257A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7056935B2 (en) | 1995-06-07 | 2006-06-06 | Gpi Nil Holdings, Inc. | Rotamase enzyme activity inhibitors |
| US8653131B2 (en) | 2008-08-22 | 2014-02-18 | Baxter Healthcare S.A. | Polymeric benzyl carbonate-derivatives |
-
1980
- 1980-04-09 CA CA349,476A patent/CA1103257A/en not_active Expired
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7056935B2 (en) | 1995-06-07 | 2006-06-06 | Gpi Nil Holdings, Inc. | Rotamase enzyme activity inhibitors |
| US8653131B2 (en) | 2008-08-22 | 2014-02-18 | Baxter Healthcare S.A. | Polymeric benzyl carbonate-derivatives |
| US8962549B2 (en) | 2008-08-22 | 2015-02-24 | Baxter International Inc. | Polymeric benzyl carbonate-derivatives |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| MKEX | Expiry | ||
| MKEX | Expiry |
Effective date: 19980616 |