WO2020186840A1 - Compound containing fluorene ring structure and application thereof - Google Patents

Compound containing fluorene ring structure and application thereof Download PDF

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Publication number
WO2020186840A1
WO2020186840A1 PCT/CN2019/125143 CN2019125143W WO2020186840A1 WO 2020186840 A1 WO2020186840 A1 WO 2020186840A1 CN 2019125143 W CN2019125143 W CN 2019125143W WO 2020186840 A1 WO2020186840 A1 WO 2020186840A1
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ring structure
fluorene ring
amino acid
protected
compound containing
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PCT/CN2019/125143
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French (fr)
Chinese (zh)
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郑庆泉
梁伟周
唐齐
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广州同隽医药科技有限公司
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Publication of WO2020186840A1 publication Critical patent/WO2020186840A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C233/00—Carboxylic acid amides
    • C07C233/01—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms
    • C07C233/16—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms
    • C07C233/17—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom
    • C07C233/18—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom having the carbon atom of the carboxamide group bound to a hydrogen atom or to a carbon atom of an acyclic saturated carbon skeleton
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C233/00—Carboxylic acid amides
    • C07C233/01—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms
    • C07C233/16—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms
    • C07C233/17—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom
    • C07C233/22—Carboxylic acid amides having carbon atoms of carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms having the nitrogen atom of at least one of the carboxamide groups bound to a carbon atom of a hydrocarbon radical substituted by singly-bound oxygen atoms with the substituted hydrocarbon radical bound to the nitrogen atom of the carboxamide group by an acyclic carbon atom having the carbon atom of the carboxamide group bound to an acyclic carbon atom of a carbon skeleton containing six-membered aromatic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C235/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms
    • C07C235/02—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton
    • C07C235/04—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton the carbon skeleton being acyclic and saturated
    • C07C235/18—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton the carbon skeleton being acyclic and saturated having at least one of the singly-bound oxygen atoms further bound to a carbon atom of a six-membered aromatic ring, e.g. phenoxyacetamides
    • C07C235/20—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to acyclic carbon atoms and singly-bound oxygen atoms bound to the same carbon skeleton the carbon skeleton being acyclic and saturated having at least one of the singly-bound oxygen atoms further bound to a carbon atom of a six-membered aromatic ring, e.g. phenoxyacetamides having the nitrogen atoms of the carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C235/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms
    • C07C235/42—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to carbon atoms of six-membered aromatic rings and singly-bound oxygen atoms bound to the same carbon skeleton
    • C07C235/44—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to carbon atoms of six-membered aromatic rings and singly-bound oxygen atoms bound to the same carbon skeleton with carbon atoms of carboxamide groups and singly-bound oxygen atoms bound to carbon atoms of the same non-condensed six-membered aromatic ring
    • C07C235/58—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to carbon atoms of six-membered aromatic rings and singly-bound oxygen atoms bound to the same carbon skeleton with carbon atoms of carboxamide groups and singly-bound oxygen atoms bound to carbon atoms of the same non-condensed six-membered aromatic ring with carbon atoms of carboxamide groups and singly-bound oxygen atoms, bound in ortho-position to carbon atoms of the same non-condensed six-membered aromatic ring
    • C07C235/60—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by oxygen atoms having carbon atoms of carboxamide groups bound to carbon atoms of six-membered aromatic rings and singly-bound oxygen atoms bound to the same carbon skeleton with carbon atoms of carboxamide groups and singly-bound oxygen atoms bound to carbon atoms of the same non-condensed six-membered aromatic ring with carbon atoms of carboxamide groups and singly-bound oxygen atoms, bound in ortho-position to carbon atoms of the same non-condensed six-membered aromatic ring having the nitrogen atoms of the carboxamide groups bound to hydrogen atoms or to acyclic carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C237/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups
    • C07C237/02—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C237/22—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton having nitrogen atoms of amino groups bound to the carbon skeleton of the acid part, further acylated
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/06—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length using protecting groups or activating agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2603/00—Systems containing at least three condensed rings
    • C07C2603/02—Ortho- or ortho- and peri-condensed systems
    • C07C2603/04—Ortho- or ortho- and peri-condensed systems containing three rings
    • C07C2603/06—Ortho- or ortho- and peri-condensed systems containing three rings containing at least one ring with less than six ring members
    • C07C2603/10—Ortho- or ortho- and peri-condensed systems containing three rings containing at least one ring with less than six ring members containing five-membered rings
    • C07C2603/12—Ortho- or ortho- and peri-condensed systems containing three rings containing at least one ring with less than six ring members containing five-membered rings only one five-membered ring
    • C07C2603/18—Fluorenes; Hydrogenated fluorenes

Definitions

  • the present invention relates to the technical field of compound and polypeptide synthesis, in particular to a compound containing a fluorene ring structure and its application.
  • the synthetic manufacturing methods of peptides can be roughly divided into three categories at present: solid-phase carrier synthesis, conventional liquid-phase synthesis and liquid-phase carrier synthesis.
  • the solid-phase carrier synthesis method can complete the separation and purification work through simple solid-liquid separation, and has the advantages of strong universality and short development cycle.
  • the solid-liquid two-phase reaction has serious mass transfer problems due to the poor affinity between the solvent and the solid carrier and the spatial structure of the carrier, and the reaction speed is limited.
  • a large excess of reagents is required under conventional reaction conditions.
  • the intermediate is attached to a solid-phase carrier, which is difficult to perform routine analysis and purification.
  • the final product can only be purified by reversed-phase liquid chromatography.
  • the conventional liquid phase synthesis method is generally a homogeneous reaction with good reactivity.
  • the intermediate can be purified by washing, crystallization and other means, and the final product has a high purity.
  • the properties of the products and impurities of each reaction are quite different, and no universal method can be found for separation and purification, the development cycle is long, and the manufacturing steps are complicated.
  • the liquid carrier synthesis method is a new peptide synthesis method developed in the past ten years to improve the above two methods.
  • the more representative ones include JITSUBO's Molecular Hiving TM technology and Ajinomoto's Ajiphase technology.
  • JITSUBO Molecular Hiving TM technology uses long-chain alkoxy benzyl alcohols, such as 3,5-bis(docosyloxy)benzyl alcohol, 2,4-bis(docosyloxy) Benzyl alcohol, 3,4,5-tris(octadecyloxy)benzyl alcohol, etc. are used as liquid carrier. These carriers are used as C-terminal protective reagents for peptide synthesis.
  • the reaction is carried out in a homogeneous phase.
  • the product is precipitated by changing the solvent composition or temperature change, and separation and purification are achieved through the steps of precipitation-filtration-washing.
  • this method has the following problems: 1The solubility of the carrier in solvents such as medium and low polarity acetate and toluene is not large, and the reaction concentration is low, which is not conducive to scale-up production; 2The solvent can be changed through quantitative distillation and other operations. The composition process is complicated to operate; 3The precipitation of the solvent system is generally amorphous solid, which is very difficult to filter and wash, and the effect of removing impurities is poor and time-consuming. Therefore, it cannot be said that this method is a universal peptide synthesis method with good reproducibility.
  • Ajinomoto's Ajiphase technology has modified the former, using multi-branched alkyl groups instead of linear alkyl groups, effectively improving the solubility of the carrier in solvents such as isopropyl acetate.
  • the reaction can be carried out at a higher concentration. Water & organic polar solvent washing can achieve separation and purification.
  • this technology still has the following problems: 1The reaction time is long.
  • the patent publication No. CN107011132A uses isopropyl acetate or methylcyclopentyl ether as a solvent for condensation to react overnight, and deprotection requires 4-6 hours.
  • the purpose of the present invention is to provide a compound containing a fluorene ring structure and its application.
  • this compound as a protective carrier for peptide liquid phase synthesis, in a homogeneous or heterogeneous solvent system, especially in a heterogeneous solvent system, it can increase the reaction speed and reagent utilization, simplify post-processing operations, and improve product purity. Improving the versatility of the operating process can be developed into a universal production method.
  • One of the objectives of the present invention is to provide a compound containing a fluorene ring structure, the structure of which is shown in the general formula (1):
  • X is selected from OH, halogen, sulfonate, NHR a ;
  • Y is selected from hydrogen and aryl
  • R a is selected from hydrogen, alkyl or aralkyl
  • Q is independently selected from O, NH, NHCO, CO, CONH, S, SO or SO 2 ;
  • n 1 and n 2 are independently selected from an integer of 0 to 3, and n 1 and n 2 are not 0 at the same time;
  • R is independently selected from the group represented by the general formula (2):
  • * means connected with Q
  • R 1 is selected from a C 1 ⁇ C 25 alkyl group or a group represented by the general formula (3):
  • * means it is connected to the carbonyl group
  • n1 represents an integer of 1 to 3;
  • R 1a is selected from C 6 -C 25 alkyl groups, and the total carbon number of m1 R 1a is not less than 8;
  • k a is selected from an integer of 0 to 3;
  • Ring B except that one R 1a O having m1 substituent groups, but may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ⁇ C 5 alkyl group, a halogen atom free of C 1 ⁇ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ⁇ C 5 alkoxy group, a halogen atom free of C 1 ⁇ C 5 alkoxy group substituent;
  • R 2 is selected from hydrogen, a C 1 to C 25 alkyl group or the group represented by the general formula (4):
  • n2 is selected from an integer of 0 to 3;
  • R 2a is selected from C 6 ⁇ C 25 alkyl groups
  • k b is selected from an integer of 1 to 6;
  • an outer ring C m2 having a substituent group R 2a O may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ⁇ C 5 alkyl group, a halogen atom free of C 1 ⁇ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ⁇ C 5 alkoxy group, a halogen atom free of C 1 ⁇ C 5 alkoxy group substituent;
  • L is selected from C 2 ⁇ C 15 organic chain groups containing O, N or S heteroatoms or C 2 ⁇ C 15 organic chain groups without O, N or S heteroatoms, when L is selected from C 2 ⁇ C 15 organic chain groups that do not contain O, N or When the C 2 ⁇ C 15 organic chain group of S heteroatom, R 2 ⁇ H;
  • n 1 and n 2 th RQ substituents can also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ⁇ C 5 alkyl group, a halogen atom free of C 1 ⁇ C 5 alkyl group, substituent group substituted with halogen atom-containing C 1 ⁇ C 5 alkoxy group or a halogen atom free of C 1 ⁇ C 5 alkoxy.
  • the aforementioned R is selected from the group represented by the general formula (5):
  • * means connected with Q
  • the aforementioned R is selected from the group represented by the general formula (6):
  • * means connected with Q
  • R 4 is selected from hydrogen, a C 1 to C 25 alkyl group or the group represented by the above general formula (4);
  • k 2 is selected from an integer of 1 to 4.
  • k 3 is selected from an integer of 1-4.
  • the above R is selected from the group represented by the general formula (7):
  • * means connected with Q
  • R 6 is selected from hydrogen, C 1 to C 25 alkyl group or the group represented by the above general formula (4);
  • k 4 is selected from an integer from 0 to 3;
  • k 5 is selected from an integer of 0 to 3; preferably, k 5 is selected from an integer of 1 to 3;
  • R 5 is selected from hydrogen, a side chain group of a natural amino acid, an alkyl group or a group represented by the general formula (8):
  • * represents the connection site
  • k 6 is selected from an integer of 1 to 4.
  • R 1 ′ is selected from a C 1 ⁇ C 25 alkyl group or a group represented by the general formula (3);
  • R 2 ' is selected from hydrogen, C 1 to C 25 alkyl group, or the group represented by the general formula (4).
  • the above Q is selected from O.
  • the R a is selected from hydrogen, methyl, ethyl, propyl, benzyl or methoxybenzyl.
  • the n 1 +n 2 is equal to 1-3.
  • m1 is selected from 2 or 3, and the total carbon number of m1 R 1a is 12-60.
  • the aforementioned R 1a is selected from C 8 to C 22 alkyl groups.
  • m2 is selected from 2 or 3, and the total carbon number of m2 R 2a is 12-60.
  • R 2a is selected from C 8 to C 22 alkyl groups.
  • R 2 is selected from hydrogen, methyl, ethyl, propyl, isopropyl, isooctyl, benzyl or 4-methoxybenzyl.
  • the halogen in X in the general formula (1) is selected from Cl, Br or I.
  • the sulfonate in X in the general formula (1) is selected from methanesulfonate or p-toluenesulfonate.
  • the structural formula of the compound containing a fluorene ring structure is selected from the following:
  • the C 8 H 17 alkyl group in the above structural formula is isooctyl
  • the C 9 H 19 alkyl group is isononyl
  • the C 10 H 21 alkyl group is isodecyl
  • the C 13 H 27 alkyl group is iso Structure tridecyl
  • C 20 H 41 alkyl is 2,3-dihydrophytyl.
  • the compound containing the fluorene ring structure is easily soluble in at least one of hydrocarbon organic solvents, aromatic hydrocarbon organic solvents, ester organic solvents, ether organic solvents, and water-soluble aprotic polar organic solvents.
  • the above-mentioned hydrocarbon organic solvent is selected from at least one of heptane, hexane, petroleum ether, cyclohexane, and methylcyclohexane.
  • the above-mentioned aromatic hydrocarbon organic solvent is selected from at least one of toluene, ethylbenzene, and xylene.
  • the above-mentioned ester organic solvent is selected from at least one of isopropyl acetate, tert-butyl acetate, and ethyl acetate.
  • the ether organic solvent is at least one selected from the group consisting of diethyl ether, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, and tetrahydrofuran.
  • the water-soluble aprotic polar organic solvent is selected from the group consisting of N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, and N-methyl- Pyrrolidone, N-ethyl-pyrrolidone, dimethyl sulfoxide, sulfolane, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidine At least one of ketones.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >1% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >5% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >30% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >50% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >80% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >100% at 25-30°C.
  • the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 30% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 30%.
  • the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is >50% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is >50%.
  • the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is greater than 60% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is greater than 60%.
  • the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 80% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 80%.
  • the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 100% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 100%.
  • the solubility of the compound containing the fluorene ring structure in the ester organic solvent is >50%; more preferably, the solubility of the compound containing the fluorene ring structure in the isopropyl acetate is >50%.
  • the solubility of the compound containing the fluorene ring structure in the ester organic solvent is> 100% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the isopropyl acetate is> 100 %.
  • the solubility of the compound containing the fluorene ring structure in the ether organic solvent is greater than 30% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether >30%.
  • the solubility of the compound containing the fluorene ring structure in ether organic solvents is >50% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in methyl tert-butyl ether >50%.
  • the solubility of the compound containing the fluorene ring structure in the ether organic solvent is greater than 60% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether >60%.
  • the solubility of the compound containing the fluorene ring structure in the ether organic solvent is> 80% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether >80%.
  • the solubility of the compound containing the fluorene ring structure in the ether organic solvent is> 100% at 25-30°C; more preferably, the compound containing the fluorene ring structure is more than 100% in methyl tert-butyl ether. Solubility>100%.
  • Compound 1-7a can convert OH into halogen, sulfonate and other compounds through conventional reaction through steps 1-8.
  • the above halogen is selected from Cl, Br, and I; the above sulfonate is selected from methanesulfonate or p-toluenesulfonate.
  • R in general formulas (5), (6) and (7) can be synthesized.
  • Another object of the present invention is to provide an amino acid or peptide C-terminal protection reagent, which comprises the above-mentioned compound containing a fluorene ring structure.
  • the present invention also provides the application of the compound containing the fluorene ring structure in the synthesis of peptide reagents in a homogeneous or heterogeneous solvent system.
  • the present invention also provides a method for synthesizing peptides, which method includes using the above-mentioned compound containing a fluorene ring structure.
  • the above-mentioned method of peptide synthesis includes the following steps:
  • Carrier access The above-mentioned compound containing fluorene ring structure is used as a carrier to connect with N-protected amino acid or N-protected peptide compound through conventional reaction to obtain N-protected amino acid or N-protected C-terminal of the carrier containing fluorene ring structure Peptide compound
  • N-terminal deprotection Dissolve the N-protected amino acid or N-protected peptide compound containing the C-terminal protection of the fluorene ring structure carrier in a solvent, and add the N-terminal protection deprotection reagent solution to form a homogeneous or heterogeneous system. N-terminal deprotection, adding a highly polar solvent for extraction, to obtain N-deprotected amino acid or N-deprotected peptide compound solution containing fluorene ring structure carrier C-terminal protection;
  • Peptide chain extension Add N-protected amino acid or N-protected peptide solution to the C-terminal protected N-deprotected amino acid or N-deprotected peptide compound solution of the fluorene ring structure carrier, and then add the condensation reagent solution to form a homogeneous solution. Condensation reaction in a phase or heterogeneous system, and a highly polar solvent is added for extraction to obtain an N-protected amino acid or N-protected peptide compound solution containing the C-terminal protection of the fluorene ring structure carrier;
  • step 2) Repeat step 2) and step 3) to insert the next amino acid until a complete peptide chain is obtained.
  • the solution containing the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier in step 2) and the N-terminal protected deprotection reagent solution in step 2) form a heterogeneous system.
  • the solution containing the N-deprotected amino acid or N-deprotected peptide compound protected by the C-terminal of the fluorene ring structure carrier in step 3) and the N-protected amino acid or N-protected peptide and condensation reagent solution in step 3) Into a heterogeneous system.
  • step 2) the solvent containing the N-protected amino acid or N-protected peptide compound protected at the C-terminal of the fluorene ring structure carrier and the N-deprotected amino acid or N-protected amino acid or N-protected at the C terminal of the fluorene ring structure carrier in the step 3) -
  • the solvent in the solution of the deprotected peptide compound is independently selected from hydrocarbons or mixed solvents formed by hydrocarbons and at least one of esters, ethers, and halogenated hydrocarbons.
  • hydrocarbons in the mixed solvent formed by at least one of hydrocarbons and esters, ethers, and halogenated hydrocarbons shall not affect the formation of a heterogeneous system.
  • step 2) dissolve the solvent containing the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier and step 3) the N-deprotected amino acid or N-protected amino acid or N-protected C terminal of the fluorene ring structure carrier.
  • the solvent in the solution of the deprotected peptide compound is independently selected from at least hexane, heptane, cyclohexane, methylcyclohexane, petroleum ether or hexane, heptane, cyclohexane, methylcyclohexane, and petroleum ether.
  • the solvent for dissolving the N-terminal protected deprotection reagent in step 2), the solvent for dissolving the N-protected amino acid or N-protected peptide, and the condensation reagent in step 3) are independently selected from amide solvents.
  • the above-mentioned amide solvent is selected from N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, N-methyl-pyrrolidone, N-ethyl -At least one of pyrrolidone, 1,3-dimethylimidazolinone, and 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone.
  • the highly polar solvent in step 2) and step 3) is selected from at least one of water, alcohols, nitriles, amides, sulfoxides, sulfones, and water-soluble alcohol ethers.
  • the highly polar solvent in step 2) and step 3) is selected from water, methanol, acetonitrile, N,N-dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazoline At least one of ketone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone, dimethyl sulfoxide, and sulfolane.
  • step 2) when the N-protected amino acid or N-protected peptide compound containing the C-terminal protection of the fluorene ring structure carrier is dissolved in a solvent, the N-terminal protection deprotection reagent solution is added to form a homogeneous system.
  • the solvent containing the N-protected amino acid or N-protected peptide compound protected by the C-terminal of the fluorene ring structure carrier does not contain hydrocarbons or contains some hydrocarbons, but this part of the hydrocarbons does not affect the formation of a homogeneous system.
  • the solvent for dissolving the N-terminal protected deprotection reagent and the solvent for dissolving the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier can be ester solvents, ether solvents, halogenated hydrocarbons or amide solvents At least one of; preferably, ethyl acetate, isopropyl acetate, tert-butyl acetate, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, tetrahydrofuran, dichloromethane, chloroform, N, N-dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone In the same step 3), add the N-protected amino acid or N-protected peptide solution to the solution
  • the solvent for dissolving N-protected amino acids or N-protected peptides and condensation reagents and dissolving the N-deprotected amino acids or N-deprotected peptide compounds protected by the C-terminal of the fluorene ring structure carrier The solvent phase can be selected from at least one of ester solvents, ether solvents or amide solvents, preferably ethyl acetate, isopropyl acetate, tert-butyl acetate, dichloromethane, chloroform, tetrahydrofuran, N, N -Dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone At least one.
  • ester solvents preferably ethyl acetate, isopropyl acetate, tert-butyl acetate, dichloromethane, chloroform,
  • the target product and impurities can be well separated only by adding a highly polar solvent, especially in a heterogeneous system, thereby greatly simplifying the post-processing operation steps.
  • the amount of the N-protected amino acid or N-protected peptide is 0.8-3.0 equivalents of the compound containing the fluorene ring structure, and the amount of the condensation reagent is 0.8-3.0 equivalents of the compound containing the fluorene ring structure.
  • the amount of the N-protected amino acid or N-protected peptide is 1 to 1.1 equivalents of the compound containing the fluorene ring structure, and the amount of the condensation reagent is 1 to 1.2 equivalents of the compound containing the fluorene ring structure.
  • the compound of the present invention is introduced into an amino acid or peptide as the C-terminal protective group.
  • the operation steps can be the same as step 3. ) Same or different.
  • the present invention provides a protective carrier that can be applied to peptide liquid phase synthesis.
  • the peptide synthesis reaction using this protective carrier has fast reaction speed and high reagent utilization in a suitable solvent system; at the same time, simple Liquid-liquid extraction separation can be effectively purified, and finally a product with higher purity can be obtained, so the post-processing operation is simple; and the solubility change during the synthesis process is small, the operation flow is strong, and it can be developed into a universal production method .
  • the fluorene ring structure-containing compound of the present invention contains a non-polar and polar two-part structure, so that it can be dissolved in both non-polar solvents and polar solvents, so it is suitable for non-polar solvents and polar solvents.
  • a protective carrier for peptide synthesis reaction As a protective carrier for peptide synthesis reaction; and the peptide synthesis reaction carried out with this kind of carrier has: 1 fast reaction speed, few by-products; 2 high utilization of reagents, low reagent dosage, low cost, and less three wastes; 3after It has good separation effect for processing the extraction stratified impurities and products.
  • the product has high purity; 4The solubility and reaction rate of the intermediate compounds in the synthesis process are small, the amino acid or peptide fragments have good repeatability, simple operation, strong universality, and suitable as Advantages of universal production methods.
  • DIPEA N,N-diisopropylethylamine
  • HBTU O-benzotriazole-tetramethylurea hexafluorophosphate
  • PE petroleum ether
  • the crude intermediate 1-2 was dissolved in tetrahydrofuran (100mL), methanol (50mL) and 30% NaOH solution (67g) were added, and the reaction was stirred for 3h. Petroleum ether (100mL) and water (200mL) were added. After stirring for 10min, the mixture was separated. Lower layer; add 2N hydrochloric acid (100mL), stir for 10 min, separate the lower layer; wash the upper layer to pH 6-7, concentrate to obtain 3,4,5-tris(isooctyloxy)-benzoic acid 48.0g (1-2) ;
  • 2-hydroxy-9-fluorenone (9.8g, 0.05mol, 1-4), 2-(2-(3,4,5-tris(isooctyloxy)-benzamide)-ethoxy) -Ethyl chloride (30.6g, 0.06mol), potassium carbonate (16.6g, 0.12mol), potassium iodide (0.05g) and DMF (150mL) are mixed at room temperature, mechanically stirred, heated to 115-120°C for 8-10h, and cooled To room temperature, pour into a mixture of petroleum ether (150ml) and water (150ml) while stirring.
  • the crude oxime was dissolved in tetrahydrofuran (120mL), acetic acid (50mL) was added, heated to 55-60°C, and zinc powder (9.8g, 0.15mol) was added in batches. After the addition, keep warm and continue the reaction for 2h, filter to remove excess zinc powder, concentrate to remove tetrahydrofuran and acetic acid. Add petroleum ether (200 mL) to re-dissolve, wash with 10% NaOHaq (100 mL ⁇ 2) and saturated brine successively, and concentrate to obtain FL-001 crude product.
  • Example 2 uses isononyl bromide (3,5,5-trimethyl-hexyl bromide) to replace the one in Example 1. Isooctyl bromide;
  • Example 3 The synthesis method of FL-003 in Example 3 is the same as that in Example 1, except that: isooctyl bromide (prepared by Exxal 10 bromination) is used instead of isooctyl bromide in Example 1;
  • Exxal 10 100 mL
  • 37% HBr 300 mL
  • concentrated sulfuric acid 30 mL
  • the reaction was stopped after GC analysis of alcohol ⁇ 0.5%.
  • the temperature was lowered to below 50° C.
  • petroleum ether 100 mL was added for extraction, the petroleum ether layer was first neutralized with water, and the petroleum ether was removed under reduced pressure to obtain isodecyl bromide.
  • Example 4 The synthesis method of FL-004 in Example 4 is the same as that in Example 1, except that: n-octyl bromide, n-decyl bromide and n-dodecyl bromide (1:1:1) are used instead of the examples. 1 isooctyl bromide;
  • Example 5 The synthesis method of FL-005 in Example 5 is the same as that of Example 1, but the difference is: isotridecyl bromide (Exxal 13 is used instead of Exxal 10 in Example 3). Preparation of bromide in Example 3 Method preparation) instead of isooctyl bromide in Example 1;
  • Example 6 The synthesis method of FL-006 in Example 6 is the same as that of Example 1, except that 2,3-dihydrophytyl bromide (3,7,11,15-tetramethyl-hexadecyl Bromine) instead of isooctyl bromide in Example 1;
  • Example 7 The synthesis method of FL-007 in Example 7 is the same as that in Example 1, except that isononyl bromide is used instead of isooctyl bromide, N-benzyl-2-(2-chloroethyl)oxy-ethylamine salt Acid salt instead of 2-(2-chloroethyl)oxy)-ethylamine hydrochloride in Example 1;
  • Example 8 The synthesis method of FL-008 in Example 8 is the same as that in Example 1, except that: isooctyl bromide is replaced by iso-tridecyl bromide, and N-benzyl-2-(2-chloroethyl)oxy- Ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1;
  • the synthesis method of FL-009 in embodiment 9 is the same as that in embodiment 2, except that: 2,7-dihydroxy-9-fluorenone is used instead of 2-hydroxy-9-fluorenone;
  • intermediate 11-4 and 2-hydroxy-9-fluorenone as raw materials, according to the method steps of intermediate 1-3 to FL-001 in Example 1, synthesis (2-(2-(N'-(3, 4 -Di(isotridecyloxy)-benzyl)-acetamido)-ethoxy)-ethoxy)-fluorene-9-amine (FL-011).
  • Example 12 The synthesis method of FL-012 in Example 12 is the same as that in Example 11, except that isononanoyl chloride replaces the acetic anhydride in Example 11.
  • Example 14 The synthesis method of FL-014 in Example 14 is the same as that in Example 6, except that methyl 3,5-dihydroxybenzoate is used instead of methyl 3,4,5-trihydroxybenzoate in Example 6;
  • Example 16 The synthesis method of FL-016 in Example 16 is the same as that in Example 5, except that: N-methyl-2-chloroethylamine hydrochloride is used instead of 2-(2-chloroethyl)oxy in Example 5 -Ethylamine hydrochloride;
  • Example 17 The synthesis method of FL-017 in Example 17 is the same as that in Example 1, except that isononanoic acid replaces 3,4,5-diisooctylbenzoic acid (Intermediate 1-2) in Example 1, and N -Benzyl-2-(2-chloroethyl)oxy-ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1, with 2,7- Dihydroxy-9-fluorenone replaces 2-hydroxy-9-fluorenone;
  • Example 18 The synthesis method of FL-018 in Example 18 is the same as that in Example 1, except that: Isostearic acid replaces 3,4,5-diisooctylbenzoic acid (Intermediate 1-2) in Example 1 N-benzyl-2-(2-chloroethyl)oxy-ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1;
  • 1,3-propanediamine (11.1g, 0.15mol) was dissolved in dichloromethane (20mL), and the dichloromethane solution obtained in the previous step was added dropwise at room temperature; after the addition, react for half an hour at room temperature and wash with water (100mL ⁇ 2) Obtain the methylene chloride solution of Intermediate 19-3, which is directly used in the next reaction;
  • the dichloromethane solution obtained in the previous step was cooled to 0-5°C, and triethylamine (10g, 0.1mol) was added; the temperature was maintained at 0-5°C, and 2-chloroacetyl chloride (5.6g, 0.05mol) was added dropwise; Reacted at -5°C for 0.5h, washed with 1N hydrochloric acid (100mL), water (100mL), saturated brine (50mL), and concentrated to obtain the crude intermediate 19-4.
  • Example 20 The synthesis method of FL-020 in Example 20 is the same as that in Example 19, except that: N,N'-dibenzyl-1,3-propanediamine is used instead of 1,3-propanediamine in Example 19;
  • Boc-Ala-OH (18.9g, 0.1mol, 21-1), 2-(2-chloroethyl)oxy-ethylamine hydrochloride (16.0g, 0.1mol), HOBt (14.9, 0.11mol) and Triethylamine (15.1g, 0.15mol) was mixed with DMF (150mL) and cooled to 5-10°C; EDCI (21.1g, 0.11mol) was added, kept at 5-10°C and reacted for 0.5h, and naturally warmed to room temperature for 1h; Wash with water (100mL), saturated sodium bicarbonate (100mL), 1N hydrochloric acid (100mL) and saturated brine (50mL) successively; concentrate to obtain the crude intermediate 21-2;
  • FL-021 was synthesized according to the method of Example 5;
  • Lysine (7.3g, 0.05mol, 23-1) was dissolved in tetrahydrofuran (100mL) and water (50mL), LiOH (1.2g, 0.05mol) was added; the mixture was heated to 70°C, and active ester 19-2 ( 96.6, 0.11mol) in tetrahydrofuran (200mL); after dripping, keep the reaction for half an hour; cool to room temperature, add 1N hydrochloric acid (50mL), and extract with petroleum ether (200mL); the upper petroleum ether solution is successively water (100mL) and saturated Wash with brine (100 mL) and concentrate to obtain crude intermediate 23-2;
  • Lysine 14.6.3g, 0.1mol was dissolved in tetrahydrofuran (150mL) and water (100mL), and NaOH (4.0g, 0.1mol) was added; the reaction solution was cooled to 0-5°C; Stearyl chloride (2,2,4,8,10,10-hexamethylundecane-5-acid chloride) (66.6g, 0.22mol) and 20% NaOH (44g), control the dropping rate to maintain the reaction temperature at 0-5°C; after dripping, keep at 0-5°C and react for half an hour, adjust the pH to 1 with 1N hydrochloric acid; extract with petroleum ether (300mL); wash petroleum ether solution with water (100mL) and saturated brine (100mL) successively ; Concentrated and purified by column chromatography to obtain 50.5g of intermediate diisostearoyl lysine;
  • Example 23 Using diisostearoyl lysine instead of 23-2 in Example 23 to synthesize FL-025 according to the steps in Example 23;
  • Example 27 The synthesis method of FL-027 in Example 27 is the same as that in Example 26, except that the intermediate 27-1 replaces the intermediate 26-5 in Example 26;
  • Example 31 The synthesis method of FL-031 in Example 31 is the same as that in Example 27, except that: 2,7-dihydroxy-fluorenone replaces 2-hydroxy-fluorenone;
  • heterogeneous synthesis of H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using carrier FL-002 includes the following steps:
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-004, and the HPLC purity is 87.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-005, and the HPLC purity is 90.0%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-006, and the HPLC purity is 85.0%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-008, and the HPLC purity is 91.6%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-009, and the HPLC purity is 86.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-012, and the HPLC purity is 87.0%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-014, and the HPLC purity is 88.0%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-018, and the HPLC purity is 85.1%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-020, and the HPLC purity is 87.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-022, and the HPLC purity is 85.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-023, and the HPLC purity is 85.3%.
  • heterogeneous synthesis of H-Arg-Lys-Gly-Thr-Lys-Ser-OH using carrier FL-027 includes the following steps:
  • the synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-028, and the HPLC purity is 95.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-029, and the HPLC purity is 93.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-031, and the HPLC purity is 92.0%.
  • step 2) The isopropyl acetate solution obtained in step 1), add diethylenetriamine (6.2g, 60mmol) and mercaptopropionic acid (4.3g, 40mmol), heat to 40-50°C to react, TLC analysis after the reaction is complete, use acetonitrile /Water (9:1) washing (add isopropyl acetate as needed) to remove by-products, isopropyl acetate solution is used for the next amino acid condensation.
  • the crude H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 was obtained by cleavage, with an HPLC purity of 70.2%.
  • the synthesis method of this embodiment is the same as that of embodiment 48, except that the carrier of embodiment 48 is replaced with FL-020, and the HPLC purity is 68.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 48, except that the carrier of embodiment 48 is replaced with FL-023, and the HPLC purity is 71.5%.
  • the synthesis method of the embodiment is the same as that of embodiment 51, except that the carrier of embodiment 51 is replaced with FL-031, and the HPLC purity is 80.0%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with REF-001, and the HPLC purity is 73.5%.
  • the synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with REF-002, and the HPLC purity is 67.8%.
  • the synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with REF-003, and the HPLC purity is 83.0%.
  • the synthesis method of this example is the same as that of Example 44, except that: the carrier of Example 44 is replaced with REF-004, and the HPLC purity is 80.5%.
  • the method of carrier activation in this example is the same as step 1) of Example 44, the method of connecting the first amino acid is the same as step 2) of Example 44, and the method of removing Fmoc and other amino acids is the same as that of Comparative Example 9. It is: the carrier used is REF-003, and the HPLC purity is 50.5%.
  • FL-022 >50 >50 >50 >100 ⁇ 20 >10 FL-023 >100 >100 >100 >80 ⁇ 1 >20 FL-024 >50 >100 >50 >100 ⁇ 5 >10 FL-025 >30 >50 >30 >100 ⁇ 5 >20 FL-026 >100 >100 >100 >100 >100 ⁇ 1 ⁇ 1 FL-027 >100 >100 >100 >100 >100 ⁇ 1 ⁇ 1 FL-028 >100 >100 >100 >100 >100 >100 ⁇ 1 ⁇ 1 FL-029 >100 >100 >100 >100 >100 >100 ⁇ 1 ⁇ 1 FL-030 >100 >100 >100 >100 >30 ⁇ 1 ⁇ 1 FL-031 >100 >100 >100 >100 >30 ⁇ 1 ⁇ 1 REF-001 >100 >100 >100 >100 ⁇ 5 ⁇ 1 ⁇ 1 REF-002 >100 >100 >100 >100 >100 ⁇ 1 ⁇ 1 ⁇ 1 REF-003 >100 >100 >100 >100 ⁇ 5
  • the compound of the present invention has better properties in heptane, isopropyl acetate (i-PrOAc), methyl tert-butyl ether (MTBE), and N,N-dimethylformamide (DMF).
  • i-PrOAc isopropyl acetate
  • MTBE methyl tert-butyl ether
  • DMF N,N-dimethylformamide
  • the deprotection reaction time is more than 2.5 hours, and the post-deprotection treatment time is more than 2 hours; and the condensation reaction time of the compound of the present invention is 0.5-1.0 hours, the post-condensation treatment time is 0.5-1.0 hours, and the deprotection reaction time is 0.5-1.0 hours.
  • the processing time after deprotection reaction is 0.5 ⁇ 1.0 hours, the time required for each amino acid fragment access step is very small, and the repeatability is good; and the comparative example REF-001 ⁇ 002 requires each amino acid fragment access step The time varies greatly.
  • the same result is obtained in the heterogeneous system compared to the comparative examples REF-003 ⁇ 004: the condensation reaction time, The processing time after the condensation reaction, the deprotection reaction time, and the post-processing time of the deprotection reaction are significantly shortened.
  • the access time of each amino acid in the peptide chain elongation stage (AA 2 -AA 5 ) changes very little.
  • Each post-treatment time is 0.5 ⁇ 1.0h, with good repeatability.
  • the solubility of the compounds in Comparative Examples REF-001 ⁇ 004 decreased rapidly during the process of peptide synthesis. During the post-treatment process, the solvent needs to be continuously added to maintain the solution state, and the gelation phenomenon began to appear from AA 4 .
  • the solubility of the compound of the present invention is basically unchanged and has a good solubility; the compound of the present invention is used as a carrier to synthesize peptides in a heterogeneous system, and the purity of the peptide with an amide at the carbon end is greater than 85%, and some are greater than 90%.
  • the purity of the peptides with carboxyl at the carbon end is greater than 90%, and some are greater than 95%, while the purity of the products with amide REF-001 ⁇ REF-002 at the carbon end of the comparative example is not higher than 75%, and the purity of the product with carboxyl at the carbon end of REF-003 ⁇ REF -004 purity is not higher than 85%.
  • the condensation reaction time, post-condensation treatment time, and deprotection of peptide synthesis as a carrier The reaction time and the post-treatment time of the deprotection reaction are shorter, and the product purity is higher.
  • the condensation reaction time using the compound of the present invention as a carrier is also Obviously shortened, generally less than 5h, while the comparison carrier is generally greater than 5h, some amino acids are 10h, and the product purity of the product using the compound of the invention as a carrier is also significantly higher.
  • the compound of the present invention as a carrier is suitable for both homogeneous and heterogeneous systems, and has better effects than the compounds in the comparative example.
  • the compounds of the present invention are more effective than those in heterogeneous systems. It has better effect in homogeneous system, especially in the two-phase system (heterogeneous system) formed by medium and low polar solvents and amide solvents, for peptide synthesis, it can significantly shorten the condensation reaction time and post-condensation treatment Time, deprotection reaction time, and post-treatment time of deprotection reaction, and finally obtain a product with higher purity.

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Abstract

Disclosed are a compound containing a fluorene ring structure and an application thereof. The compound containing the fluorene ring structure of the present invention contains a hydroxyl group, an amino group, a substituted amino group, and an active group, and can be used as an amino acid or peptide C-terminal protection reagent. A synthesis reaction of a peptide using such a protective carrier has a high reaction speed and a high reagent utilization rate in a suitable solvent system; post-treatment is performed by means of simple liquid-liquid extraction separation to achieve effective purification, so that a product having high purity is finally obtained; a change in solubility is small in the synthesis process, the operating process has strong universality, and a universal production method can be developed into.

Description

一种含有芴环结构的化合物及其应用A compound containing fluorene ring structure and its application 技术领域Technical field
本发明涉及化合物及多肽合成技术领域，尤其涉及一种含有芴环结构的化合物及其应用。The present invention relates to the technical field of compound and polypeptide synthesis, in particular to a compound containing a fluorene ring structure and its application.
背景技术Background technique
肽的合成制造方法，目前大体可以分成三类：固相载体合成法、常规液相合成法和液相载体合成法。The synthetic manufacturing methods of peptides can be roughly divided into three categories at present: solid-phase carrier synthesis, conventional liquid-phase synthesis and liquid-phase carrier synthesis.
固相载体合成法通过简单的固液分离即可完成分离纯化的工作，具有普适性强、开发周期短等优点。但固-液两相反应由于溶剂和固相载体间的亲和性较差、载体的空间结构等原因，存在较为严重的传质问题，反应速度受到限制。为了弥补低反应性，在常规反应条件下，反应试剂需要大大过量。中间体附载在固相载体上，难以进行常规分析，也无法进行纯化，最终产物一般情况下只能通过反相液相制备色谱纯化。The solid-phase carrier synthesis method can complete the separation and purification work through simple solid-liquid separation, and has the advantages of strong universality and short development cycle. However, the solid-liquid two-phase reaction has serious mass transfer problems due to the poor affinity between the solvent and the solid carrier and the spatial structure of the carrier, and the reaction speed is limited. In order to compensate for the low reactivity, a large excess of reagents is required under conventional reaction conditions. The intermediate is attached to a solid-phase carrier, which is difficult to perform routine analysis and purification. The final product can only be purified by reversed-phase liquid chromatography.
常规液相合成法一般为均相反应，反应性良好，中间体可通过洗涤、结晶等手段进行纯化，最终产物纯度较高。但每个反应的产物和杂质性质差异较大，无法找到普适的方法进行分离纯化，开发周期长，制造步骤复杂。The conventional liquid phase synthesis method is generally a homogeneous reaction with good reactivity. The intermediate can be purified by washing, crystallization and other means, and the final product has a high purity. However, the properties of the products and impurities of each reaction are quite different, and no universal method can be found for separation and purification, the development cycle is long, and the manufacturing steps are complicated.
液相载体合成法是近十年来为了改善上述两种方法而开发出来的一种新的肽合成方法。其中，较为有代表性的包括JITSUBO公司的Molecular Hiving TM技术和味之素的Ajiphase技术。 The liquid carrier synthesis method is a new peptide synthesis method developed in the past ten years to improve the above two methods. Among them, the more representative ones include JITSUBO's Molecular Hiving TM technology and Ajinomoto's Ajiphase technology.
JITSUBO公司的Molecular Hiving TM技术使用长链烷氧基苄醇，如：3，5-二(二十二烷基氧基)苄醇、2，4--二(二十二烷基氧基)苄醇、3，4，5—三(十八烷基氧基)苄醇等，作为液相载体。使用这些载体作为C端保护试剂进行肽的合成，反应在均相中进行，通过改变溶剂组成或温度变化等使产物析出，通过沉析—过滤—洗涤步骤实现分离纯化。但该方法存在以下问题：①载体在中低极性的乙酸酯和甲苯等溶剂中的溶解度不大，反应浓度较低，不利于放大生产；②要通过定量蒸馏等操作才可以完成改变溶剂组成的工序，操作复杂；③改变溶剂体系析出的一般为非晶体状固体，过滤—洗涤十分困难，杂质除去效果差，耗时长。因此，不能说该方法是一种重复性好的普适型肽合成方法。 JITSUBO’s Molecular Hiving TM technology uses long-chain alkoxy benzyl alcohols, such as 3,5-bis(docosyloxy)benzyl alcohol, 2,4-bis(docosyloxy) Benzyl alcohol, 3,4,5-tris(octadecyloxy)benzyl alcohol, etc. are used as liquid carrier. These carriers are used as C-terminal protective reagents for peptide synthesis. The reaction is carried out in a homogeneous phase. The product is precipitated by changing the solvent composition or temperature change, and separation and purification are achieved through the steps of precipitation-filtration-washing. However, this method has the following problems: ①The solubility of the carrier in solvents such as medium and low polarity acetate and toluene is not large, and the reaction concentration is low, which is not conducive to scale-up production; ②The solvent can be changed through quantitative distillation and other operations. The composition process is complicated to operate; ③The precipitation of the solvent system is generally amorphous solid, which is very difficult to filter and wash, and the effect of removing impurities is poor and time-consuming. Therefore, it cannot be said that this method is a universal peptide synthesis method with good reproducibility.
味之素的Ajiphase技术通过对前者进行改造，使用了多支链烷基代替直链烷基，有效改善了载体在醋酸异丙酯等溶剂中的溶解度，反应可以在较高浓度进行，通过使用水&有机极性溶剂洗涤即可实现分离纯化。但该技术还存在以下问题：①反应时间长，公开号为 CN107011132A的专利中使用醋酸异丙酯或甲基环戊基醚作为溶剂缩合需要反应过夜，脱保护需要4～6小时，其它公开的文献资料中使用氯仿作为溶剂的缩合反应也需要3小时以上，反应时间长导致副产物杂质增多；②由于液液分离时两相极性相差较小，杂质难以除去，产物纯度低；③使用此类型载体合成肽的过程中，溶解度下降较快，一般的肽链氨基酸数大于4时即出现凝胶化现象，因此，该方法也很难说是一种重复性好的普适型肽合成方法。Ajinomoto's Ajiphase technology has modified the former, using multi-branched alkyl groups instead of linear alkyl groups, effectively improving the solubility of the carrier in solvents such as isopropyl acetate. The reaction can be carried out at a higher concentration. Water & organic polar solvent washing can achieve separation and purification. However, this technology still has the following problems: ①The reaction time is long. The patent publication No. CN107011132A uses isopropyl acetate or methylcyclopentyl ether as a solvent for condensation to react overnight, and deprotection requires 4-6 hours. Other publications The condensation reaction using chloroform as a solvent in the literature also requires more than 3 hours, and the long reaction time leads to an increase in by-product impurities; ②Due to the small difference in polarity between the two phases during liquid-liquid separation, the impurities are difficult to remove and the product purity is low; ③Use this In the process of peptide synthesis by the type carrier, the solubility decreases rapidly, and gelation occurs when the number of amino acids in the general peptide chain is greater than 4. Therefore, this method can hardly be said to be a universal peptide synthesis method with good repeatability.
因此，合成一种具有普适型的液相合成保护载体化合物，进而得到一种重复性好的普适型肽合成方法具有重要的意义。Therefore, it is of great significance to synthesize a universal type of liquid phase synthetic protective carrier compound, and then obtain a universal peptide synthesis method with good reproducibility.
发明内容Summary of the invention
本发明的目的在于提供一种含有芴环结构的化合物及其应用。使用该化合物作为肽液相合成的保护载体，在均相或非均相的溶剂体系，特别在非均相溶剂体系中，能提高反应速度和试剂利用率、简化后处理操作、提高产品纯度、提高操作流程的通用性，可以发展成一种普适型的生产方法。The purpose of the present invention is to provide a compound containing a fluorene ring structure and its application. Using this compound as a protective carrier for peptide liquid phase synthesis, in a homogeneous or heterogeneous solvent system, especially in a heterogeneous solvent system, it can increase the reaction speed and reagent utilization, simplify post-processing operations, and improve product purity. Improving the versatility of the operating process can be developed into a universal production method.
本发明所采取的技术方案是：The technical scheme adopted by the present invention is:
本发明的目的之一在于提供一种含有芴环结构的化合物，该化合物的结构如通式(1)所示：One of the objectives of the present invention is to provide a compound containing a fluorene ring structure, the structure of which is shown in the general formula (1):
Figure PCTCN2019125143-appb-000001
Figure PCTCN2019125143-appb-000001
其中：among them:
X选自OH、卤素、磺酸酯、NHR a； X is selected from OH, halogen, sulfonate, NHR a ;
Y选自氢、芳基；Y is selected from hydrogen and aryl;
其中，R a选自氢、烷基或芳烷基； Wherein, R a is selected from hydrogen, alkyl or aralkyl;
Q独立选自O、NH、NHCO、CO、CONH、S、SO或SO 2； Q is independently selected from O, NH, NHCO, CO, CONH, S, SO or SO 2 ;
n 1，n 2独立选自0～3的整数，且n 1、n 2不同时为0； n 1 and n 2 are independently selected from an integer of 0 to 3, and n 1 and n 2 are not 0 at the same time;
R独立选自通式(2)所示的基团：R is independently selected from the group represented by the general formula (2):
Figure PCTCN2019125143-appb-000002
Figure PCTCN2019125143-appb-000002
其中，*表示与Q连接；Among them, * means connected with Q;
R 1选自C 1～C 25的烷基或通式(3)所示的基团： R 1 is selected from a C 1 ～C 25 alkyl group or a group represented by the general formula (3):
Figure PCTCN2019125143-appb-000003
Figure PCTCN2019125143-appb-000003
其中，*表示与羰基连接；Wherein, * means it is connected to the carbonyl group;
m1表示1～3的整数；m1 represents an integer of 1 to 3;
R 1a选自C 6～C 25的烷基，且m1个R 1a的总碳数不少于8； R 1a is selected from C 6 -C 25 alkyl groups, and the total carbon number of m1 R 1a is not less than 8;
k a选自0～3的整数； k a is selected from an integer of 0 to 3;
环B除了具有m1个R 1aO取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基、不含卤素原子取代的C 1～C 5烷氧基的取代基； Ring B except that one R 1a O having m1 substituent groups, but may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ~ C 5 alkoxy group, a halogen atom free of C 1 ~ C 5 alkoxy group substituent;
通式(2)中，R 2选自氢、C 1～C 25的烷基或通式(4)所示的基团： In the general formula (2), R 2 is selected from hydrogen, a C 1 to C 25 alkyl group or the group represented by the general formula (4):
Figure PCTCN2019125143-appb-000004
Figure PCTCN2019125143-appb-000004
通式(4)中，*表示与N连接；In the general formula (4), * means connected with N;
m2选自0～3的整数；m2 is selected from an integer of 0 to 3;
R 2a选自C 6～C 25的烷基； R 2a is selected from C 6 ～C 25 alkyl groups;
k b选自1～6的整数； k b is selected from an integer of 1 to 6;
环C除了具有m2个R 2aO取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基、不含卤素原子取代的C 1～C 5烷氧基的取代基； In addition to an outer ring C m2 having a substituent group R 2a O, may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ~ C 5 alkoxy group, a halogen atom free of C 1 ~ C 5 alkoxy group substituent;
L选自含有O、N或S杂原子的C 2～C 15有机链基或不含有O、N或S杂原子的C 2～C 15有机链基，当L选自不含有O、N或S杂原子的C 2～C 15有机链基时，R 2≠H； L is selected from C 2 ～C 15 organic chain groups containing O, N or S heteroatoms or C 2 ～C 15 organic chain groups without O, N or S heteroatoms, when L is selected from C 2 ～C 15 organic chain groups that do not contain O, N or When the C 2 ～C 15 organic chain group of S heteroatom, R 2 ≠H;
芴环除了具有n 1和n 2个RQ取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基或不含卤素原子取代的C 1～C 5烷氧基的取代基。 In addition to having a fluorene ring n 1 and n 2 th RQ substituents, they can also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, substituent group substituted with halogen atom-containing C 1 ~ C 5 alkoxy group or a halogen atom free of C 1 ~ C 5 alkoxy.
优选地，上述R选自通式(5)所示的基团：Preferably, the aforementioned R is selected from the group represented by the general formula (5):
Figure PCTCN2019125143-appb-000005
Figure PCTCN2019125143-appb-000005
其中，通式(5)中，*表示与Q连接；Among them, in the general formula (5), * means connected with Q;
k 1选自0～3的整数，并且当k 1＝0时，R 2≠H，更优选地，k 1选自1～3的整数。优选地，上述R选自通式(6)所示的基团： k 1 is selected from an integer of 0 to 3, and when k 1 =0, R 2 ≠H, more preferably, k 1 is selected from an integer of 1 to 3. Preferably, the aforementioned R is selected from the group represented by the general formula (6):
Figure PCTCN2019125143-appb-000006
Figure PCTCN2019125143-appb-000006
其中，通式(6)中，*表示与Q连接；Among them, in the general formula (6), * means connected with Q;
R 4选自氢、C 1～C 25的烷基或上述通式(4)所示的基团； R 4 is selected from hydrogen, a C 1 to C 25 alkyl group or the group represented by the above general formula (4);
k 2选自1～4的整数； k 2 is selected from an integer of 1 to 4;
k 3选自1～4的整数。 k 3 is selected from an integer of 1-4.
优选地，上述R选自通式(7)所示的基团：Preferably, the above R is selected from the group represented by the general formula (7):
Figure PCTCN2019125143-appb-000007
Figure PCTCN2019125143-appb-000007
其中，通式(7)中，*表示与Q连接；Among them, in the general formula (7), * means connected with Q;
R 6选自氢、C 1～C 25的烷基或上述通式(4)所示的基团； R 6 is selected from hydrogen, C 1 to C 25 alkyl group or the group represented by the above general formula (4);
k 4选自0～3的整数； k 4 is selected from an integer from 0 to 3;
k 5选自0～3的整数；优选地，k 5选自1～3的整数； k 5 is selected from an integer of 0 to 3; preferably, k 5 is selected from an integer of 1 to 3;
R 5选自氢、天然氨基酸的侧链基、烷基或通式(8)所示的基团： R 5 is selected from hydrogen, a side chain group of a natural amino acid, an alkyl group or a group represented by the general formula (8):
Figure PCTCN2019125143-appb-000008
Figure PCTCN2019125143-appb-000008
其中，*表示连接位点；Among them, * represents the connection site;
k 6选自1～4的整数； k 6 is selected from an integer of 1 to 4;
R 1＇选自C 1～C 25的烷基或通式(3)所示的基团； R 1 ′ is selected from a C 1 ～C 25 alkyl group or a group represented by the general formula (3);
R 2＇选自氢、C 1～C 25的烷基或通式(4)所示的基团。 R 2 'is selected from hydrogen, C 1 to C 25 alkyl group, or the group represented by the general formula (4).
优选地，上述Q选自O。Preferably, the above Q is selected from O.
优选地，上述R a选自氢、甲基、乙基、丙基、苄基或甲氧苄基。 Preferably, the R a is selected from hydrogen, methyl, ethyl, propyl, benzyl or methoxybenzyl.
优选地，所述n 1+n 2等于1～3。 Preferably, the n 1 +n 2 is equal to 1-3.
优选地，上述m1选自2或3，m1个R 1a的总碳数为12～60。 Preferably, m1 is selected from 2 or 3, and the total carbon number of m1 R 1a is 12-60.
优选地，上述R 1a选自C 8～C 22的烷基。 Preferably, the aforementioned R 1a is selected from C 8 to C 22 alkyl groups.
优选地，上述m2选自2或3，m2个R 2a的总碳数为12～60。 Preferably, m2 is selected from 2 or 3, and the total carbon number of m2 R 2a is 12-60.
优选地，上述R 2a选自C 8～C 22的烷基。 Preferably, the above-mentioned R 2a is selected from C 8 to C 22 alkyl groups.
优选地，上述R 2选自氢、甲基、乙基、丙基、异丙基、异辛基、苄基或4-甲氧苄基。 Preferably, the above-mentioned R 2 is selected from hydrogen, methyl, ethyl, propyl, isopropyl, isooctyl, benzyl or 4-methoxybenzyl.
优选地，通式(1)中的X中的卤素选自Cl、Br或I。Preferably, the halogen in X in the general formula (1) is selected from Cl, Br or I.
优选地，通式(1)中的X中的磺酸酯选自甲磺酸酯或对甲苯磺酸酯。Preferably, the sulfonate in X in the general formula (1) is selected from methanesulfonate or p-toluenesulfonate.
优选地，上述含有芴环结构的化合物的结构式选自如下：Preferably, the structural formula of the compound containing a fluorene ring structure is selected from the following:
Figure PCTCN2019125143-appb-000009
Figure PCTCN2019125143-appb-000009
Figure PCTCN2019125143-appb-000010
Figure PCTCN2019125143-appb-000010
Figure PCTCN2019125143-appb-000011
Figure PCTCN2019125143-appb-000011
Figure PCTCN2019125143-appb-000012
Figure PCTCN2019125143-appb-000012
优选地，上述结构式中的C 8H 17烷基为异辛基，C 9H 19烷基为异壬基，C 10H 21烷基为异构十烷基，C 13H 27烷基为异构十三烷基，C 20H 41烷基为2,3-二氢植烷基。 Preferably, the C 8 H 17 alkyl group in the above structural formula is isooctyl, the C 9 H 19 alkyl group is isononyl, the C 10 H 21 alkyl group is isodecyl, and the C 13 H 27 alkyl group is iso Structure tridecyl, C 20 H 41 alkyl is 2,3-dihydrophytyl.
优选地，上述含有芴环结构的化合物易溶于烃类有机溶剂、芳香烃类有机溶剂、酯类有机溶剂、醚类有机溶剂、水溶性非质子类极性有机溶剂中的至少一种。Preferably, the compound containing the fluorene ring structure is easily soluble in at least one of hydrocarbon organic solvents, aromatic hydrocarbon organic solvents, ester organic solvents, ether organic solvents, and water-soluble aprotic polar organic solvents.
优选地，上述烃类有机溶剂选自庚烷、己烷、石油醚、环己烷、甲基环己烷中的至少一种。Preferably, the above-mentioned hydrocarbon organic solvent is selected from at least one of heptane, hexane, petroleum ether, cyclohexane, and methylcyclohexane.
优选地，上述芳香烃类有机溶剂选自甲苯、乙苯、二甲苯中的至少一种。Preferably, the above-mentioned aromatic hydrocarbon organic solvent is selected from at least one of toluene, ethylbenzene, and xylene.
优选地，上述酯类有机溶剂选自乙酸异丙酯、乙酸叔丁酯、乙酸乙酯中的至少一种。Preferably, the above-mentioned ester organic solvent is selected from at least one of isopropyl acetate, tert-butyl acetate, and ethyl acetate.
优选地，上述醚类有机溶剂选自***、异丙醚、甲基叔丁基醚、甲基环戊基醚、四氢呋喃中的至少一种。Preferably, the ether organic solvent is at least one selected from the group consisting of diethyl ether, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, and tetrahydrofuran.
优选地，上述水溶性非质子类极性有机溶剂选自N,N-二甲基甲酰胺、N,N-二乙基甲酰胺、N,N-二甲基乙酰胺、N-甲基-吡咯烷酮、N-乙基-吡咯烷酮、二甲基亚砜、环丁砜、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种。Preferably, the water-soluble aprotic polar organic solvent is selected from the group consisting of N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, and N-methyl- Pyrrolidone, N-ethyl-pyrrolidone, dimethyl sulfoxide, sulfolane, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidine At least one of ketones.
优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞1％。Preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >1% at 25-30°C.
优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞5％。Preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >5% at 25-30°C.
优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞30％。Preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >30% at 25-30°C.
优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度 ＞50％。Preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >50% at 25-30°C.
优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞80％。Preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >80% at 25-30°C.
更优选地，在25～30℃时，上述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞100％。More preferably, the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >100% at 25-30°C.
优选地，在25～30℃时，上述含有芴环结构的化合物在烃类有机溶剂中的溶解度＞30％；更优选地，上述含有芴环结构的化合物在庚烷中的溶解度＞30％。Preferably, the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 30% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 30%.
优选地，在25～30℃时，上述含有芴环结构的化合物在烃类有机溶剂中的溶解度＞50％；更优选地，上述含有芴环结构的化合物在庚烷中的溶解度＞50％。Preferably, the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is >50% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is >50%.
优选地，在25～30℃时，上述含有芴环结构的化合物在烃类有机溶剂中的溶解度＞60％；更优选地，上述含有芴环结构的化合物在庚烷中的溶解度＞60％。Preferably, the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is greater than 60% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is greater than 60%.
优选地，在25～30℃时，上述含有芴环结构的化合物在烃类有机溶剂中的溶解度＞80％；更优选地，上述含有芴环结构的化合物在庚烷中的溶解度＞80％。Preferably, the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 80% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 80%.
优选地，在25～30℃时，上述含有芴环结构的化合物在烃类有机溶剂中的溶解度＞100％；更优选地，上述含有芴环结构的化合物在庚烷中的溶解度＞100％。Preferably, the solubility of the compound containing the fluorene ring structure in a hydrocarbon organic solvent is> 100% at 25-30° C.; more preferably, the solubility of the compound containing the fluorene ring structure in heptane is> 100%.
优选地，上述含有芴环结构的化合物在酯类有机溶剂中的溶解度＞50％；更优选地，上述含有芴环结构的化合物在乙酸异丙酯中的溶解度＞50％。Preferably, the solubility of the compound containing the fluorene ring structure in the ester organic solvent is >50%; more preferably, the solubility of the compound containing the fluorene ring structure in the isopropyl acetate is >50%.
优选地，在25～30℃时，上述含有芴环结构的化合物在酯类有机溶剂中的溶解度＞100％；更优选地，上述含有芴环结构的化合物在乙酸异丙酯中的溶解度＞100％。Preferably, the solubility of the compound containing the fluorene ring structure in the ester organic solvent is> 100% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the isopropyl acetate is> 100 %.
优选地，在25～30℃时，上述含有芴环结构的化合物在醚类有机溶剂中的溶解度＞30％；更优选地，上述含有芴环结构的化合物在甲基叔丁基醚中的溶解度＞30％。Preferably, the solubility of the compound containing the fluorene ring structure in the ether organic solvent is greater than 30% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether ＞30%.
优选地，在25～30℃时，上述含有芴环结构的化合物在醚类有机溶剂中的溶解度＞50％；更优选地，上述含有芴环结构的化合物在甲基叔丁基醚中的溶解度＞50％。Preferably, the solubility of the compound containing the fluorene ring structure in ether organic solvents is >50% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in methyl tert-butyl ether >50%.
优选地，在25～30℃时，上述含有芴环结构的化合物在醚类有机溶剂中的溶解度＞60％；更优选地，上述含有芴环结构的化合物在甲基叔丁基醚中的溶解度＞60％。Preferably, the solubility of the compound containing the fluorene ring structure in the ether organic solvent is greater than 60% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether ＞60%.
优选地，在25～30℃时，上述含有芴环结构的化合物在醚类有机溶剂中的溶解度＞80％；更优选地，上述含有芴环结构的化合物在甲基叔丁基醚中的溶解度＞80％。Preferably, the solubility of the compound containing the fluorene ring structure in the ether organic solvent is> 80% at 25-30°C; more preferably, the solubility of the compound containing the fluorene ring structure in the methyl tert-butyl ether >80%.
更优选地，在25～30℃时，上述含有芴环结构的化合物在醚类有机溶剂中的溶解度＞100％；更优选地，上述含有芴环结构的化合物在甲基叔丁基醚中的溶解度＞100％。More preferably, the solubility of the compound containing the fluorene ring structure in the ether organic solvent is> 100% at 25-30°C; more preferably, the compound containing the fluorene ring structure is more than 100% in methyl tert-butyl ether. Solubility>100%.
本发明的上述含有芴环结构的化合物可以通过以下合成路线制备得到：The above-mentioned compound containing fluorene ring structure of the present invention can be prepared by the following synthetic route:
Figure PCTCN2019125143-appb-000013
Figure PCTCN2019125143-appb-000013
芴酮1-1a经过步骤1-1→步骤1-2，或经过步骤1-3→步骤1-4→步骤1-5合成X＝NHRa的化合物；Fluorenone 1-1a goes through Step 1-1→Step 1-2, or Step 1-3→Step 1-4→Step 1-5 to synthesize X=NHRa compound;
芴酮1-1a经过步骤1-6→步骤1-7合成X＝OH的化合物；Fluorenone 1-1a undergoes step 1-6→step 1-7 to synthesize X=OH compound;
化合物1-7a经过步骤1-8可将OH可经过常规反应可转化为卤素、磺酸酯等化合物。Compound 1-7a can convert OH into halogen, sulfonate and other compounds through conventional reaction through steps 1-8.
上述卤素选自Cl、Br、I；上述磺酸酯选自甲磺酸酯或对甲苯磺酸酯。The above halogen is selected from Cl, Br, and I; the above sulfonate is selected from methanesulfonate or p-toluenesulfonate.
Figure PCTCN2019125143-appb-000014
Figure PCTCN2019125143-appb-000014
按上述反应路线可合成如通式(5)、(6)和(7)中的R。According to the above reaction route, R in general formulas (5), (6) and (7) can be synthesized.
本发明的另一目的在于提供一种氨基酸或肽C端保护试剂，该保护试剂包含上述的含有芴环结构的化合物。Another object of the present invention is to provide an amino acid or peptide C-terminal protection reagent, which comprises the above-mentioned compound containing a fluorene ring structure.
本发明还提供了上述含有芴环结构的化合物在均相或非均相溶剂体系中合成肽试剂中的应用。The present invention also provides the application of the compound containing the fluorene ring structure in the synthesis of peptide reagents in a homogeneous or heterogeneous solvent system.
另外，本领域的技术人员可以依据常规的选择在上述试剂中添加其他物质及依据需要制备成所需的化合物或复合物。In addition, those skilled in the art can add other substances to the above reagents according to conventional selection and prepare the required compounds or complexes according to needs.
本发明同时还提供了一种肽的合成方法，该方法包括使用上述含有芴环结构的化合物。The present invention also provides a method for synthesizing peptides, which method includes using the above-mentioned compound containing a fluorene ring structure.
优选地，上述肽合成的方法，包括如下步骤：Preferably, the above-mentioned method of peptide synthesis includes the following steps:
1)载体接入：将上述含有芴环结构的化合物作为载体与N-保护氨基酸或N-保护肽化合物通过常规反应进行连接得到含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物；1) Carrier access: The above-mentioned compound containing fluorene ring structure is used as a carrier to connect with N-protected amino acid or N-protected peptide compound through conventional reaction to obtain N-protected amino acid or N-protected C-terminal of the carrier containing fluorene ring structure Peptide compound
2)N端去保护：将含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶解于溶剂中，加入N端保护的脱保护试剂溶液形成均相或非均相体系进行N端脱保护，加入高极性溶剂进行萃取，得含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液；2) N-terminal deprotection: Dissolve the N-protected amino acid or N-protected peptide compound containing the C-terminal protection of the fluorene ring structure carrier in a solvent, and add the N-terminal protection deprotection reagent solution to form a homogeneous or heterogeneous system. N-terminal deprotection, adding a highly polar solvent for extraction, to obtain N-deprotected amino acid or N-deprotected peptide compound solution containing fluorene ring structure carrier C-terminal protection;
3)肽链延长：在含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶 液中，加入N-保护氨基酸或N-保护肽溶液，后加入缩合试剂溶液形成均相或非均相体系进行缩合反应，加入高极性溶剂进行萃取，得到含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶液；3) Peptide chain extension: Add N-protected amino acid or N-protected peptide solution to the C-terminal protected N-deprotected amino acid or N-deprotected peptide compound solution of the fluorene ring structure carrier, and then add the condensation reagent solution to form a homogeneous solution. Condensation reaction in a phase or heterogeneous system, and a highly polar solvent is added for extraction to obtain an N-protected amino acid or N-protected peptide compound solution containing the C-terminal protection of the fluorene ring structure carrier;
4)重复步骤2)和步骤3)接入下一个氨基酸，直至得到完整的肽链。4) Repeat step 2) and step 3) to insert the next amino acid until a complete peptide chain is obtained.
优选地，步骤2)中的含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶液与步骤2)中的N端保护的脱保护试剂溶液成非均相体系。Preferably, the solution containing the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier in step 2) and the N-terminal protected deprotection reagent solution in step 2) form a heterogeneous system.
优选地，步骤3)中的含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶液与步骤3)中的N-保护氨基酸或N-保护肽、缩合试剂溶液成非均相体系。Preferably, the solution containing the N-deprotected amino acid or N-deprotected peptide compound protected by the C-terminal of the fluorene ring structure carrier in step 3) and the N-protected amino acid or N-protected peptide and condensation reagent solution in step 3) Into a heterogeneous system.
优选地，步骤2)中溶解含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶剂和步骤3)中含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶液中的溶剂独立选自烃类或烃类与酯类、醚类、卤代烃中的至少一种形成的混合溶剂。Preferably, in step 2) the solvent containing the N-protected amino acid or N-protected peptide compound protected at the C-terminal of the fluorene ring structure carrier and the N-deprotected amino acid or N-protected amino acid or N-protected at the C terminal of the fluorene ring structure carrier in the step 3) -The solvent in the solution of the deprotected peptide compound is independently selected from hydrocarbons or mixed solvents formed by hydrocarbons and at least one of esters, ethers, and halogenated hydrocarbons.
需要说明的是烃类与酯类、醚类、卤代烃中的至少一种形成的混合溶剂中的烃类的含量以不影响形成非均相体系为准。It should be noted that the content of hydrocarbons in the mixed solvent formed by at least one of hydrocarbons and esters, ethers, and halogenated hydrocarbons shall not affect the formation of a heterogeneous system.
优选地，步骤2)溶解含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶剂和步骤3)中含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶液中的溶剂独立选自己烷、庚烷、环己烷、甲基环己烷、石油醚或己烷、庚烷、环己烷、甲基环己烷、石油醚中至少一种与醋酸异丙酯、乙酸叔丁酯、乙酸乙酯、***、异丙醚、甲基叔丁醚、甲基环戊基醚、二氯甲烷、氯仿中的至少一种形成的混合溶剂中的至少一种。Preferably, step 2) dissolve the solvent containing the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier and step 3) the N-deprotected amino acid or N-protected amino acid or N-protected C terminal of the fluorene ring structure carrier. The solvent in the solution of the deprotected peptide compound is independently selected from at least hexane, heptane, cyclohexane, methylcyclohexane, petroleum ether or hexane, heptane, cyclohexane, methylcyclohexane, and petroleum ether. A mixed solvent formed with at least one of isopropyl acetate, tert-butyl acetate, ethyl acetate, diethyl ether, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, dichloromethane, and chloroform At least one of them.
优选地，步骤2)中溶解N端保护的脱保护试剂的溶剂、步骤3)中溶解N-保护氨基酸或N-保护肽、缩合试剂的溶剂独立选自酰胺类溶剂。Preferably, the solvent for dissolving the N-terminal protected deprotection reagent in step 2), the solvent for dissolving the N-protected amino acid or N-protected peptide, and the condensation reagent in step 3) are independently selected from amide solvents.
优选地，上述酰胺类溶剂选自N,N-二甲基甲酰胺、N,N-二乙基甲酰胺、N,N-二甲基乙酰胺、N-甲基-吡咯烷酮、N-乙基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种。Preferably, the above-mentioned amide solvent is selected from N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, N-methyl-pyrrolidone, N-ethyl -At least one of pyrrolidone, 1,3-dimethylimidazolinone, and 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone.
优选地，步骤2)和步骤3)中的高极性溶剂选自水、醇类、腈类、酰胺类、亚砜类、砜类、水溶性醇醚类中的至少一种。Preferably, the highly polar solvent in step 2) and step 3) is selected from at least one of water, alcohols, nitriles, amides, sulfoxides, sulfones, and water-soluble alcohol ethers.
优选地，步骤2)和步骤3)中的高极性溶剂选自水、甲醇、乙腈、N,N-二甲基甲酰胺、N-甲基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮、二甲基亚砜中、环丁砜中的至少一种。Preferably, the highly polar solvent in step 2) and step 3) is selected from water, methanol, acetonitrile, N,N-dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazoline At least one of ketone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone, dimethyl sulfoxide, and sulfolane.
需要说明的是，步骤2)中，当将含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶解于溶剂中，加入N端保护的脱保护试剂溶液形成均相体系进行N端脱保护时， 溶解含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶剂不含有烃类或含有部分烃类，但该部分烃类不影响形成均相体系，溶解N端保护的脱保护试剂的溶剂与溶解芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶剂可以为酯类溶剂、醚类溶剂、卤代烃类或酰胺类溶剂中的至少一种；优选地，醋酸乙酯，醋酸异丙酸、醋酸叔丁酯、异丙醚、甲基叔丁醚、甲基环戊基醚、四氢呋喃、二氯甲烷、氯仿、N,N-二甲基甲酰胺、N-甲基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种；同样的步骤3)中，在含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶液中，加入N-保护氨基酸或N-保护肽溶液，后加入缩合试剂溶液形成均相体系进行缩合反应时，含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶液的溶剂不含有烃类或含有部分烃类，但该部分烃类不影响形成均相体系，溶解N-保护氨基酸或N-保护肽和缩合试剂的溶剂与溶解含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物的溶剂相可以选自酯类溶剂、醚类溶剂或酰胺类溶剂中的至少一种，优选地，醋酸乙酯，醋酸异丙酯、醋酸叔丁酯、二氯甲烷、氯仿、四氢呋喃、N,N-二甲基甲酰胺、N-甲基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种。It should be noted that in step 2), when the N-protected amino acid or N-protected peptide compound containing the C-terminal protection of the fluorene ring structure carrier is dissolved in a solvent, the N-terminal protection deprotection reagent solution is added to form a homogeneous system. When the N-terminal is deprotected, the solvent containing the N-protected amino acid or N-protected peptide compound protected by the C-terminal of the fluorene ring structure carrier does not contain hydrocarbons or contains some hydrocarbons, but this part of the hydrocarbons does not affect the formation of a homogeneous system. The solvent for dissolving the N-terminal protected deprotection reagent and the solvent for dissolving the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier can be ester solvents, ether solvents, halogenated hydrocarbons or amide solvents At least one of; preferably, ethyl acetate, isopropyl acetate, tert-butyl acetate, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, tetrahydrofuran, dichloromethane, chloroform, N, N-dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone In the same step 3), add the N-protected amino acid or N-protected peptide solution to the solution containing the N-deprotected amino acid or N-deprotected peptide compound protected by the C-terminal of the fluorene ring structure carrier, After adding the condensation reagent solution to form a homogeneous system for the condensation reaction, the solvent of the solution containing the N-deprotected amino acid or N-deprotected peptide compound protected by the C-terminal of the fluorene ring structure carrier does not contain hydrocarbons or contains part of hydrocarbons, but This part of the hydrocarbon does not affect the formation of a homogeneous system. The solvent for dissolving N-protected amino acids or N-protected peptides and condensation reagents and dissolving the N-deprotected amino acids or N-deprotected peptide compounds protected by the C-terminal of the fluorene ring structure carrier The solvent phase can be selected from at least one of ester solvents, ether solvents or amide solvents, preferably ethyl acetate, isopropyl acetate, tert-butyl acetate, dichloromethane, chloroform, tetrahydrofuran, N, N -Dimethylformamide, N-methyl-pyrrolidone, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone At least one.
本发明的肽合成方法中，仅通过加入高极性溶剂就可以将目标产物和杂质进行很好的分离，特别是在非均相体系中，从而大大简化了后处理的操作步骤。In the peptide synthesis method of the present invention, the target product and impurities can be well separated only by adding a highly polar solvent, especially in a heterogeneous system, thereby greatly simplifying the post-processing operation steps.
优选地，上述N-保护氨基酸或N-保护肽的用量为上述含有芴环结构的化合物的0.8～3.0当量，上述缩合试剂的用量为上述含有芴环结构的化合物的0.8～3.0当量。Preferably, the amount of the N-protected amino acid or N-protected peptide is 0.8-3.0 equivalents of the compound containing the fluorene ring structure, and the amount of the condensation reagent is 0.8-3.0 equivalents of the compound containing the fluorene ring structure.
优选地，上述N-保护氨基酸或N-保护肽的用量为上述含有芴环结构的化合物的1～1.1当量，上述缩合试剂的用量为上述含有芴环结构的化合物的1～1.2当量。Preferably, the amount of the N-protected amino acid or N-protected peptide is 1 to 1.1 equivalents of the compound containing the fluorene ring structure, and the amount of the condensation reagent is 1 to 1.2 equivalents of the compound containing the fluorene ring structure.
另外，在肽合成反应中，将本发明的化合物导入氨基酸或肽，作为C端的保护基，在导入本发明的化合物过程中，可以将X＝OH的化合物先转化成等价的活性物，如卤代物、磺酸酯，再与氨基酸或肽反应，也可以直接酯化或酰胺化；导入X＝NHR a的化合物通过常规的缩合方法与氨基酸和肽反应即可完成，操作步骤可与步骤3)相同或不同。反应可在均相和非均相溶剂体系进行，其中X＝OH的载体接入反应优选在均相中进行，X＝NHRa的载体接入反应优选在非均相溶剂中进行。 In addition, in the peptide synthesis reaction, the compound of the present invention is introduced into an amino acid or peptide as the C-terminal protective group. In the process of introducing the compound of the present invention, the compound with X=OH can be first converted into an equivalent active substance, such as Halogenated compounds, sulfonate esters, and then reacted with amino acids or peptides, can also be directly esterified or amidated; the compound introduced into X=NHR a can be completed by reacting with amino acids and peptides by conventional condensation methods. The operation steps can be the same as step 3. ) Same or different. The reaction can be carried out in homogeneous and heterogeneous solvent systems, wherein the carrier incorporation reaction of X=OH is preferably carried out in a homogeneous phase, and the carrier incorporation reaction of X=NHRa is preferably carried out in a heterogeneous solvent.
若需要，本领域的技术人员可以依据常规的方法对肽链进行后续C端修饰或N端修饰、或通过选择性侧链去保护再进行如环合或引入其他基团的反应。If necessary, those skilled in the art can perform subsequent C-terminal modification or N-terminal modification on the peptide chain according to conventional methods, or through selective side chain deprotection, and then perform reactions such as cyclization or introduction of other groups.
本发明的有益效果是：The beneficial effects of the present invention are:
1、本发明提供了一种能应用于肽液相合成的保护载体，使用这种保护载体的肽合成反应， 在合适的溶剂体系中，反应速度快，试剂利用率高；同时，通过简单的液液萃取分离即能够进行有效纯化，最终可获得纯度较高的产品，因而后处理操作简单；并且，合成过程中溶解度变化小，操作流程通用性强，可以发展成一种普适型的生产方法。1. The present invention provides a protective carrier that can be applied to peptide liquid phase synthesis. The peptide synthesis reaction using this protective carrier has fast reaction speed and high reagent utilization in a suitable solvent system; at the same time, simple Liquid-liquid extraction separation can be effectively purified, and finally a product with higher purity can be obtained, so the post-processing operation is simple; and the solubility change during the synthesis process is small, the operation flow is strong, and it can be developed into a universal production method .
2、本发明的含有芴环结构的化合物含有非极性和极性两部分结构，使其既能溶于非极性溶剂，又溶于极性溶剂，因此适合在非极性溶剂与极性溶剂组成的均相或非均相混合溶剂体系中，特别是烃类或烃类与醚类、酯类中的至少一种溶剂形成的混合溶剂与酰胺类极性溶剂组成的非均相体系，作为保护载体进行肽的合成反应；并且以此类载体进行的肽合成反应具有：①反应速度快，副产物少；②反应试剂利用率高，试剂用量少，成本低，三废少；③后处理萃取分层杂质和产物的分离效果好产物纯度高；④合成过程中中间体化合物的溶解度和反应速率变化小，氨基酸或肽片断接入重复性好，操作简便，普适性强，适合作为通用型生产方法的优势。2. The fluorene ring structure-containing compound of the present invention contains a non-polar and polar two-part structure, so that it can be dissolved in both non-polar solvents and polar solvents, so it is suitable for non-polar solvents and polar solvents. In a homogeneous or heterogeneous mixed solvent system composed of solvents, particularly a heterogeneous system composed of a mixed solvent of hydrocarbons or hydrocarbons and at least one of ethers and esters and amide polar solvents, As a protective carrier for peptide synthesis reaction; and the peptide synthesis reaction carried out with this kind of carrier has: ① fast reaction speed, few by-products; ② high utilization of reagents, low reagent dosage, low cost, and less three wastes; ③after It has good separation effect for processing the extraction stratified impurities and products. The product has high purity; ④The solubility and reaction rate of the intermediate compounds in the synthesis process are small, the amino acid or peptide fragments have good repeatability, simple operation, strong universality, and suitable as Advantages of universal production methods.
具体实施方式detailed description
下面进一步列举实施例以详细说明本发明。同样应理解，以下实施例只用于对本发明进行进一步说明，不能理解为对本发明保护范围的限制，本领域技术人员根据本发明阐述的原理做出的一些非本质的改进和调整均属于本发明的保护范围。下述示例具体的工艺参数等也仅是合适范围中的一个示例，即本领域技术人员可以通过本文的说明做合适范围内的选择，而并非要限定于下文示例的具体数据。Examples are further listed below to illustrate the present invention in detail. It should also be understood that the following examples are only used to further illustrate the present invention, and cannot be construed as limiting the protection scope of the present invention. Some non-essential improvements and adjustments made by those skilled in the art based on the principles set forth in the present invention belong to the present invention. The scope of protection. The specific process parameters in the following examples are only an example of the appropriate range, that is, those skilled in the art can make selections within the appropriate range through the description herein, and are not limited to the specific data illustrated below.
本说明书中和以下的实施例中，下述缩写所代表的物质为：In this specification and in the following examples, the substances represented by the following abbreviations are:
DCM：二氯甲烷DCM: Dichloromethane
DIPEA：N，N-二异丙基乙胺DIPEA: N,N-diisopropylethylamine
DMAP：4-二甲氨基吡啶DMAP: 4-Dimethylaminopyridine
DMF：N，N-二甲基甲酰胺DMF: N,N-dimethylformamide
EA：乙酸乙酯EA: ethyl acetate
EDCI:1-乙基-(3-二甲基氨基丙基)碳酰二亚胺盐酸盐EDCI: 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride
HBTU：O-苯并三氮唑-四甲基脲六氟磷酸酯HBTU: O-benzotriazole-tetramethylurea hexafluorophosphate
HOBt：1-羟基苯并***HOBt: 1-Hydroxybenzotriazole
HONB：N-羟基-5-降冰片烯-2,3-二甲酰亚胺HONB: N-hydroxy-5-norbornene-2,3-dicarboximide
PE：石油醚PE: petroleum ether
TFA：三氟乙酸TFA: Trifluoroacetic acid
TIS：三异丙基硅烷TIS: Triisopropylsilane
实施例1Example 1
2-(2-(2-(3，4，5-三(异辛氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-001)的合成：2-(2-(2-(3,4,5-Tris(isooctyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL-001) synthesis:
Figure PCTCN2019125143-appb-000015
Figure PCTCN2019125143-appb-000015
3，4，5-三羟基苯甲酸甲酯(18.4g，0.1mol，1-1)、异辛基溴(2-乙基-己基溴)(63.7g，0.33mol)、碳酸钾(55.2g，0.4mol)和DMF(150mL)常温混合，加热至110-120℃反应12h。冷却至室温，搅拌下倒入石油醚(150mL)和水(150mL)混合液中。分去下层，上层石油醚溶液用水(200mL×2)洗涤，浓缩得油状中间体1-2粗品；3,4,5-Trihydroxybenzoic acid methyl ester (18.4g, 0.1mol, 1-1), isooctyl bromide (2-ethyl-hexyl bromide) (63.7g, 0.33mol), potassium carbonate (55.2g , 0.4mol) and DMF (150mL) are mixed at room temperature and heated to 110-120°C for 12h. Cool to room temperature and pour into a mixture of petroleum ether (150 mL) and water (150 mL) while stirring. Separate the lower layer, wash the upper layer with petroleum ether solution with water (200mL×2), and concentrate to obtain the crude oily intermediate 1-2;
将中间体1-2粗品溶于四氢呋喃(100mL)，加入甲醇(50mL)和30％NaOH溶液(67g)，搅拌反应3h，加入石油醚(100mL)和水(200mL)，搅拌10min后，分去下层；加入2N盐酸(100mL)，再搅拌10min，分去下层；上层水洗至pH 6-7，浓缩得3，4，5-三(异辛氧基)-苯甲酸48.0g(1-2)；The crude intermediate 1-2 was dissolved in tetrahydrofuran (100mL), methanol (50mL) and 30% NaOH solution (67g) were added, and the reaction was stirred for 3h. Petroleum ether (100mL) and water (200mL) were added. After stirring for 10min, the mixture was separated. Lower layer; add 2N hydrochloric acid (100mL), stir for 10 min, separate the lower layer; wash the upper layer to pH 6-7, concentrate to obtain 3,4,5-tris(isooctyloxy)-benzoic acid 48.0g (1-2) ；
3，4，5-三(异辛氧基)-苯甲酸(45.5g，0.09mol)、2-(2-氯乙氧基)-乙胺盐酸盐(16.0g，0.1mol)、和二氯甲烷(200mL)混合，磁力搅拌，冷却至5-10℃；依次加入HOBt(13.5g，0.1mol)、二异丙基乙胺(18g，0.14mol)和EDCI(19.2，0.1mol)；在5-10℃下保温10min，撤去冷浴，自然升至室温反应3h。依次用水(100mL)、饱和碳酸氢钠(100mL×2)、1N盐酸(50mL)和饱和食盐水(50mL)洗涤，浓缩得2-(2-(3，4，5-三(异辛氧基)-苯甲酰胺基)-乙氧基)-乙基氯55.1g(1-3)；3,4,5-Tris(isooctyloxy)-benzoic acid (45.5g, 0.09mol), 2-(2-chloroethoxy)-ethylamine hydrochloride (16.0g, 0.1mol), and two Mix chloromethane (200mL), magnetically stir, and cool to 5-10°C; add HOBt (13.5g, 0.1mol), diisopropylethylamine (18g, 0.14mol) and EDCI (19.2, 0.1mol) in sequence; Incubate at 5-10°C for 10 minutes, remove the cold bath, and naturally rise to room temperature for 3 hours. Wash with water (100mL), saturated sodium bicarbonate (100mL×2), 1N hydrochloric acid (50mL) and saturated brine (50mL) successively, and concentrate to obtain 2-(2-(3,4,5-tris(isooctyloxy) )-Benzamido)-ethoxy)-ethyl chloride 55.1g (1-3);
2-羟基-9-芴酮(9.8g，0.05mol，1-4)、2-(2-(3，4，5-三(异辛氧基)-苯甲酰胺基)-乙氧基)-乙基氯(30.6g，0.06mol)、碳酸钾(16.6g，0.12mol)、碘化钾(0.05g)和DMF(150mL)常温混合，机械搅拌，加热至115-120℃反应8-10h，冷却至室温，搅拌下倒入石油醚(150ml)和水(150ml)混合液中。分去下层，石油醚层用水(200mL×2)洗，减压浓缩得2-(2-(2-(3，4，5-三(异辛氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-芴酮(1-5)粗品。2-hydroxy-9-fluorenone (9.8g, 0.05mol, 1-4), 2-(2-(3,4,5-tris(isooctyloxy)-benzamide)-ethoxy) -Ethyl chloride (30.6g, 0.06mol), potassium carbonate (16.6g, 0.12mol), potassium iodide (0.05g) and DMF (150mL) are mixed at room temperature, mechanically stirred, heated to 115-120℃ for 8-10h, and cooled To room temperature, pour into a mixture of petroleum ether (150ml) and water (150ml) while stirring. The lower layer was separated, the petroleum ether layer was washed with water (200mL×2), and concentrated under reduced pressure to obtain 2-(2-(2-(3,4,5-tris(isooctyloxy)-benzamide)-ethoxy (Yl)-ethoxy)-9-fluorenone (1-5) crude product.
将以上芴酮(1-5)粗品溶解于无水乙醇(100mL)和四氢呋喃(100mL)，加入盐酸羟胺(5.2g，0.075mol)和三乙胺(7.6g，0.075mol)，加热至75-80℃反应4-5h；冷却至室温，将反应液倒入石油醚(150mL)和水(300mL)混合液中。分去下层，石油醚层用水(200mL×2)洗，减压浓缩得中间体肟(1-6)粗品。将肟粗品溶解于四氢呋喃(120mL)，加入醋酸(50mL)，加热至55-60℃，分批加入锌粉(9.8g，0.15mol)。加毕，保温继续反应2h，过滤除去过量的锌粉，浓缩除去四氢呋喃和醋酸。加入石油醚(200mL)重新溶解，依次用10％NaOHaq(100mL×2)和饱和食盐水洗，浓缩得FL-001粗品。柱层析纯化(100％PE→100％EA→95％EA+5％MeOH)得2-(2-(2-(3，4，5-三(异辛氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-001)22.5g。Dissolve the above crude fluorenone (1-5) in absolute ethanol (100mL) and tetrahydrofuran (100mL), add hydroxylamine hydrochloride (5.2g, 0.075mol) and triethylamine (7.6g, 0.075mol), and heat to 75- React at 80°C for 4-5h; cool to room temperature, pour the reaction solution into a mixture of petroleum ether (150mL) and water (300mL). The lower layer was separated, the petroleum ether layer was washed with water (200 mL×2), and concentrated under reduced pressure to obtain the crude intermediate oxime (1-6). The crude oxime was dissolved in tetrahydrofuran (120mL), acetic acid (50mL) was added, heated to 55-60°C, and zinc powder (9.8g, 0.15mol) was added in batches. After the addition, keep warm and continue the reaction for 2h, filter to remove excess zinc powder, concentrate to remove tetrahydrofuran and acetic acid. Add petroleum ether (200 mL) to re-dissolve, wash with 10% NaOHaq (100 mL×2) and saturated brine successively, and concentrate to obtain FL-001 crude product. Purification by column chromatography (100%PE→100%EA→95%EA+5%MeOH) to obtain 2-(2-(2-(3,4,5-tris(isooctyloxy)-benzamide) -Ethoxy)-Ethoxy)-fluorene-9-amine (FL-001) 22.5g.
1H-NMR(400MHz，CDCl 3)：0.85-1.00(m，18H)，1.30-1.80(m，27H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(s，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：773.5461。 1 H-NMR (400MHz, CDCl 3 ): 0.85-1.00 (m, 18H), 1.30-1.80 (m, 27H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m , 1H), 6.90-6.97 (d, 1H), 7.00 (s, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[ M+1] + : 773.5461.
实施例2Example 2
2-(2-(2-(3，4，5-三(异壬氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-002)的合成：2-(2-(2-(3,4,5-Tris(isononyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL-002) synthesis:
实施例2中的FL-002的合成方法与实施例1一致，不同之处在于：实施例2以异壬基溴(3，5，5-三甲基-己基溴)代替实施例1中的异辛基溴；The synthesis method of FL-002 in Example 2 is the same as that in Example 1, except that: Example 2 uses isononyl bromide (3,5,5-trimethyl-hexyl bromide) to replace the one in Example 1. Isooctyl bromide;
Figure PCTCN2019125143-appb-000016
Figure PCTCN2019125143-appb-000016
1H-NMR(400MHz，CDCl 3)：0.85-1.00(m，36H)，1.15-1.25(m，3H)，1.20-1.30(m，3H)，1.55-1.65(m，3H)，1.70-1.90(m，6H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(s，2H)，7.15-7.25(m，2H)， 7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：815.9930。 1 H-NMR (400MHz, CDCl 3 ): 0.85-1.00 (m, 36H), 1.15-1.25 (m, 3H), 1.20-1.30 (m, 3H), 1.55-1.65 (m, 3H), 1.70-1.90 (m, 6H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (s, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 815.9930.
实施例3Example 3
2-(2-(2-(3，4，5-三(异构十烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-003)的合成：2-(2-(2-(3,4,5-tris(isodecyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL-003 )Synthesis:
实施例3中的FL-003的合成方法与实施例1一致，不同之处在于：以异构十烷基溴(通过Exxal 10溴化制备)代替实施例1中的异辛基溴；The synthesis method of FL-003 in Example 3 is the same as that in Example 1, except that: isooctyl bromide (prepared by Exxal 10 bromination) is used instead of isooctyl bromide in Example 1;
Figure PCTCN2019125143-appb-000017
Figure PCTCN2019125143-appb-000017
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，57H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：857.6401。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 57H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 857.6401 .
异构十烷基溴的制备(溴化物的制备)：Preparation of isodecyl bromide (preparation of bromide):
异构十醇(Exxal 10)(100mL)和37％HBr(300mL)在三口烧瓶中混合，机械搅拌下，缓慢加入浓硫酸(30mL)，混合反应液加热到115-120℃反应。GC分析醇<0.5％后停止反应。降温至50℃以下，加入石油醚(100mL)萃取，石油醚层用水先至中性，减压除去石油醚后得到异构十烷基溴。Exxal 10 (100 mL) and 37% HBr (300 mL) were mixed in a three-necked flask. Under mechanical stirring, concentrated sulfuric acid (30 mL) was slowly added, and the mixed reaction solution was heated to 115-120°C for reaction. The reaction was stopped after GC analysis of alcohol <0.5%. The temperature was lowered to below 50° C., petroleum ether (100 mL) was added for extraction, the petroleum ether layer was first neutralized with water, and the petroleum ether was removed under reduced pressure to obtain isodecyl bromide.
实施例4Example 4
2-(2-(2-(3，4，5-三(八/十/十二烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-004)的合成：2-(2-(2-(3,4,5-Tris(octa/dec/dodecyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine ( FL-004) Synthesis:
实施例4中的FL-004的合成方法与实施例1一致，不同之处在于：以正辛烷基溴、正癸烷基溴和正十二烷基溴(1：1：1)代替实施例1中的异辛基溴；The synthesis method of FL-004 in Example 4 is the same as that in Example 1, except that: n-octyl bromide, n-decyl bromide and n-dodecyl bromide (1:1:1) are used instead of the examples. 1 isooctyl bromide;
Figure PCTCN2019125143-appb-000018
Figure PCTCN2019125143-appb-000018
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，57H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：829.6087，857.6401，885.6716。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 57H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + :829.6087 , 857.6401, 885.6716.
实施例5Example 5
2-(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-005)的合成：2-(2-(2-(3,4,5-Tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL- 005) Synthesis:
实施例5中的FL-005的合成方法与实施例1一致，不同之处在于：以异构十三烷基溴(通过Exxal 13代替实施例3中的Exxal 10按实施例3溴化物的制备方法制备)代替实施例1中的异辛基溴；The synthesis method of FL-005 in Example 5 is the same as that of Example 1, but the difference is: isotridecyl bromide (Exxal 13 is used instead of Exxal 10 in Example 3). Preparation of bromide in Example 3 Method preparation) instead of isooctyl bromide in Example 1;
Figure PCTCN2019125143-appb-000019
Figure PCTCN2019125143-appb-000019
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：983.7810。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 983.7810 .
实施例6Example 6
2-(2-(2-(3，4，5-三(2，3-二氢植烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-006)的合成：2-(2-(2-(3,4,5-Tris(2,3-dihydrophytaalkoxy)-benzamide)-ethoxy)-ethoxy)-fluoren-9-amine Synthesis of (FL-006):
实施例6中的FL-006的合成方法与实施例1一致，不同之处在于：以2，3-二氢植烷基溴(3，7，11，15-四甲基－十六烷基溴)代替实施例1中的异辛基溴；The synthesis method of FL-006 in Example 6 is the same as that of Example 1, except that 2,3-dihydrophytyl bromide (3,7,11,15-tetramethyl-hexadecyl Bromine) instead of isooctyl bromide in Example 1;
Figure PCTCN2019125143-appb-000020
Figure PCTCN2019125143-appb-000020
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，117H)，3.60-4.20(m，14H)，4.71(s，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1278.1096。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 117H), 3.60-4.20 (m, 14H), 4.71 (s, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + :1278.1096 .
实施例7Example 7
2-(2-(2-(N'-苄基-3，4，5-三(异壬氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-007)的合成2-(2-(2-(N'-Benzyl-3,4,5-tris(isononyloxy)-benzamide)-ethoxy)-ethoxy)-fluoren-9-amine (FL-007) Synthesis
Figure PCTCN2019125143-appb-000021
Figure PCTCN2019125143-appb-000021
实施例7的FL-007合成方法同实施例1，不同之处在于：以异壬基溴代替异辛基溴，N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐代替实施例1中的2-(2-氯乙基)氧基)-乙胺盐酸盐；The synthesis method of FL-007 in Example 7 is the same as that in Example 1, except that isononyl bromide is used instead of isooctyl bromide, N-benzyl-2-(2-chloroethyl)oxy-ethylamine salt Acid salt instead of 2-(2-chloroethyl)oxy)-ethylamine hydrochloride in Example 1;
1H-NMR(400MHz，CDCl 3)：δ0.80-1.40(m，42H)，1.50-2.00(m，9H)，3.40-4.20(m，14)，4.71(s，1H)，4.80-4.90(m，2H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.40(m，8H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：905.6402。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-1.40 (m, 42H), 1.50-2.00 (m, 9H), 3.40-4.20 (m, 14), 4.71 (s, 1H), 4.80-4.90 (m, 2H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.40 (m, 8H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 905.6402 .
N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐的制备Preparation of N-benzyl-2-(2-chloroethyl)oxy-ethylamine hydrochloride
二甘醇胺(21.0g，0.2mol)溶解于无水乙醇(100mL)，加入苯甲醛(21.2g，0.2mol)，常温反应半小时；降温至5-10℃，加入硼氢化钠(11.4g，0.3mol)，升至室温反应2h；降温至5-10℃，用浓盐酸调pH至1，减压除去乙醇；余下物加水(200mL)溶解，用二氯甲烷(50mL)萃取，丢弃二氯甲烷层，水层用10％NaOH溶液调pH值至>14；用二氯甲烷(100mL×2)萃取，合并二氯甲烷萃取液，用水(50mL)和饱和食盐水(50mL)洗，无水硫酸钠干燥，浓缩得N-苄基-二乙醇胺粗品(23.4g)；Diglycolamine (21.0g, 0.2mol) was dissolved in absolute ethanol (100mL), added benzaldehyde (21.2g, 0.2mol), reacted at room temperature for half an hour; cooled to 5-10℃, added sodium borohydride (11.4g , 0.3mol), warm to room temperature and react for 2h; cool to 5-10°C, adjust pH to 1 with concentrated hydrochloric acid, remove ethanol under reduced pressure; add water (200mL) to dissolve the remainder, extract with dichloromethane (50mL), discard the second The methyl chloride layer and the aqueous layer were adjusted to pH >14 with 10% NaOH solution; extracted with dichloromethane (100mL×2), combined the dichloromethane extracts, washed with water (50mL) and saturated brine (50mL), no Dry with sodium sulfate and concentrate to obtain crude N-benzyl-diethanolamine (23.4g);
将N-苄基-二乙醇胺粗品(23.4g，0.12mol)溶解于二氯甲烷(150mL)，冷却至0-5℃，滴加氯化亚砜(42.8g，0.36mol)；滴加完毕后，加热回流反应3h，减压除去二氯甲烷和未反应的氯化亚砜；余下物加入甲苯(200mL)打浆，过滤干燥得N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐(25.0g)。Dissolve the crude N-benzyl-diethanolamine (23.4g, 0.12mol) in dichloromethane (150mL), cool to 0-5°C, add thionyl chloride (42.8g, 0.36mol) dropwise; after the addition is complete , Heating and refluxing for 3h, the dichloromethane and unreacted thionyl chloride were removed under reduced pressure; the remainder was added to toluene (200mL) to make a slurry, filtered and dried to obtain N-benzyl-2-(2-chloroethyl)oxy- Ethylamine hydrochloride (25.0g).
实施例8Example 8
2-(2-(2-(N'-苄基-3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-008)的合成2-(2-(2-(N'-Benzyl-3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene- Synthesis of 9-amine (FL-008)
Figure PCTCN2019125143-appb-000022
Figure PCTCN2019125143-appb-000022
实施例8的FL-008合成方法同实施例1，不同之处在于：以异构十三烷基溴代替异辛基溴，N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐代替实施例1中的2-(2-氯乙基)氧基-乙胺盐酸 盐；The synthesis method of FL-008 in Example 8 is the same as that in Example 1, except that: isooctyl bromide is replaced by iso-tridecyl bromide, and N-benzyl-2-(2-chloroethyl)oxy- Ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，57H)，3.40-4.20(m，14)，4.71(s，1H)，4.80-4.90(m，2H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.40(m，8H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1073.8278。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 57H), 3.40-4.20 (m, 14), 4.71 (s, 1H), 4.80-4.90 (m, 2H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.40 (m, 8H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 1073.8278.
实施例9Example 9
2，7-二(2-(2-(3，4，5-三(异壬氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-009)的合成：2,7-Bis(2-(2-(3,4,5-tris(isononyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL- 009) Synthesis:
Figure PCTCN2019125143-appb-000023
Figure PCTCN2019125143-appb-000023
实施例9的FL-009合成方法同实施例2，不同之处在于：以2，7-二羟基-9-芴酮代替2-羟基-9-芴酮；The synthesis method of FL-009 in embodiment 9 is the same as that in embodiment 2, except that: 2,7-dihydroxy-9-fluorenone is used instead of 2-hydroxy-9-fluorenone;
1H-NMR(400MHz，CDCl 3)：0.85-1.00(m，72H)，1.15-1.25(m，6H)，1.20-1.30(m，6H)，1.55-1.65(m，6H)，1.70-1.90(m，12H)，3.60-4.20(m，28H)，4.71(s，1H)，6.60-6.65(m，2H)，6.90-6.97(d，2H)，7.00(s，4H)，7.15-7.20(m，2H)，7.40-7.50(d，2H)；HRMS TOF[M+1] +：1449.0900。 1 H-NMR (400MHz, CDCl 3 ): 0.85-1.00 (m, 72H), 1.15-1.25 (m, 6H), 1.20-1.30 (m, 6H), 1.55-1.65 (m, 6H), 1.70-1.90 (m, 12H), 3.60-4.20 (m, 28H), 4.71 (s, 1H), 6.60-6.65 (m, 2H), 6.90-6.97 (d, 2H), 7.00 (s, 4H), 7.15-7.20 (m, 2H), 7.40-7.50 (d, 2H); HRMS TOF[M+1] + : 1449.0900.
实施例10Example 10
2，7-二(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-009)的合成：2,7-Bis(2-(2-(3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-fluoren-9-amine Synthesis of (FL-009):
Figure PCTCN2019125143-appb-000024
Figure PCTCN2019125143-appb-000024
实施例10的FL-010合成方法同实施例5，不同之处在于：以2，7-二羟基-9-芴酮代替2-羟基-9-芴酮；The synthesis method of FL-010 of embodiment 10 is the same as that of embodiment 5, except that: 2,7-dihydroxy-9-fluorenone is used instead of 2-hydroxy-9-fluorenone;
1H-NMR(400MHz，CDCl 3)：0.85-2.00(m，150H)，3.60-4.20(m，28H)，4.71(s，1H)，6.60-6.65(m，2H)，6.90-6.97(d，2H)，7.00(m，4H)，7.15-7.20(m，2H)，7.40-7.50(d，2H)；HRMS TOF[M+1] +：1784.4582。 1 H-NMR (400MHz, CDCl 3 ): 0.85-2.00 (m, 150H), 3.60-4.20 (m, 28H), 4.71 (s, 1H), 6.60-6.65 (m, 2H), 6.90-6.97 (d , 2H), 7.00 (m, 4H), 7.15-7.20 (m, 2H), 7.40-7.50 (d, 2H); HRMS TOF[M+1] + : 1784.4582.
实施例11Example 11
2-(2-(2-(N'-(3，4-二(异构十三烷氧基)-苄基)-乙酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-011)的合成：2-(2-(2-(N'-(3,4-bis(isotridecyloxy)-benzyl)-acetamido)-ethoxy)-ethoxy)-fluorene-9 -Synthesis of amine (FL-011):
Figure PCTCN2019125143-appb-000025
Figure PCTCN2019125143-appb-000025
3，4-二羟基-苯甲醛(13.8g，0.1mol，11-1)、异构十三烷基溴(63.1g，0.24mol)、碳酸钾(41.4g，0.3mol)和DMF(200mL)混合，加热至90-100℃反应10h；搅拌下倒入预先放入石油醚(200mL)和水(200mL)的反应瓶中，分去下层，上层用水(100mL)和饱和食盐水(100mL)洗涤，浓缩得中间体11-2粗品；3,4-Dihydroxy-benzaldehyde (13.8g, 0.1mol, 11-1), isotridecyl bromide (63.1g, 0.24mol), potassium carbonate (41.4g, 0.3mol) and DMF (200mL) Mix, heat to 90-100°C for 10 hours; pour into a reaction flask pre-placed with petroleum ether (200mL) and water (200mL) under stirring, separate the lower layer, and wash the upper layer with water (100mL) and saturated brine (100mL) , Concentrated to obtain crude intermediate 11-2;
以上粗品溶解于四氢呋喃(100mL)和乙醇(200mL)，加入2-(2-氯乙基)氧基-乙胺盐酸盐(16.0g，0.1mol)和三乙胺(10.0g，0.1mol)，常温下反应3h；降温至0-5℃，加入硼氢化钠(3.8g，0.1mol)；自然升至室温反应2h，加入1N盐酸至无气泡产生，pH 7-8；加入石油醚(300mL)和水(300mL)，搅拌10min后静置分层，分去下层，上层用水(100mL×2)和饱和食盐水洗，浓缩得中间体11-3粗品；The above crude product was dissolved in tetrahydrofuran (100mL) and ethanol (200mL), and 2-(2-chloroethyl)oxy-ethylamine hydrochloride (16.0g, 0.1mol) and triethylamine (10.0g, 0.1mol) were added , React at room temperature for 3h; cool to 0-5℃, add sodium borohydride (3.8g, 0.1mol); naturally warm to room temperature and react for 2h, add 1N hydrochloric acid until no bubbles are generated, pH 7-8; add petroleum ether (300mL ) And water (300mL), stirred for 10min and then allowed to stand to separate the layers, separate the lower layer, wash the upper layer with water (100mL×2) and saturated brine, and concentrate to obtain the crude intermediate 11-3;
将中间体11-3粗品溶解于乙酸乙酯(300mL)；加入三乙胺(15.0g，0.15mol)和醋酐(10.3g，0.1mol)，常温下反应半小时；加入水搅拌10min，分去水层；上层乙酸乙酯溶液水(100mL×2)洗，浓缩得中间体11-4粗品，柱层析纯化(石油醚→石油醚：乙酸乙酯＝10:1)得中间体11-4 38.0g；Dissolve the crude intermediate 11-3 in ethyl acetate (300mL); add triethylamine (15.0g, 0.15mol) and acetic anhydride (10.3g, 0.1mol), react for half an hour at room temperature; add water and stir for 10min, divide Dewatering layer; the upper layer was washed with ethyl acetate solution (100mL×2), concentrated to obtain crude intermediate 11-4, and purified by column chromatography (petroleum ether→petroleum ether: ethyl acetate=10:1) to obtain intermediate 11- 4 38.0g;
以中间体11-4和2-羟基-9-芴酮为原料，按实施例1的中间体1-3至FL-001的方法步骤合成(2-(2-(N’-(3，4-二(异构十三烷氧基)-苄基)-乙酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-011)。Using intermediate 11-4 and 2-hydroxy-9-fluorenone as raw materials, according to the method steps of intermediate 1-3 to FL-001 in Example 1, synthesis (2-(2-(N'-(3, 4 -Di(isotridecyloxy)-benzyl)-acetamido)-ethoxy)-ethoxy)-fluorene-9-amine (FL-011).
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，50H)，2.10-2.20(s，3H)，3.60-4.20(m，12H)，4.68(s，2H)，4.71(s，1H)，6.60-6.65(m，2H)，6.70-6.75(m，1H)，6.90-6.97(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：813.6140。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 50H), 2.10-2.20 (s, 3H), 3.60-4.20 (m, 12H), 4.68 (s, 2H), 4.71 (s , 1H), 6.60-6.65 (m, 2H), 6.70-6.75 (m, 1H), 6.90-6.97 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55 -7.65 (m, 3H); HRMS TOF[M+1] + : 813.6140.
实施例12Example 12
2-(2-(2-(N'-(3，4-二(异构十三烷氧基)-苄基)-异壬酰胺基)-乙氧基)-乙氧基)-芴-9-胺的合成2-(2-(2-(N'-(3,4-bis(isotridecyloxy)-benzyl)-isononamido)-ethoxy)-ethoxy)-fluorene- Synthesis of 9-amine
Figure PCTCN2019125143-appb-000026
Figure PCTCN2019125143-appb-000026
实施例12的FL-012合成方法同实施例11，不同之处是以异壬酰氯代替实施例11的中的醋酐。The synthesis method of FL-012 in Example 12 is the same as that in Example 11, except that isononanoyl chloride replaces the acetic anhydride in Example 11.
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，65H)，2.10-2.40(m，2H)，3.60-4.20(m，12H)，4.68(s，2H)，4.71(s，1H)，6.60-6.65(m，2H)，6.70-6.75(m，1H)，6.90-6.97(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：911.7234。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 65H), 2.10-2.40 (m, 2H), 3.60-4.20 (m, 12H), 4.68 (s, 2H), 4.71 (s , 1H), 6.60-6.65 (m, 2H), 6.70-6.75 (m, 1H), 6.90-6.97 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55 -7.65 (m, 3H); HRMS TOF[M+1] + : 911.7234.
实施例13Example 13
2-(2-(2-(N'-(3，4-二(异构十三烷氧基)-苄基)-(3，4，5-三(异壬氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-013)的合成：2-(2-(2-(N'-(3,4-bis(isotridecyloxy)-benzyl)-(3,4,5-tris(isononyloxy)-benzamide Synthesis of (yl)-ethoxy)-ethoxy)-fluorene-9-amine (FL-013):
Figure PCTCN2019125143-appb-000027
Figure PCTCN2019125143-appb-000027
以3，4，5-三异壬氧基苯甲酸(实施例2的中间体)代替中间体1-2和中间体11-3代替2-(2-氯乙氧基)-乙胺盐酸盐，按照实施例1的方法合成3-(2-(2-(N'-(3，4-二(异构十三烷氧基)-苄基)-(3，4，5-三(异壬氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-013)；Replace Intermediate 1-2 and Intermediate 11-3 with 3,4,5-triisononyloxybenzoic acid (intermediate of Example 2) instead of 2-(2-chloroethoxy)-ethylamine hydrochloride Salt, according to the method of Example 1 to synthesize 3-(2-(2-(N'-(3,4-bis(isotridecyloxy)-benzyl)-(3,4,5-tri( (Isononyloxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL-013);
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，101H)，3.40-4.20(m，18H)，4.60-4.75 (m，3H)，6.60-6.65(m，2H)，6.70-6.75(m，1H)，6.90-6.97(m，4H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1302.0366。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 101H), 3.40-4.20 (m, 18H), 4.60-4.75 (m, 3H), 6.60-6.65 (m, 2H), 6.70 -6.75 (m, 1H), 6.90-6.97 (m, 4H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1 ] + : 1302.0366.
实施例14Example 14
2-(2-(2-(3，5-二(2，3-二氢植烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-014)的合成：2-(2-(2-(3,5-bis(2,3-dihydrophytalkoxy)-benzamide)-ethoxy)-ethoxy)-fluorene-9-amine (FL -014) Synthesis:
Figure PCTCN2019125143-appb-000028
Figure PCTCN2019125143-appb-000028
实施例14的FL-014合成方法同实施例6，不同之处在于：以3，5-二羟基苯甲酸甲酯代替实施例6中的3，4，5-三羟基苯甲酸甲酯；The synthesis method of FL-014 in Example 14 is the same as that in Example 6, except that methyl 3,5-dihydroxybenzoate is used instead of methyl 3,4,5-trihydroxybenzoate in Example 6;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，78H)，3.60-4.20(m，12H)，4.70(s，1H)，6.50-6.60(m，1H)，6.60-6.65(m，1H)，6.85-6.97(d，3H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：981.8018。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 78H), 3.60-4.20 (m, 12H), 4.70 (s, 1H), 6.50-6.60 (m, 1H), 6.60-6.65 (m, 1H), 6.85-6.97 (d, 3H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 981.8018.
实施例15Example 15
2-(2-(2-(3，4-二(异构十三烷氧基)-苯乙酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-015)的合成：2-(2-(2-(3,4-bis(isotridecyloxy)-phenylacetamido)-ethoxy)-ethoxy)-fluorene-9-amine (FL-015) Synthesis:
Figure PCTCN2019125143-appb-000029
Figure PCTCN2019125143-appb-000029
实施例15的FL-015合成方法同实施例5，不同之处在于：以3，4-二羟基苯乙酸甲酯代替实施例5中的3，4，5-三羟基苯甲酸甲酯；The synthesis method of FL-015 in embodiment 15 is the same as that in embodiment 5, except that methyl 3,4-dihydroxyphenylacetate is used instead of methyl 3,4,5-trihydroxybenzoate in embodiment 5;
1H-NMR(400MHz，CDCl 3)：δ0.70-1.90(m，50H)，3.40-4.10(m，14H)，4.70(s，1H)，6.60-6.65(m，1H)，6.70-6.75(m，2H)，6.90-7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：799.5982。 1 H-NMR (400MHz, CDCl 3 ): δ 0.70-1.90 (m, 50H), 3.40-4.10 (m, 14H), 4.70 (s, 1H), 6.60-6.65 (m, 1H), 6.70-6.75 (m, 2H), 6.90-7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 799.5982.
实施例16Example 16
2-(2-(N'-甲基-3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-芴-9-胺(FL-016)的合成：2-(2-(N'-Methyl-3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-fluorene-9-amine (FL-016) Synthesis:
Figure PCTCN2019125143-appb-000030
Figure PCTCN2019125143-appb-000030
实施例16的FL-016合成方法同实施例5，不同之处在于：以N-甲基-2-氯乙胺盐酸盐代替实施例5中的2-(2-氯乙基)氧基-乙胺盐酸盐；The synthesis method of FL-016 in Example 16 is the same as that in Example 5, except that: N-methyl-2-chloroethylamine hydrochloride is used instead of 2-(2-chloroethyl)oxy in Example 5 -Ethylamine hydrochloride;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，2.90-3.05(m，3H)，3.60-4.20(m，10H)，4.71(s，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：953.7703。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.90-3.05 (m, 3H), 3.60-4.20 (m, 10H), 4.71 (s, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 953.7703 .
实施例17Example 17
2，7-二(2-(N'-苄基-2-异壬酰胺基-乙氧基)-乙氧基)-芴-9-胺(FL-017)的合成：Synthesis of 2,7-bis(2-(N'-benzyl-2-isononamido-ethoxy)-ethoxy)-fluorene-9-amine (FL-017):
Figure PCTCN2019125143-appb-000031
Figure PCTCN2019125143-appb-000031
实施例17的FL-017合成方法同实施例1，不同之处在于：异壬酸代替实施例1中的3，4，5-二异辛基苯甲酸(中间体1-2)，以N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐代替实施例1中的2-(2-氯乙基)氧基-乙胺盐酸盐，以2，7-二羟基-9-芴酮代替2-羟基-9-芴酮；The synthesis method of FL-017 in Example 17 is the same as that in Example 1, except that isononanoic acid replaces 3,4,5-diisooctylbenzoic acid (Intermediate 1-2) in Example 1, and N -Benzyl-2-(2-chloroethyl)oxy-ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1, with 2,7- Dihydroxy-9-fluorenone replaces 2-hydroxy-9-fluorenone;
1H-NMR(400MHz，CDCl 3)：δ0.8-1.90(m，30H)，2.10-2.35(m，4H)3.40-3.80(m，12H)，4.10-4.20(m，4H)，4.60-4.80(m，5H)，6.90-6.97(d，2H)，7.00(m，4H)，7.15-7.40(m，12H)，7.40-7.50(d，2H)；HRMS TOF[M+1] +：848.5572。 1 H-NMR (400MHz, CDCl 3 ): δ 0.8-1.90 (m, 30H), 2.10-2.35 (m, 4H) 3.40-3.80 (m, 12H), 4.10-4.20 (m, 4H), 4.60- 4.80 (m, 5H), 6.90-6.97 (d, 2H), 7.00 (m, 4H), 7.15-7.40 (m, 12H), 7.40-7.50 (d, 2H); HRMS TOF[M+1] + : 848.5572.
实施例18Example 18
2-(2-(N'-苄基-2-异硬脂酰胺基-乙氧基)-乙氧基)-芴-9-胺(FL-018)的合成：Synthesis of 2-(2-(N'-benzyl-2-isostearylamino-ethoxy)-ethoxy)-fluorene-9-amine (FL-018):
Figure PCTCN2019125143-appb-000032
Figure PCTCN2019125143-appb-000032
实施例18的FL-018合成方法同实施例1，不同之处在于：异硬脂酸代替实施例1中的3，4，5-二异辛基苯甲酸(中间体1-2)，以N-苄基-2-(2-氯乙基)氧基-乙胺盐酸盐代替实施例1中的2-(2-氯乙基)氧基-乙胺盐酸盐；The synthesis method of FL-018 in Example 18 is the same as that in Example 1, except that: Isostearic acid replaces 3,4,5-diisooctylbenzoic acid (Intermediate 1-2) in Example 1 N-benzyl-2-(2-chloroethyl)oxy-ethylamine hydrochloride instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride in Example 1;
1H-NMR(400MHz，CDCl 3)：δ0.8-1.90(m，33H)，2.30-2.50(m，1H)3.40-4.10(m，8H)，4.60-4.80(m，3H)，6.90-6.97(d，1H)，7.15-7.40(m，8H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：641.4675。 1 H-NMR (400MHz, CDCl 3 ): δ 0.8-1.90 (m, 33H), 2.30-2.50 (m, 1H) 3.40-4.10 (m, 8H), 4.60-4.80 (m, 3H), 6.90- 6.97 (d, 1H), 7.15-7.40 (m, 8H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 641.4675.
实施例19Example 19
2-((5-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-2-酮-3-氮代-己基)氧基)-芴-9-胺(FL-019)的合成：2-((5-(3,4,5-Tris(isotridecyloxy)-benzamide)-2-one-3-aza-hexyl)oxy)-fluoren-9-amine Synthesis of (FL-019):
Figure PCTCN2019125143-appb-000033
Figure PCTCN2019125143-appb-000033
酸(19-1)(35.8g，0.05mol)、HONB(10.8g，0.06mol)溶解于DCM(150mL)；加入EDCI(11.5g，0.06mol)常温反应1h；反应液依次用水(100mL)、饱和碳酸氢钠水溶液(100mL)、1N盐酸(100mL)和饱和食盐水洗涤得中间体19-2的二氯甲烷溶液，此溶液直接用于下一步反应；Acid (19-1) (35.8g, 0.05mol), HONB (10.8g, 0.06mol) were dissolved in DCM (150mL); EDCI (11.5g, 0.06mol) was added to react at room temperature for 1h; the reaction solution was water (100mL), Wash with saturated sodium bicarbonate aqueous solution (100mL), 1N hydrochloric acid (100mL) and saturated brine to obtain a dichloromethane solution of Intermediate 19-2, which is directly used in the next reaction;
1，3-丙二胺(11.1g，0.15mol)溶解于二氯甲烷(20mL)，常温下滴入上一步所得二氯甲烷溶液；加毕，常温反应半小时，用水(100mL×2)洗涤得中间体19-3的二氯甲烷溶液，直接用于下一步反应；1,3-propanediamine (11.1g, 0.15mol) was dissolved in dichloromethane (20mL), and the dichloromethane solution obtained in the previous step was added dropwise at room temperature; after the addition, react for half an hour at room temperature and wash with water (100mL×2) Obtain the methylene chloride solution of Intermediate 19-3, which is directly used in the next reaction;
上一步所得二氯甲烷溶液冷却至0-5℃，加入三乙胺(10g，0.1mol)；保温0-5℃，滴加2-氯乙酰氯(5.6g，0.05mol)；滴毕保温0-5℃反应0.5h，依次用1N盐酸(100mL)，水(100mL)，饱和食盐水(50mL)洗涤，浓缩得中间体19-4粗品。The dichloromethane solution obtained in the previous step was cooled to 0-5°C, and triethylamine (10g, 0.1mol) was added; the temperature was maintained at 0-5°C, and 2-chloroacetyl chloride (5.6g, 0.05mol) was added dropwise; Reacted at -5°C for 0.5h, washed with 1N hydrochloric acid (100mL), water (100mL), saturated brine (50mL), and concentrated to obtain the crude intermediate 19-4.
以中间体19-4代替实施例1的中间体1-3，按实施例1方法步骤合成FL-019；Use Intermediate 19-4 instead of Intermediate 1-3 of Example 1, and synthesize FL-019 according to the method of Example 1;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.0(m，78H)，3.40-3.50(m，4H)，4.00-4.10(m，6H)，4.45-4.50(m，2H)，4.71(s，1H)，6.60-6.65(m，2H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1010.7920。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.0 (m, 78H), 3.40-3.50 (m, 4H), 4.00-4.10 (m, 6H), 4.45-4.50 (m, 2H), 4.71 (s, 1H), 6.60-6.65 (m, 2H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55 -7.65 (m, 3H); HRMS TOF[M+1] + : 1010.7920.
实施例20Example 20
2-(2-(N′-苄基-(3-(N″-苄基-3，4，5-三异构十三烷氧基-苯甲酰胺基)-丙胺基)-2-酮-乙氧基)-芴-9-胺(FL-020)的合成：2-(2-(N′-benzyl-(3-(N″-benzyl-3,4,5-triisotridecyloxy-benzamide)-propylamino)-2-one Synthesis of -ethoxy)-fluorene-9-amine (FL-020):
Figure PCTCN2019125143-appb-000034
Figure PCTCN2019125143-appb-000034
实施例20的FL-020合成方法同实施例19，不同之处在于：以N,N'-二苄基-1，3-丙二胺代替实施例19中的1，3-丙二胺；The synthesis method of FL-020 in Example 20 is the same as that in Example 19, except that: N,N'-dibenzyl-1,3-propanediamine is used instead of 1,3-propanediamine in Example 19;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.0(m，78H)，3.40-3.50(m，4H)，4.00-4.10(m，6H)，4.45-4.50(m，2H)，4.7-4.9(m，5H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.40(m，13H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1190.8858。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.0 (m, 78H), 3.40-3.50 (m, 4H), 4.00-4.10 (m, 6H), 4.45-4.50 (m, 2H), 4.7 -4.9 (m, 5H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.40 (m, 13H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 1190.8858.
实施例21Example 21
2-((2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-丙酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-021)的合成：2-((2-(2-(3,4,5-Tris(isotridecyloxy)-benzamide)-propionamido)-ethoxy)-ethoxy)-fluorene- Synthesis of 9-amine (FL-021):
Figure PCTCN2019125143-appb-000035
Figure PCTCN2019125143-appb-000035
Boc-Ala-OH(18.9g，0.1mol，21-1)、2-(2-氯乙基)氧基-乙胺盐酸盐(16.0g，0.1mol)、HOBt(14.9，0.11mol)和三乙胺(15.1g，0.15mol)混合于DMF(150mL)，冷 却至5-10℃；加入EDCI(21.1g，0.11mol)，保温5-10℃反应0.5h，自然升至室温反应1h；依次用水(100mL)、饱和碳酸氢钠(100mL)、1N盐酸(100mL)和饱和食盐水(50mL)洗涤；浓缩得中间体21-2粗品；Boc-Ala-OH (18.9g, 0.1mol, 21-1), 2-(2-chloroethyl)oxy-ethylamine hydrochloride (16.0g, 0.1mol), HOBt (14.9, 0.11mol) and Triethylamine (15.1g, 0.15mol) was mixed with DMF (150mL) and cooled to 5-10°C; EDCI (21.1g, 0.11mol) was added, kept at 5-10°C and reacted for 0.5h, and naturally warmed to room temperature for 1h; Wash with water (100mL), saturated sodium bicarbonate (100mL), 1N hydrochloric acid (100mL) and saturated brine (50mL) successively; concentrate to obtain the crude intermediate 21-2;
将以上粗品用***(200mL)溶解，滴加4N HCl/***溶液(200mL)，常温搅拌5h，过滤收集固体为中间体21-3；Dissolve the above crude product with ether (200mL), add dropwise 4N HCl/ether solution (200mL), stir at room temperature for 5h, filter and collect the solid as intermediate 21-3;
以中间体21-3代替2-(2-氯乙基)氧基-乙胺盐酸盐，按实施例5的方法合成FL-021；Using the intermediate 21-3 instead of 2-(2-chloroethyl)oxy-ethylamine hydrochloride, FL-021 was synthesized according to the method of Example 5;
1H-NMR(400MHz，CDCl 3):δ0.80-2.0(m，78H)，3.45-4.15(m，14)，4.60-4.75(m，2H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1054.8180。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.0 (m, 78H), 3.45-4.15 (m, 14), 4.60-4.75 (m, 2H), 6.60-6.65 (m, 1H), 6.90 -6.97(d, 1H), 7.00(m, 2H), 7.15-7.25(m, 2H), 7.30-7.40(m, 1H), 7.55-7.65(m, 3H); HRMS TOF[M+1] + : 1054.8180.
实施例22Example 22
2-(2-(2-(N″-甲基-3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-022)的合成：2-(2-(2-(N″-Methyl-3,4,5-tris(isotridecyloxy)-benzamide)-acetamido)-ethoxy)-ethoxy Synthesis of fluorene-9-amine (FL-022):
Figure PCTCN2019125143-appb-000036
Figure PCTCN2019125143-appb-000036
实施例22的FL-021合成方法同实施例21，不同之处在于：Boc-(Me)Gly-OH代替Boc-Ala-OH；The synthesis method of FL-021 in embodiment 22 is the same as that in embodiment 21, except that Boc-(Me)Gly-OH replaces Boc-Ala-OH;
1H-NMR(400MHz，CDCl 3):δ0.80-2.00(m，75H)，2.90-3.00(m，3H)，3.45-4.15(m，14)，4.55-4.60(s，2H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1054.8180。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.90-3.00 (m, 3H), 3.45-4.15 (m, 14), 4.55-4.60 (s, 2H), 6.90 -6.97(d, 1H), 7.00(m, 2H), 7.15-7.25(m, 2H), 7.30-7.40(m, 1H), 7.55-7.65(m, 3H); HRMS TOF[M+1] + : 1054.8180.
实施例23Example 23
2-(2-(2-((2，6-二(3，4，5-三(异壬烷氧基)-苯甲酰胺基)-己酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-023)的合成：2-(2-(2-((2,6-bis(3,4,5-tris(isononyloxy)-benzamide)-hexanamido)-ethoxy)-ethoxy Synthesis of )-fluorene-9-amine (FL-023):
Figure PCTCN2019125143-appb-000037
Figure PCTCN2019125143-appb-000037
赖氨酸(7.3g，0.05mol，23-1)溶解于四氢呋喃(100mL)和水(50mL)，加入LiOH(1.2g，0.05mol)；混合物加热至70℃，滴加活性酯19-2(96.6，0.11mol)的四氢呋喃(200mL)溶液；滴毕，保温反应半小时；冷至室温，加入1N盐酸(50mL)，用石油醚(200mL)萃取；上层石油醚溶液依次用水(100mL)和饱和食盐水(100mL)洗涤，浓缩得中间体23-2粗品；Lysine (7.3g, 0.05mol, 23-1) was dissolved in tetrahydrofuran (100mL) and water (50mL), LiOH (1.2g, 0.05mol) was added; the mixture was heated to 70°C, and active ester 19-2 ( 96.6, 0.11mol) in tetrahydrofuran (200mL); after dripping, keep the reaction for half an hour; cool to room temperature, add 1N hydrochloric acid (50mL), and extract with petroleum ether (200mL); the upper petroleum ether solution is successively water (100mL) and saturated Wash with brine (100 mL) and concentrate to obtain crude intermediate 23-2;
以中间体23-2代替实施例1的中间体1-2，按实施例1的方法步骤合成FL-023；Use Intermediate 23-2 instead of Intermediate 1-2 of Example 1, and synthesize FL-023 according to the method of Example 1;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，154H)，3.65-4.15(m，22)，4.55-4.60(m，1H)，6.30-6.35(m，1H)，6.90-6.97(m，2H)，7.00(m，4H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，4H)；HRMS TOF[M+1] +：1810.4973。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 154H), 3.65-4.15 (m, 22), 4.55-4.60 (m, 1H), 6.30-6.35 (m, 1H), 6.90 -6.97 (m, 2H), 7.00 (m, 4H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 4H); HRMS TOF[M+1] + : 1810.4973.
实施例24Example 24
2-(2-(2-(4-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-丁酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-024)的合成：2-(2-(2-(4-(3,4,5-Tris(isotridecyloxy)-benzamide)-butyramido)-ethoxy)-ethoxy)- Synthesis of fluorene-9-amine (FL-024):
Figure PCTCN2019125143-appb-000038
Figure PCTCN2019125143-appb-000038
实施例24的FL-024合成方法同实施例23，不同之处在于：以4-氨基-丁酸代替实施例23中的赖氨酸；The synthesis method of FL-024 in embodiment 24 is the same as that in embodiment 23, except that 4-amino-butyric acid is used instead of lysine in embodiment 23;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，77H)，2.25-2.35(m，2H)，3.40-3.50 (m，4H)，3.55-4.10(m，12H)，4.71(s，1H)，6.15-6.25(m，1H)，6.60-6.65(m，1H)，6.90-6.97(d，1H)，7.00(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：1068.8337。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 77H), 2.25-2.35 (m, 2H), 3.40-3.50 (m, 4H), 3.55-4.10 (m, 12H), 4.71 (s, 1H), 6.15-6.25 (m, 1H), 6.60-6.65 (m, 1H), 6.90-6.97 (d, 1H), 7.00 (m, 2H), 7.15-7.25 (m, 2H), 7.30 -7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 1068.8337.
实施例25Example 25
2-(2-(2-(2，6-二异硬脂酰胺基-己酰胺基)-乙氧基)-乙氧基)-芴-9-胺(FL-025)的合成：Synthesis of 2-(2-(2-(2,6-Diisostearamido-hexanamido)-ethoxy)-ethoxy)-fluorene-9-amine (FL-025):
Figure PCTCN2019125143-appb-000039
Figure PCTCN2019125143-appb-000039
赖氨酸(14.6.3g，0.1mol)溶解于四氢呋喃(150mL)和水(100mL)，加入NaOH(4.0g，0.1mol)；将反应液冷却至0-5℃；剧烈搅拌下同时滴加异硬脂酰氯(2，2，4，8，10，10-六甲基十一烷-5-酰氯)(66.6g，0.22mol)和20％NaOH(44g)，控制滴加速度使反应温度维持在0-5℃；滴毕，保温0-5℃反应半小时，用1N盐酸调pH值至1；用石油醚(300mL)萃取；石油醚溶液依次用水(100mL)和饱和食盐水(100mL)洗涤；浓缩，柱层析纯化得中间体二异硬脂酰赖氨酸50.5g；Lysine (14.6.3g, 0.1mol) was dissolved in tetrahydrofuran (150mL) and water (100mL), and NaOH (4.0g, 0.1mol) was added; the reaction solution was cooled to 0-5°C; Stearyl chloride (2,2,4,8,10,10-hexamethylundecane-5-acid chloride) (66.6g, 0.22mol) and 20% NaOH (44g), control the dropping rate to maintain the reaction temperature at 0-5℃; after dripping, keep at 0-5℃ and react for half an hour, adjust the pH to 1 with 1N hydrochloric acid; extract with petroleum ether (300mL); wash petroleum ether solution with water (100mL) and saturated brine (100mL) successively ; Concentrated and purified by column chromatography to obtain 50.5g of intermediate diisostearoyl lysine;
以二异硬脂酰赖氨酸代替实施例23的23-2按实施例23的步骤合成FL-025；Using diisostearoyl lysine instead of 23-2 in Example 23 to synthesize FL-025 according to the steps in Example 23;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，74H)，2.30-2.50(m，2H)，3.55-4.20(m，10H)，4.30-4.40(m，1H)，4.71(s，1H)，6.15-6.25(m，1H)，6.90-6.97(m，2H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，4H)；HRMS TOF[M+1] +：945.7766。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 74H), 2.30-2.50 (m, 2H), 3.55-4.20 (m, 10H), 4.30-4.40 (m, 1H), 4.71 (s, 1H), 6.15-6.25 (m, 1H), 6.90-6.97 (m, 2H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 4H) ; HRMS TOF[M+1] + : 945.7766.
实施例26Example 26
2-(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-苯基-芴-9-醇(FL-026)的合成：2-(2-(2-(3,4,5-Tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9-phenyl-fluorene-9 -Synthesis of alcohol (FL-026):
Figure PCTCN2019125143-appb-000040
Figure PCTCN2019125143-appb-000040
格氏试剂制备：Preparation of Grignard reagent:
镁粉(10.6g，0.44mol)浸泡于干燥四氢呋喃(50mL)，氮气保护；加入少量碘后，搅拌下缓慢滴加溴苯(62.8g，0.4mol，26-1)的干燥四氢呋喃(150mL)溶液；控制滴加速度使反应温度保持在40℃以下；加毕，保温40℃反应4h，得26-2。在反应液中加入2-羟基-9-芴酮(9.8g，0.05mol，26-3)的无水四氢呋喃(50mL)溶液，回流反应20h；将反应液冷却至室温，用1N氯化铵水溶液终止反应；用二氯甲烷(200mL×2)萃取；合并的萃取液用饱和食盐水(200mL×2)洗涤；减压浓缩得粗品；柱层析纯化得中间体26-4 7.8g。Magnesium powder (10.6g, 0.44mol) was soaked in dry tetrahydrofuran (50mL), protected by nitrogen; after adding a small amount of iodine, slowly add bromobenzene (62.8g, 0.4mol, 26-1) in dry tetrahydrofuran (150mL) solution with stirring ; Control the dropping rate to keep the reaction temperature below 40℃; after the addition, keep the temperature at 40℃ and react for 4h to obtain 26-2. Add 2-hydroxy-9-fluorenone (9.8g, 0.05mol, 26-3) in anhydrous tetrahydrofuran (50mL) solution to the reaction solution, reflux for 20h; cool the reaction solution to room temperature and use 1N ammonium chloride aqueous solution The reaction was terminated; extracted with dichloromethane (200 mL×2); the combined extracts were washed with saturated brine (200 mL×2); concentrated under reduced pressure to obtain a crude product; purified by column chromatography to obtain intermediate 26-4 7.8 g.
取中间体26-4(2.7g，0.01mol)溶解于DMF(50mL)，加入碳酸钾(5.5g，0.04mol)、碘化钾(0.1g)和中间体26-5(9.9g，0.012mol)，加热至80-85℃反应过夜；加入石油醚(200mL)和水(100mL)，搅拌10min后静置分层；分去水层，石油醚层用水(100mL)洗，浓缩得粗品。柱层析纯化(100％PE→100％EA)得FL-026 8.2g。Intermediate 26-4 (2.7g, 0.01mol) was dissolved in DMF (50mL), potassium carbonate (5.5g, 0.04mol), potassium iodide (0.1g) and intermediate 26-5 (9.9g, 0.012mol) were added, Heat to 80-85°C and react overnight; add petroleum ether (200 mL) and water (100 mL), stir for 10 min and then stand for separation; separate the water layer, wash the petroleum ether layer with water (100 mL), and concentrate to obtain a crude product. Column chromatography purification (100% PE→100% EA) obtained FL-026 8.2g.
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，2.80-2.90(m，1H)，3.60-4.20(m，14H)，6.60-6.65(m，1H)，6.80-6.91(m，3H)，7.00(m，2H)，7.15-7.40(m，7H)，7.55-7.65(m，2H)；HRMS TOF[M+1] +：1060.7965，[M-17] +：1042.7855。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.80-2.90 (m, 1H), 3.60-4.20 (m, 14H), 6.60-6.65 (m, 1H), 6.80 -6.91 (m, 3H), 7.00 (m, 2H), 7.15-7.40 (m, 7H), 7.55-7.65 (m, 2H); HRMS TOF[M+1] + : 1060.7965, [M-17] + : 1042.7855.
实施例27Example 27
2-(2-(2-(N′-苄基-3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-苯基-芴-9-醇(FL-027)的合成：2-(2-(2-(N′-Benzyl-3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9- Synthesis of phenyl-fluorene-9-ol (FL-027):
Figure PCTCN2019125143-appb-000041
Figure PCTCN2019125143-appb-000041
实施例27的FL-027合成方法同实施例26，不同之处在于：中间体27-1代替实施例26中的中间体26-5；The synthesis method of FL-027 in Example 27 is the same as that in Example 26, except that the intermediate 27-1 replaces the intermediate 26-5 in Example 26;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，2.80-2.90(m，1H)，3.60-4.20(m，14H)，4.70-4.75(m，2H)，6.80-6.91(m，3H)，7.00(m，2H)，7.15-7.40(m，12H)，7.55-7.65(m，2H)；HRMS TOF[M+1] +：1150.8430。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.80-2.90 (m, 1H), 3.60-4.20 (m, 14H), 4.70-4.75 (m, 2H), 6.80 -6.91 (m, 3H), 7.00 (m, 2H), 7.15-7.40 (m, 12H), 7.55-7.65 (m, 2H); HRMS TOF[M+1] + : 1150.8430.
实施例28Example 28
2-(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-(3-氟苯基)-芴-9-醇(FL-028)的合成：2-(2-(2-(3,4,5-Tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9-(3-fluorophenyl Synthesis of )-fluorene-9-ol (FL-028):
Figure PCTCN2019125143-appb-000042
Figure PCTCN2019125143-appb-000042
实施例28的FL-028合成方法同实施例26，不同之处在于：3-氟-1-溴苯代替溴苯；The synthesis method of FL-028 in embodiment 28 is the same as that in embodiment 26, except that: 3-fluoro-1-bromobenzene replaces bromobenzene;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，2.80-2.90(m，1H)，3.60-4.20(m，14H)，4.70-4.75(m，2H)，6.80-6.91(m，3H)，7.00(m，2H)，7.05-7.35(m，6H)，7.55-7.60(m，2H)；HRMS TOF[M+1] +：1078.7865。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.80-2.90 (m, 1H), 3.60-4.20 (m, 14H), 4.70-4.75 (m, 2H), 6.80 -6.91 (m, 3H), 7.00 (m, 2H), 7.05-7.35 (m, 6H), 7.55-7.60 (m, 2H); HRMS TOF[M+1] + : 1078.7865.
实施例29Example 29
2-(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-(4-氯苯基)-芴-9-醇(FL-029)的合成：2-(2-(2-(3,4,5-Tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9-(4-chlorophenyl Synthesis of )-fluorene-9-ol (FL-029):
Figure PCTCN2019125143-appb-000043
Figure PCTCN2019125143-appb-000043
实施例29的FL-029合成方法同实施例26，不同之处在于：4-氯-1-溴苯代替溴苯；The synthesis method of FL-029 in embodiment 29 is the same as that in embodiment 26, except that 4-chloro-1-bromobenzene replaces bromobenzene;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，75H)，2.80-2.90(m，1H)，3.60-4.20(m，14H)，6.60-6.65(m，1H)，6.80-6.91(m，2H)，7.00(m，2H)，7.15-7.45(m，7H)，7.55-7.60(m，2H)；HRMS TOF[M+1] +：1094.7572。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 75H), 2.80-2.90 (m, 1H), 3.60-4.20 (m, 14H), 6.60-6.65 (m, 1H), 6.80 -6.91 (m, 2H), 7.00 (m, 2H), 7.15-7.45 (m, 7H), 7.55-7.60 (m, 2H); HRMS TOF[M+1] + : 1094.7572.
实施例30Example 30
2,7-二(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-(3-氟苯基)-芴-9-醇(FL-030)的合成：2,7-Bis(2-(2-(3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9-(3- Synthesis of fluorophenyl)-fluorene-9-ol (FL-030):
Figure PCTCN2019125143-appb-000044
Figure PCTCN2019125143-appb-000044
实施例30的FL-030合成方法同实施例26，不同之处在于：2,7-二羟基-芴酮代替2-羟基-芴酮；The synthesis method of FL-030 in embodiment 30 is the same as that in embodiment 26, except that 2,7-dihydroxy-fluorenone replaces 2-hydroxy-fluorenone;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，150H)，3.00-3.05(m，1H)，3.60-4.10(m，28H)，6.60-6.65(m，2H)，6.85-6.91(m，2H)，7.00(m，4H)，7.10-7.20(s，2H)，7.20-7.35(m，5H),7.45-7.50(m，2H)；HRMS TOF[M+1] +：1862.4806。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 150H), 3.00-3.05 (m, 1H), 3.60-4.10 (m, 28H), 6.60-6.65 (m, 2H), 6.85 -6.91(m, 2H), 7.00(m, 4H), 7.10-7.20(s, 2H), 7.20-7.35(m, 5H), 7.45-7.50(m, 2H); HRMS TOF[M+1] + : 1862.4806.
实施例31Example 31
2,7-二(2-(2-(3，4，5-三(异构十三烷氧基)-苯甲酰胺基)-乙氧基)-乙氧基)-9-(3-氟苯基)-芴-9-醇(FL-031)的合成：2,7-Bis(2-(2-(3,4,5-tris(isotridecyloxy)-benzamide)-ethoxy)-ethoxy)-9-(3- Synthesis of fluorophenyl)-fluorene-9-ol (FL-031):
Figure PCTCN2019125143-appb-000045
Figure PCTCN2019125143-appb-000045
实施例31的FL-031合成方法同实施例27，不同之处在于：2,7-二羟基-芴酮代替2-羟基-芴酮；The synthesis method of FL-031 in Example 31 is the same as that in Example 27, except that: 2,7-dihydroxy-fluorenone replaces 2-hydroxy-fluorenone;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，150H)，3.10-3.15(m，1H)，3.60-4.20(m，28H)，6.60-6.65(m，2H)，6.80-6.90(m，4H)，6.90-7.00(m，4H)，7.00-7.05 (m，2H)，7.10-7.20(m，2H),7.40-7.45(m，2H)；HRMS TOF[M+1] +：1880.4717。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 150H), 3.10-3.15 (m, 1H), 3.60-4.20 (m, 28H), 6.60-6.65 (m, 2H), 6.80 -6.90 (m, 4H), 6.90-7.00 (m, 4H), 7.00-7.05 (m, 2H), 7.10-7.20 (m, 2H), 7.40-7.45 (m, 2H); HRMS TOF[M+1 ] + : 1880.4717.
实施例32Example 32
利用载体FL-002非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2，包括如下步骤： The heterogeneous synthesis of H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using carrier FL-002 includes the following steps:
1)将FL-002溶解于庚烷配成溶液，搅拌降温至15-20℃；Fmoc-Arg(Pbf)-OH(FL-002的1～1.5当量)、二异丙基乙胺(FL-002的1～2当量)和HOBt(FL-002的1～1.5当量)的DMF溶液加入与载体溶液混合；保持15-20℃，加入HBTU(FL-002的1～1.5当量)DMF溶液进行反应。1) Dissolve FL-002 in heptane to make a solution, stir and cool to 15-20℃; Fmoc-Arg(Pbf)-OH (1～1.5 equivalent of FL-002), diisopropylethylamine (FL- Add 1-2 equivalents of 002) and HOBt (1~1.5 equivalents of FL-002) DMF solution and mix with the carrier solution; keep at 15-20℃, add HBTU (1~1.5 equivalents of FL-002) DMF solution to react .
反应完全后加入水(庚烷和水的体积比为(1～15)：1)，继续搅拌10min，静置分层；分去下层，加入DMF和水混合溶液(DMF与水的体积比为(1～8)：1)，搅拌5min，静置分层；分去下层，上层直接用于脱Fmoc步骤。After the reaction is complete, add water (the volume ratio of heptane and water is (1～15):1), continue to stir for 10 minutes, stand still for layering; divide the lower layer, add a mixed solution of DMF and water (the volume ratio of DMF to water is (1-8): 1), stir for 5 min, stand still for layering; divide the lower layer, and use the upper layer directly in the Fmoc removal step.
2)上一步骤所得上层溶液加热至40-50℃，加入巯基丙酸、二乙烯三胺和DMF混合液(巯基丙酸和二乙烯三胺的用量为FL-002的3～5当量)，保温40-50℃反应。反应完全后加入DMF和水混合溶液(DMF和水的体积比为(1～5)：(5～1))，搅拌5min，静置分层；分去下层，上层加入DMF和水混合溶液(DMF与水的体积比为(1～8)：1)，搅拌5min，静置分层；分去下层，上层重复以上洗涤步骤至洗出液中性，上层直接用于下一个氨基酸缩合；2) The upper layer solution obtained in the previous step is heated to 40-50°C, and the mixture of mercaptopropionic acid, diethylenetriamine and DMF is added (the amount of mercaptopropionic acid and diethylenetriamine is 3 to 5 equivalents of FL-002), Incubate at 40-50℃ to react. After the reaction is complete, add a mixed solution of DMF and water (the volume ratio of DMF and water is (1～5): (5～1)), stir for 5 minutes, and let stand for layering; divide the lower layer, add the mixed solution of DMF and water ( The volume ratio of DMF to water is (1-8): 1), stir for 5 min, stand still for layering; divide the lower layer, repeat the above washing steps for the upper layer until the eluate is neutral, and use the upper layer directly for the next amino acid condensation;
3)按以上方法连续接入Fmoc-Arg(Pbf)-OH，Fmoc-Gln(Trt)-OH，Fmoc-Glu(OtBu)-OH，Fmoc-Met-OH，Fmoc-Glu(OtBu)-OH，并脱除Fmoc后得H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002)庚烷溶液，浓缩得固体；3) Continuously connect to Fmoc-Arg(Pbf)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Met-OH, Fmoc-Glu(OtBu)-OH according to the above method, After removing Fmoc, H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002) heptane solution is obtained, which is concentrated to obtain a solid;
4)将以上固体溶解于TFA：TIS：水(94：3：3，裂解液，是固体重量的5-10倍)，常温搅拌2h。缓慢滴加进0-10℃甲基叔丁基醚(甲基叔丁基醚的重量为裂解液的2-20倍)，保温0-10℃搅拌半小时；过滤，滤饼用甲基叔丁基醚洗至洗出液中性，干燥得H-Glu-Met-Glu-Gln-Arg-Arg-NH 2粗品，HPLC纯度87.3％，HRMS TOF[M+1] +:889.2309。 4) Dissolve the above solid in TFA:TIS:water (94:3:3, lysis solution, 5-10 times the weight of the solid), and stir for 2h at room temperature. Slowly add methyl tert-butyl ether at 0-10°C (the weight of methyl tert-butyl ether is 2-20 times of the lysate), keep it at 0-10°C and stir for half an hour; filter, filter cake with methyl tertiary The butyl ether was washed until the eluate was neutral, and dried to obtain a crude H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 product with an HPLC purity of 87.3% and HRMS TOF[M+1] + : 889.2309.
实施例33Example 33
利用载体FL-004非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-004 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-004，HPLC纯度87.5％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-004, and the HPLC purity is 87.5%.
实施例34Example 34
利用载体FL-005非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-005 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-005，HPLC纯度90.0％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-005, and the HPLC purity is 90.0%.
实施例35Example 35
利用载体FL-006非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-006 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-006，HPLC纯度85.0％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-006, and the HPLC purity is 85.0%.
实施例36Example 36
利用载体FL-008非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-008 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-008，HPLC纯度91.6％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-008, and the HPLC purity is 91.6%.
实施例37Example 37
利用载体FL-009非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-009 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-009，HPLC纯度86.5％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-009, and the HPLC purity is 86.5%.
实施例38Example 38
利用载体FL-012非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-012 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-012，HPLC纯度87.0％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-012, and the HPLC purity is 87.0%.
实施例39Example 39
利用载体FL-014非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-014 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-014，HPLC纯度88.0％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-014, and the HPLC purity is 88.0%.
实施例40Example 40
利用载体FL-018非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-018 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-018，HPLC纯度85.1％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-018, and the HPLC purity is 85.1%.
实施例41Example 41
利用载体FL-020非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-020 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-020，HPLC纯度87.5％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-020, and the HPLC purity is 87.5%.
实施例42Example 42
利用载体FL-022非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 by the heterogeneous method of carrier FL-022:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-022，HPLC纯度85.5％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-022, and the HPLC purity is 85.5%.
实施例43Example 43
利用载体FL-023非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-023 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成FL-023，HPLC纯度85.3％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with FL-023, and the HPLC purity is 85.3%.
实施例44Example 44
利用载体FL-027非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH，包括如下步骤：The heterogeneous synthesis of H-Arg-Lys-Gly-Thr-Lys-Ser-OH using carrier FL-027 includes the following steps:
1)载体活化：FL-027溶解于二氯甲烷；加入乙酰溴(FL-027的2-3当量)，常温反应1-3小时；反应结束后减压除去二氯甲烷和过量的乙酰溴；用二氯甲烷重新溶解，再次浓缩除去余下的乙酰溴得到FL-027溴化中间体。1) Support activation: FL-027 is dissolved in dichloromethane; acetyl bromide (2-3 equivalents of FL-027) is added, and reacted at room temperature for 1-3 hours; after the reaction, the dichloromethane and excess acetyl bromide are removed under reduced pressure; Re-dissolved in dichloromethane, concentrated again to remove the remaining acetyl bromide to obtain FL-027 brominated intermediate.
2)接入第一个氨基酸：FL-027溴化中间体溶解于醋酸异丙酯(2-4体积)，加入Fmoc-Ser(t-Bu)-OH(FL-027的1.5-2当量)、二异丙基乙胺(FL-027的1.5-3当量)；混合反应液加热至50-60℃反应6-8h；冷却至室温，加入庚烷(醋酸异丙酯5-10倍体积)；加入DMF和水混合溶液(DMF与水的体积比为(1～8)：1)，搅拌5min，静置分层；分去下层，上层直接用于脱Fmoc反应；2) Connect the first amino acid: FL-027 brominated intermediate is dissolved in isopropyl acetate (2-4 volumes), add Fmoc-Ser(t-Bu)-OH (1.5-2 equivalents of FL-027) , Diisopropylethylamine (1.5-3 equivalents of FL-027); The mixed reaction solution is heated to 50-60°C for 6-8h; cooled to room temperature, add heptane (5-10 times the volume of isopropyl acetate) ; Add a mixed solution of DMF and water (the volume ratio of DMF to water is (1-8):1), stir for 5 minutes, and stand for layering; separate the lower layer, and the upper layer is directly used for the Fmoc reaction;
3)脱Fmoc反应：上一步骤所得上层溶液加热至40-50℃，加入巯基丙酸、二乙烯三胺和DMF混合液(巯基丙酸和二乙烯三胺的用量为FL-027的3～5当量)，保温40-50℃反应。反应完全后加入DMF和水混合溶液(DMF和水的体积比为(1～5)：(5～1))，搅拌5min，静置分层；分去下层，上层加入DMF和水混合溶液(DMF与水的体积比为(1～8)：1)，搅拌5min，静置分层；分去下层，上层重复以上洗涤步骤至洗出液中性，上层直接用于下一个氨基酸缩合；3) Fmoc removal reaction: The upper layer solution obtained in the previous step is heated to 40-50°C, and the mixture of mercaptopropionic acid, diethylenetriamine and DMF is added (the amount of mercaptopropionic acid and diethylenetriamine is 3～3 of FL-027 5 equivalents), and react at 40-50°C. After the reaction is complete, add a mixed solution of DMF and water (the volume ratio of DMF and water is (1～5): (5～1)), stir for 5 minutes, and let stand for layering; divide the lower layer, add the mixed solution of DMF and water ( The volume ratio of DMF to water is (1-8): 1), stir for 5 min, stand still for layering; divide the lower layer, repeat the above washing steps for the upper layer until the eluate is neutral, and use the upper layer directly for the next amino acid condensation;
4)接入第二个氨基酸：上一步骤所得上层溶液，加入Fmoc-Lys(Boc)-OH(FL-027的1～1.5当量)和DIPEA(DIPEA的用量为FL-027的2～3当量)的DMF溶液，搅拌降温至15-20℃；加入HBTU/DMF溶液(其中HBTU的用量为FL-027的1～1.5当量)，保温15-20℃反应。反应完全后，加入水，搅拌5min，静置分层；分去下层，上层加入DMF和水混合溶液(DMF和水的体积比为(1～8)：1)，搅拌5min，静置分层；分去下层，上层直接用于脱Fmoc反应；4) Access to the second amino acid: add Fmoc-Lys(Boc)-OH (1～1.5 equivalent of FL-027) and DIPEA (the amount of DIPEA is 2～3 equivalent of FL-027) to the upper solution obtained in the previous step ) DMF solution, stir and cool to 15-20°C; add HBTU/DMF solution (wherein the amount of HBTU is 1 to 1.5 equivalents of FL-027), keep at 15-20°C for reaction. After the reaction is complete, add water, stir for 5 minutes, stand for layering; divide the lower layer, add a mixed solution of DMF and water (the volume ratio of DMF to water is (1-8):1) in the upper layer, stir for 5 minutes, stand for layering ; Separate the lower layer, and the upper layer is directly used for the Fmoc reaction;
5)重复以上步骤3)和4)依次接入Fmoc-Thr(t-Bu)-OH，Fmoc-Gly-OH，Fmoc-Lys(Boc)-OH，Fmoc-Arg(Pbf)-OH，并脱除Fmoc后得H-Arg(Pbf)-Lys(Boc)-Gly-Thr(t-Bu)-Lys(Boc)-Ser(t-Bu)-(FL-027)庚烷溶液，浓缩得固体；5) Repeat the above steps 3) and 4) to connect Fmoc-Thr(t-Bu)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Arg(Pbf)-OH in sequence, and remove After removing Fmoc, obtain H-Arg(Pbf)-Lys(Boc)-Gly-Thr(t-Bu)-Lys(Boc)-Ser(t-Bu)-(FL-027) heptane solution, which is concentrated to obtain a solid;
6)将以上固体溶解于TFA：TIS：水(94:3:3，裂解液，是固体重量的5-10倍)，常温搅拌2h。缓慢滴加进0-10℃甲基叔丁基醚(甲基叔丁基醚的重量为裂解液的2-20倍)，保温0-10℃搅拌半小时；过滤，滤饼用甲基叔丁基醚洗至洗出液中性，干燥得H-Arg-Lys-Gly-Thr-Lys-Ser-OH粗品，HPLC纯度94.5％，HRMS TOF[M+1] +：676.4102。 6) Dissolve the above solid in TFA:TIS:water (94:3:3, lysate, 5-10 times the weight of the solid), and stir for 2h at room temperature. Slowly add methyl tert-butyl ether at 0-10°C (the weight of methyl tert-butyl ether is 2-20 times of the lysate), keep it at 0-10°C and stir for half an hour; filter, filter cake with methyl tertiary The butyl ether was washed until the eluate was neutral, and dried to obtain crude H-Arg-Lys-Gly-Thr-Lys-Ser-OH, with a HPLC purity of 94.5%, and HRMS TOF[M+1] + : 676.4102.
实施例45Example 45
利用载体FL-028非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH by heterogeneous method using carrier FL-028:
本实施例的合成方法同实施例44，不同之处在于：将实施例44的载体换成FL-028，HPLC纯度95.5％。The synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-028, and the HPLC purity is 95.5%.
实施例46Example 46
利用载体FL-029非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH by heterogeneous method using carrier FL-029:
本实施例的合成方法同实施例44，不同之处在于：将实施例44的载体换成FL-029，HPLC纯度93.5％。The synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-029, and the HPLC purity is 93.5%.
实施例47Example 47
利用载体FL-031非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH by heterogeneous method using carrier FL-031:
本实施例的合成方法同实施例44，不同之处在于：将实施例44的载体换成FL-031，HPLC纯度92.0％。The synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with FL-031, and the HPLC purity is 92.0%.
实施例48Example 48
利用载体FL-002均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-002 homogeneous method:
A)将FL-002(8.2g，10.0mmol)溶解于醋酸异丙酯(100mL)，加入Fmoc-Arg(Pbf)-OH(9.7g，15.0mmol)、HOBt(2.0g，15mmol)、和DIPEA(3.24g，25.0mmol)，搅拌降温至5-10℃，加入HBTU(5.62g，15mmol)进行反应；反应完全后加入乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)，醋酸异丙酯溶液用于下一步脱Fmoc反应。A) Dissolve FL-002 (8.2g, 10.0mmol) in isopropyl acetate (100mL), add Fmoc-Arg(Pbf)-OH (9.7g, 15.0mmol), HOBt (2.0g, 15mmol), and DIPEA (3.24g, 25.0mmol), stir and cool to 5-10℃, add HBTU (5.62g, 15mmol) for reaction; after the reaction is complete, add acetonitrile/water (9:1) to wash (add isopropyl acetate as needed) , The isopropyl acetate solution is used in the next step of the Fmoc removal reaction.
B)步骤1)所得醋酸异丙酯溶液，加入二乙烯三胺(6.2g，60mmol)和巯基丙酸(4.3g，40mmol)，加热至40-50℃反应，TLC分析反应完全后，用乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)除去副产物，醋酸异丙酯溶液用于下一个氨基酸缩合。B) The isopropyl acetate solution obtained in step 1), add diethylenetriamine (6.2g, 60mmol) and mercaptopropionic acid (4.3g, 40mmol), heat to 40-50℃ to react, TLC analysis after the reaction is complete, use acetonitrile /Water (9:1) washing (add isopropyl acetate as needed) to remove by-products, isopropyl acetate solution is used for the next amino acid condensation.
C)按以上方法连续接入Fmoc-Arg(Pbf)-OH，Fmoc-Gln(Trt)-OH，Fmoc-Glu(OtBu)-OH，Fmoc-Met-OH，Fmoc-Glu(OtBu)-OH，并脱除Fmoc后得 H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002)醋酸异丙酯，浓缩得固体；C) Continuously connect to Fmoc-Arg(Pbf)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Met-OH, Fmoc-Glu(OtBu)-OH according to the above method, After removing Fmoc, H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002) isopropyl acetate is obtained, which is concentrated to obtain a solid;
D)按实施32的方法裂解得H-Glu-Met-Glu-Gln-Arg-Arg-NH 2粗品，HPLC纯度70.2％。 D) According to the method of implementation 32, the crude H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 was obtained by cleavage, with an HPLC purity of 70.2%.
实施例49Example 49
利用载体FL-020均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-020 homogeneous method:
本实施例的合成方法同实施例48，不同之处在于：将实施例48的载体换成FL-020，HPLC纯度68.5％。The synthesis method of this embodiment is the same as that of embodiment 48, except that the carrier of embodiment 48 is replaced with FL-020, and the HPLC purity is 68.5%.
实施例50Example 50
利用载体FL-023均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier FL-023 homogeneous method:
本实施例的合成方法同实施例48，不同之处在于：将实施例48的载体换成FL-023，HPLC纯度71.5％。The synthesis method of this embodiment is the same as that of embodiment 48, except that the carrier of embodiment 48 is replaced with FL-023, and the HPLC purity is 71.5%.
实施例51Example 51
利用载体FL-027均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH using carrier FL-027 homogeneous method:
A)载体FL-027(11.5g，10mmol)的活化和第一个氨基酸接入反应同实施例44，反应完全后，用乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)，醋酸异丙酯溶液用于下一步脱Fmoc反应。A) The activation of carrier FL-027 (11.5g, 10mmol) and the first amino acid incorporation reaction are the same as in Example 44. After the reaction is complete, wash with acetonitrile/water (9:1) (add isopropyl acetate as needed) ), the isopropyl acetate solution is used in the next step of the Fmoc removal reaction.
B)所得醋酸异丙酯溶液，加入二乙烯三胺(6.2g，60mmol)和巯基丙酸(4.3g，40mmol)，加热至40-50℃反应，TLC分析反应完全后，用乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)除去副产物，醋酸异丙酯溶液用于下一个氨基酸缩合。B) The obtained isopropyl acetate solution, add diethylenetriamine (6.2g, 60mmol) and mercaptopropionic acid (4.3g, 40mmol), heat to 40-50℃ to react, TLC analysis after the reaction is complete, use acetonitrile/water ( 9:1) Wash (add isopropyl acetate as needed) to remove by-products, and the isopropyl acetate solution is used for the next amino acid condensation.
C)将上一步所得Fmoc反应所得醋酸异丙酯溶液，加入Fmoc-Lys(Boc)-OH(7.0g，15.0mmol)、HOBt(2.0g，15mmol)、和DIPEA(3.24g，25.0mmol)，搅拌降温至5-10℃，加入HBTU(5.62g，15mmol)进行反应；反应完全后加入乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)，醋酸异丙酯溶液用于下一步脱Fmoc反应。C) Add Fmoc-Lys(Boc)-OH (7.0g, 15.0mmol), HOBt (2.0g, 15mmol), and DIPEA (3.24g, 25.0mmol) to the isopropyl acetate solution obtained from the Fmoc reaction obtained in the previous step, Stir and cool to 5-10°C, add HBTU (5.62g, 15mmol) to react; after the reaction is complete, add acetonitrile/water (9:1) to wash (add isopropyl acetate as needed), and isopropyl acetate solution is used The next step is the Fmoc reaction.
D)重复以上步骤2)和步骤3)依次接入Fmoc-Thr(t-Bu)-OH，Fmoc-Gly-OH，Fmoc-Lys(Boc)-OH，Fmoc-Arg(Pbf)-OH，并脱除Fmoc后得H-Arg(Pbf)-Lys(Boc)-Gly-Thr(t-Bu)-Lys(Boc)-Ser(t-Bu)-(FL-0027)醋酸异丙酯溶液，浓缩得固体；D) Repeat steps 2) and 3) above to connect to Fmoc-Thr(t-Bu)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Arg(Pbf)-OH, and After removing Fmoc, obtain H-Arg(Pbf)-Lys(Boc)-Gly-Thr(t-Bu)-Lys(Boc)-Ser(t-Bu)-(FL-0027) isopropyl acetate solution, and concentrate Get a solid
E)按实施例48的方法裂解得H-Arg-Lys-Gly-Thr-Lys-Ser-OH，HPLC纯度82.0％。E) According to the method of Example 48, H-Arg-Lys-Gly-Thr-Lys-Ser-OH was obtained by cleavage, with an HPLC purity of 82.0%.
实施例52Example 52
利用载体FL-031均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH using carrier FL-031 homogeneous method:
实施例的合成方法同实施例51，不同之处在于：将实施例51的载体换成FL-031，HPLC纯度80.0％。The synthesis method of the embodiment is the same as that of embodiment 51, except that the carrier of embodiment 51 is replaced with FL-031, and the HPLC purity is 80.0%.
对比例1Comparative example 1
2-(2，3-二氢植烷氧基)-芴-9-胺(REF-001)的合成：Synthesis of 2-(2,3-dihydrophytaalkoxy)-fluorene-9-amine (REF-001):
Figure PCTCN2019125143-appb-000046
Figure PCTCN2019125143-appb-000046
对比例1中REF-001的合成方法同实施例1，以2，3-二氢植烷基溴代替中间体1-3与2-羟基9-芴酮反应得到中间体R1-2，然后按同样的方法步骤合成REF-001；The synthesis method of REF-001 in Comparative Example 1 is the same as that of Example 1, using 2,3-dihydrophytyl bromide instead of Intermediate 1-3 to react with 2-hydroxy 9-fluorenone to obtain Intermediate R1-2, and then press Synthesize REF-001 in the same way;
1H-NMR(400MHz，CDCl 3)：δ0.80-2.00(m，39H)，4.00-4.10(m，2H)，4.76(s，1H)，6.90-6.97(d，1H)，7.15-7.25(m，2H)，7.30-7.40(m，1H)，7.55-7.65(m，3H)；HRMS TOF[M+1] +：478.4040。 1 H-NMR (400MHz, CDCl 3 ): δ 0.80-2.00 (m, 39H), 4.00-4.10 (m, 2H), 4.76 (s, 1H), 6.90-6.97 (d, 1H), 7.15-7.25 (m, 2H), 7.30-7.40 (m, 1H), 7.55-7.65 (m, 3H); HRMS TOF[M+1] + : 478.4040.
对比例2Comparative example 2
2，7-二(2，3-二氢植烷氧基)-芴-9-胺(REF-002)的合成：Synthesis of 2,7-bis(2,3-dihydrophytoxy)-fluorene-9-amine (REF-002):
Figure PCTCN2019125143-appb-000047
Figure PCTCN2019125143-appb-000047
对比例2中REF-002的合成方法同对比例1，不同之处是：以2,7-二羟基-9-芴酮代替2-羟基-9-芴酮；The synthesis method of REF-002 in Comparative Example 2 is the same as that of Comparative Example 1, except that 2,7-dihydroxy-9-fluorenone is used instead of 2-hydroxy-9-fluorenone;
1H-NMR(400MHz，CDCl 3)：0.8-1.85(m，78H)，3.90-4.00(m，4H)，4.71(s，1H)，6.60-6.70(m，2H)，6.85-6.90(d，2H)，7.10-7.15(s，2H)，7.40-7.50(d，2H)；HRMS TOF[M+1] +：774.7122。 1 H-NMR (400MHz, CDCl 3 ): 0.8-1.85 (m, 78H), 3.90-4.00 (m, 4H), 4.71 (s, 1H), 6.60-6.70 (m, 2H), 6.85-6.90 (d , 2H), 7.10-7.15 (s, 2H), 7.40-7.50 (d, 2H); HRMS TOF[M+1] + : 774.7122.
对比例3Comparative example 3
2-(2，3-二氢植烷氧基)-9-(3-氟苯基)-芴-9-醇(REF-003)的合成：Synthesis of 2-(2,3-dihydrophytaalkoxy)-9-(3-fluorophenyl)-fluorene-9-ol (REF-003):
Figure PCTCN2019125143-appb-000048
Figure PCTCN2019125143-appb-000048
中间体R1-2(9.5g，0.02mol)溶解于无水四氢呋喃(100mL)，常温下滴加格氏试剂(R3-1)(0.1mol)；滴毕，加热回流反应5h；冷却至室温，滴加饱和氯化铵水溶液中止反应；用石油醚(200mL)萃取，石油醚层用水(100mL×2)和饱和食盐水(100mL)洗涤；减压浓缩得粗品，柱层析纯化得REF-003 8.3g；Intermediate R1-2 (9.5g, 0.02mol) was dissolved in anhydrous tetrahydrofuran (100mL), and Grignard reagent (R3-1) (0.1mol) was added dropwise at room temperature; after dripping, the reaction was heated and refluxed for 5h; cooled to room temperature, The reaction was stopped by adding saturated aqueous ammonium chloride solution dropwise; extracted with petroleum ether (200mL), the petroleum ether layer was washed with water (100mL×2) and saturated brine (100mL); concentrated under reduced pressure to obtain crude product, purified by column chromatography to obtain REF-003 8.3g;
1H-NMR(400MHz，CDCl 3)：0.8-1.85(m，39H)，2.80-2.90(m，1H)，3.90-4.00(m，2H)，6.84-6.95(m，3H)，7.00-7.10(m，1H)，7.13-7.29(m，4H)，7.30-7.40(m，1H)，7.55-7.60(m，2H)；HRMS TOF[M-17] +：555.3994，HRMS TOF[M+1] +：573.4100。 1 H-NMR (400MHz, CDCl 3 ): 0.8-1.85 (m, 39H), 2.80-2.90 (m, 1H), 3.90-4.00 (m, 2H), 6.84-6.95 (m, 3H), 7.00-7.10 (m, 1H), 7.13-7.29 (m, 4H), 7.30-7.40 (m, 1H), 7.55-7.60 (m, 2H); HRMS TOF[M-17] + : 555.3994, HRMS TOF[M+1 ] + : 573.4100.
对比例4Comparative example 4
2，7-二(2,3-二氢植烷氧基)-芴-(3-氟苯基)-芴-9-醇(REF-003)的合成：Synthesis of 2,7-bis(2,3-dihydrophytoxy)-fluorene-(3-fluorophenyl)-fluorene-9-ol (REF-003):
Figure PCTCN2019125143-appb-000049
Figure PCTCN2019125143-appb-000049
对比例4中REF-004的合成方法同对比例3，不同之处是：以2,7-二(2,3-二氢植烷氧基)-9-芴酮代替2-二氢植烷氧基-9-芴酮；The synthesis method of REF-004 in Comparative Example 4 is the same as that of Comparative Example 3. The difference is that 2,7-bis(2,3-dihydrophytoxy)-9-fluorenone is used instead of 2-dihydrophytane Oxy-9-fluorenone;
1H-NMR(400MHz，CDCl 3)：0.8-1.85(m，78H)，2.80-2.90(m，1H)，3.90-4.00(m，4H)，6.80-6.90(m，4H)，6.90-7.00(m，1H)，7.10-7.15(m，1H)，7.20-7.30(m，2H)，7.45-7.50(d，2H)；HRMS TOF[M-17] +：852.3582，HRMS TOF[M+1] +：869.7180。 1 H-NMR (400MHz, CDCl 3 ): 0.8-1.85 (m, 78H), 2.80-2.90 (m, 1H), 3.90-4.00 (m, 4H), 6.80-6.90 (m, 4H), 6.90-7.00 (m, 1H), 7.10-7.15 (m, 1H), 7.20-7.30 (m, 2H), 7.45-7.50 (d, 2H); HRMS TOF[M-17] + : 852.3582, HRMS TOF[M+1 ] + : 869.7180.
对比例5Comparative example 5
利用载体REF-001非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 by the heterogeneous method of carrier REF-001:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成REF-001，HPLC纯度73.5％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with REF-001, and the HPLC purity is 73.5%.
对比例6Comparative example 6
利用载体REF-002非均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2： Synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using the carrier REF-002 heterogeneous method:
本实施例的合成方法同实施例32，不同之处在于：将实施例32的载体换成REF-002，HPLC纯度67.8％。The synthesis method of this embodiment is the same as that of embodiment 32, except that the carrier of embodiment 32 is replaced with REF-002, and the HPLC purity is 67.8%.
对比例7Comparative example 7
利用载体REF-003非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH by using carrier REF-003 heterogeneous method:
本实施例的合成方法同实施例44，不同之处在于：将实施例44的载体换成REF-003，HPLC纯度83.0％。The synthesis method of this embodiment is the same as that of embodiment 44, except that the carrier of embodiment 44 is replaced with REF-003, and the HPLC purity is 83.0%.
对比例8Comparative example 8
利用载体REF-004非均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH by heterogeneous method using carrier REF-004:
本实施例的合成方法同实施例44，不同之处在于：将实施例44的载体换成REF-004，HPLC纯度80.5％。The synthesis method of this example is the same as that of Example 44, except that: the carrier of Example 44 is replaced with REF-004, and the HPLC purity is 80.5%.
对比例9Comparative example 9
利用载体REF-001均相法合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 The homogeneous synthesis of H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 using carrier REF-001
A)缩合反应：将REF-001(4.8g，10mmol)溶解于醋酸异丙酯(100mL)，搅拌降温至5-10℃；依次加入Fmoc-Arg(Pbf)-OH(9.8g，15mmol)、二异丙基乙胺(3.25g，25mmol)、HOBt(2.0g，15mmol)、和HBTU(5.62g，15mmol)进行反应；保持5-10℃，反应半小时，自然升到室温反应至反应完全。A) Condensation reaction: Dissolve REF-001 (4.8g, 10mmol) in isopropyl acetate (100mL), stir and cool to 5-10°C; add Fmoc-Arg(Pbf)-OH (9.8g, 15mmol), Diisopropylethylamine (3.25g, 25mmol), HOBt (2.0g, 15mmol), and HBTU (5.62g, 15mmol) are reacted; keep at 5-10℃, react for half an hour, naturally warm to room temperature and react until the reaction is complete .
B)反应完全后加入乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)，TLC分析Fmoc-Arg(Pbf)-OH完全除去后，醋酸异丙酯溶液用于下一步反应。B) After the reaction is complete, add acetonitrile/water (9:1) to wash (add isopropyl acetate as needed), TLC analysis After Fmoc-Arg(Pbf)-OH is completely removed, the isopropyl acetate solution is used for the next reaction .
C)脱Fmoc反应：缩合反应所得醋酸异丙酯溶液，加入二乙烯三烯(6.2g，60mmol)和巯基丙酸(4.3g，40mmol)，加热至40-50℃反应，TLC分析反应完全后，用乙腈/水(9：1)洗涤(根据需要补加醋酸异丙酯)洗除去副产物，醋酸异丙酯溶液用于下一个氨基酸缩合。C) De-Fmoc reaction: the isopropyl acetate solution obtained from the condensation reaction, add diethylenetriene (6.2g, 60mmol) and mercaptopropionic acid (4.3g, 40mmol), heat to 40-50℃ to react, TLC analysis after the reaction is complete , Wash with acetonitrile/water (9:1) (add isopropyl acetate as needed) to remove by-products, and the isopropyl acetate solution is used for the next amino acid condensation.
D)按以上方法连续接入Fmoc-Arg(Pbf)-OH，Fmoc-Gln(Trt)-OH，Fmoc-Glu(OtBu)-OH，Fmoc-Met-OH，Fmoc-Glu(OtBu)-OH，并脱除Fmoc后得H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002)醋酸异丙酯溶液，浓缩得固体；D) Continuously connect to Fmoc-Arg(Pbf)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Met-OH, Fmoc-Glu(OtBu)-OH according to the above method, After removing Fmoc, H-Glu(OtBu)-Met-Glu(OtBu)-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-(FL-002) isopropyl acetate solution is obtained, which is concentrated to obtain a solid;
E)固体按实施例32的方法裂解得H-Glu-Met-Glu-Gln-Arg-Arg-NH 2，HPLC纯度45.0％。 E) The solid was cleaved according to the method in Example 32 to obtain H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 with an HPLC purity of 45.0%.
对比例10Comparative example 10
利用载体REF-003均相法合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH：Synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH using carrier REF-003 homogeneous method:
本实施例的载体活化的方法同实施例44的步骤1)、接第一个氨基酸的方法同实施例44的步骤2)，脱Fmoc和其它氨基酸的接入方法同对比例9，不用之处在于：使用的载体为REF-003，HPLC纯度50.5％。The method of carrier activation in this example is the same as step 1) of Example 44, the method of connecting the first amino acid is the same as step 2) of Example 44, and the method of removing Fmoc and other amino acids is the same as that of Comparative Example 9. It is: the carrier used is REF-003, and the HPLC purity is 50.5%.
测试例1：Test case 1:
将本发明实施例1～31合成的化合物及对比例1～4合成的化合物在室温下(25～30℃)对常用溶剂中的溶解度进行考察，结果如表1所示：The compounds synthesized in Examples 1 to 31 of the present invention and the compounds synthesized in Comparative Examples 1 to 4 were investigated for their solubility in common solvents at room temperature (25-30°C). The results are shown in Table 1:
表1Table 1
载体Carrier 庚烷(％)Heptane (%) i-PrOAc(％)i-PrOAc(%) MTBE(％)MTBE(%) DMF(％)DMF(%) MeOH(％)MeOH(%) DMSO(％)DMSO(%)
FL-001FL-001 >50>50 >50>50 >50>50 >100>100 <10<10 <1<1
FL-002FL-002 >60>60 >100>100 >60>60 >100>100 <5<5 <1<1
FL-003FL-003 >60>60 >100>100 >100>100 >100>100 <1<1 <1<1
FL-004FL-004 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-005FL-005 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-006FL-006 >100>100 >100>100 >100>100 >30>30 <1<1 <1<1
FL-007FL-007 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-008FL-008 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-009FL-009 >100>100 >100>100 >100>100 >30>30 <1<1 <1<1
FL-010FL-010 >100>100 >100>100 >100>100 >30>30 <1<1 <1<1
FL-011FL-011 >80>80 >100>100 >80>80 >100>100 <10<10 <1<1
FL-012FL-012 >100>100 >100>100 >100>100 >100>100 <5<5 <1<1
FL-013FL-013 >100>100 >100>100 >100>100 >50>50 <1<1 <1<1
FL-014FL-014 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-015FL-015 >50>50 >100>100 >50>50 >100>100 <1<1 <1<1
FL-016FL-016 >50>50 >50>50 >50>50 >100>100 <10<10 <1<1
FL-017FL-017 >50>50 >100>100 >50>50 >100>100 <20<20 >30>30
FL-018FL-018 >100>100 >100>100 >100>100 >100>100 <30<30 >100>100
FL-019FL-019 >50>50 >50>50 >50>50 >100>100 <20<20 >30>30
FL-020FL-020 >50>50 >100>100 >100>100 >100>100 <20<20 >10>10
FL-021FL-021 >50>50 >50>50 >50>50 >100>100 <20<20 >10>10
FL-022FL-022 >50>50 >50>50 >50>50 >100>100 <20<20 >10>10
FL-023FL-023 >100>100 >100>100 >100>100 >80>80 <1<1 >20>20
FL-024FL-024 >50>50 >100>100 >50>50 >100>100 <5<5 >10>10
FL-025FL-025 >30>30 >50>50 >30>30 >100>100 <5<5 >20>20
FL-026FL-026 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-027FL-027 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-028FL-028 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-029FL-029 >100>100 >100>100 >100>100 >100>100 <1<1 <1<1
FL-030FL-030 >100>100 >100>100 >100>100 >30>30 <1<1 <1<1
FL-031FL-031 >100>100 >100>100 >100>100 >30>30 <1<1 <1<1
REF-001REF-001 >100>100 >100>100 >100>100 <5<5 <1<1 <1<1
REF-002REF-002 >100>100 >100>100 >100>100 <1<1 <1<1 <1<1
REF-003REF-003 >100>100 >100>100 >100>100 <5<5 <1<1 <1<1
REF-004REF-004 >100>100 >100>100 >100>100 <1<1 <1<1 <1<1
由表1可知：本发明化合物在庚烷、乙酸异丙酯(i-PrOAc)、甲基叔丁基醚(MTBE)、N，N-二甲基甲酰胺(DMF)中均具有较好的溶解度，尤其在DMF中的溶解度远远高于对比例。It can be seen from Table 1 that the compound of the present invention has better properties in heptane, isopropyl acetate (i-PrOAc), methyl tert-butyl ether (MTBE), and N,N-dimethylformamide (DMF). The solubility, especially the solubility in DMF is much higher than the comparative example.
测试例2：Test case 2:
对使用本发明化合物作为载体利用非均相反应合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2六肽(实施例32-43)和合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH六肽(实施例44-47)、对比例1-2的化合物作为载体利用非均相反应合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2六肽(对比例5-6)和对比例3-4的化合物作为载体利用非均相反应合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH六肽(对比例7-8)，及使用本发明化合物作为载体利用均相反应合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2六肽(实施例48-50)和合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH六肽(实施例51-52)、对比例1的化合物作为载体利用均相反应合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2六肽(对比例9)、对比例3的化合物作为载体利用均相反应合成H-Arg-Lys-Gly-Thr-Lys-Ser-OH六肽的反应时间、纯度进行统计，结果如表2所示，其中，AA 1为第一个氨基酸，AA 2为第二个氨基酸，AA 3为第三个氨基酸，AA 4为第四个氨基酸，AA 5为第五个氨基酸，AA 6为第六个氨基酸，t R为 缩合反应时间，t W为缩合反应后处理时间，t D为去保护反应时间，t Dw为去保护反应后处理时间，单位：小时； To use the compound of the present invention as a carrier, use heterogeneous reaction to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 hexapeptide (Example 32-43) and synthesize H-Arg-Lys-Gly-Thr- Lys-Ser-OH hexapeptide (Examples 44-47) and the compound of Comparative Example 1-2 were used as carriers to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 hexapeptide (for The compounds of Examples 5-6) and Comparative Examples 3-4 were used as carriers to synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH hexapeptide (Comparative Examples 7-8) using heterogeneous reaction, and use the present invention The compound is used as a carrier to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 hexapeptide (Examples 48-50) and synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH using homogeneous reaction Hexapeptide (Examples 51-52) and the compound of Comparative Example 1 were used as carriers to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 hexapeptide (Comparative Example 9) and the compound of Comparative Example 3 using a homogeneous reaction The compound is used as a carrier to synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH hexapeptide by a homogeneous reaction. The reaction time and purity are counted. The results are shown in Table 2, where AA 1 is the first amino acid. AA 2 is the second amino acid, AA 3 is the third amino acid, AA 4 is the fourth amino acid, AA 5 is the fifth amino acid, AA 6 is the sixth amino acid, t R is the condensation reaction time, t W is The post-treatment time of the condensation reaction, t D is the deprotection reaction time, t Dw is the post-treatment time of the deprotection reaction, unit: hour;
表2Table 2
Figure PCTCN2019125143-appb-000050
Figure PCTCN2019125143-appb-000050
Figure PCTCN2019125143-appb-000051
Figure PCTCN2019125143-appb-000051
Figure PCTCN2019125143-appb-000052
Figure PCTCN2019125143-appb-000052
*表示均相法反应，其余为非均相反应。* Indicates a homogeneous reaction, the rest are heterogeneous reactions.
由表2可知，在非均相体系中，本发明化合物作为载体进行H-Glu-Met-Glu-Gln-Arg-Arg-NH 2六肽合成时，相对于对比例REF-001～002，其缩合反应时间，缩合反应后处理时间、去保护反应时间、去保护反应后处理时间明显缩短；其中，对比例REF-001～002的缩合反应时间大于2小时，缩合反应后处理时间大于1小时，去保护反应时间大于2.5小时，去保护反应后处理时间大于2小时；而本发明化合物的缩合反应时间为0.5～1.0小时，缩合反应后处理时间为0.5～1.0小时，去保护反应时间为0.5～1.0小时，去保护反应后处理时间为0.5～1.0小时，每个氨基酸片断接入步骤所需时间相差很小，重复性好；而对比例REF-001～002每个氨基酸片断接入步骤所需时间相差很大。 It can be seen from Table 2 that in a heterogeneous system, when the compound of the present invention is used as a carrier to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 hexapeptide, compared with the comparative examples REF-001～002, Condensation reaction time, condensation reaction post-treatment time, deprotection reaction time, and deprotection post-treatment time are significantly shortened; among them, the condensation reaction time of Comparative Examples REF-001～002 is more than 2 hours, and the post-condensation treatment time is more than 1 hour. The deprotection reaction time is more than 2.5 hours, and the post-deprotection treatment time is more than 2 hours; and the condensation reaction time of the compound of the present invention is 0.5-1.0 hours, the post-condensation treatment time is 0.5-1.0 hours, and the deprotection reaction time is 0.5-1.0 hours. 1.0 hour, the processing time after deprotection reaction is 0.5～1.0 hours, the time required for each amino acid fragment access step is very small, and the repeatability is good; and the comparative example REF-001～002 requires each amino acid fragment access step The time varies greatly.
本发明化合物作为载体进行H-Arg-Lys-Gly-Thr-Lys-Ser-OH六肽合成时，在非均相体系相对于对比例REF-003～004也取得一样的结果：缩合反应时间，缩合反应后处理时间、去保护反应时间、去保护反应后处理时间明显缩短，其中在肽链延长阶段(AA 2-AA 5)每个氨基酸的接入时间变化非常小，缩合、去保护和两个后处理时间都是0.5～1.0h，重复性好。 When the compound of the present invention is used as a carrier to synthesize H-Arg-Lys-Gly-Thr-Lys-Ser-OH hexapeptide, the same result is obtained in the heterogeneous system compared to the comparative examples REF-003～004: the condensation reaction time, The processing time after the condensation reaction, the deprotection reaction time, and the post-processing time of the deprotection reaction are significantly shortened. The access time of each amino acid in the peptide chain elongation stage (AA 2 -AA 5 ) changes very little. Each post-treatment time is 0.5～1.0h, with good repeatability.
此外，对比例REF-001～004的化合物在合成肽的过程中，溶解度下降较快，后处理过程中需要不断补加溶剂才可保持溶液状态，从AA 4开始出现凝胶化现象，而本发明化合物在多肽合成中，溶解度基本没有变化，具有较好的溶解度；本发明化合物作为载体在非均相体系中合成肽的产物，其中碳端为酰胺的肽纯度大于85％，部分大于90％，碳端为羧基的肽纯度大于90％，部分大于95％，而对比例碳端为酰胺REF-001～REF-002的产品纯度不高于75％，碳端为羧基的REF-003～REF-004纯度不高于85％。 In addition, the solubility of the compounds in Comparative Examples REF-001～004 decreased rapidly during the process of peptide synthesis. During the post-treatment process, the solvent needs to be continuously added to maintain the solution state, and the gelation phenomenon began to appear from AA 4 . In the synthesis of peptides, the solubility of the compound of the present invention is basically unchanged and has a good solubility; the compound of the present invention is used as a carrier to synthesize peptides in a heterogeneous system, and the purity of the peptide with an amide at the carbon end is greater than 85%, and some are greater than 90%. The purity of the peptides with carboxyl at the carbon end is greater than 90%, and some are greater than 95%, while the purity of the products with amide REF-001～REF-002 at the carbon end of the comparative example is not higher than 75%, and the purity of the product with carboxyl at the carbon end of REF-003～REF -004 purity is not higher than 85%.
并且，当本发明含有芴环结构的化合物为FL-005、FL-008、FL-014、FL-028和FL-029时，作为载体合成肽的缩合反应时间、缩合反应后处理时间、去保护反应时间、去保护反应后处理时间更短，其产品纯度更高。In addition, when the compounds of the present invention containing a fluorene ring structure are FL-005, FL-008, FL-014, FL-028, and FL-029, the condensation reaction time, post-condensation treatment time, and deprotection of peptide synthesis as a carrier The reaction time and the post-treatment time of the deprotection reaction are shorter, and the product purity is higher.
使用均相体系合成H-Glu-Met-Glu-Gln-Arg-Arg-NH 2和H-Arg-Lys-Gly-Thr-Lys-Ser-OH时，使用本发明化合物作为载体的缩合反应时间也明显缩短，一般小于5h，而对比载体一般都大于5h，部分氨基酸10h，并且使用本发明化合物作为载体产品纯度也明显较高。 When using a homogeneous system to synthesize H-Glu-Met-Glu-Gln-Arg-Arg-NH 2 and H-Arg-Lys-Gly-Thr-Lys-Ser-OH, the condensation reaction time using the compound of the present invention as a carrier is also Obviously shortened, generally less than 5h, while the comparison carrier is generally greater than 5h, some amino acids are 10h, and the product purity of the product using the compound of the invention as a carrier is also significantly higher.
综上，使用本发明化合物作为载体，在均相或非均相体系中均适用，且相对于对比例中的化合物均具有更优的效果，同时本发明的化合物在非均相体系中比在均相体系中具有更好的效果，特别是在中低极性溶剂和酰胺溶剂形成的两相体系中(非均相体系)，进行肽的合成，能显著缩短缩合反应时间、缩合反应后处理时间、去保护反应时间、去保护反应后处理时间，最终得到纯度较高的产品。In summary, using the compound of the present invention as a carrier is suitable for both homogeneous and heterogeneous systems, and has better effects than the compounds in the comparative example. At the same time, the compounds of the present invention are more effective than those in heterogeneous systems. It has better effect in homogeneous system, especially in the two-phase system (heterogeneous system) formed by medium and low polar solvents and amide solvents, for peptide synthesis, it can significantly shorten the condensation reaction time and post-condensation treatment Time, deprotection reaction time, and post-treatment time of deprotection reaction, and finally obtain a product with higher purity.

Claims (17)

  1. 一种含有芴环结构的化合物，其特征在于：所述化合物的结构如通式(1)所示：A compound containing a fluorene ring structure, characterized in that the structure of the compound is as shown in the general formula (1):
    Figure PCTCN2019125143-appb-100001
    Figure PCTCN2019125143-appb-100001
    其中：among them:
    X选自OH、卤素、磺酸酯、NHR a； X is selected from OH, halogen, sulfonate, NHR a ;
    Y选自氢、芳基；Y is selected from hydrogen and aryl;
    其中，R a选自氢、烷基或芳烷基； Wherein, R a is selected from hydrogen, alkyl or aralkyl;
    Q独立选自O、NH、NHCO、CO、CONH、S、SO或SO 2； Q is independently selected from O, NH, NHCO, CO, CONH, S, SO or SO 2 ;
    n 1，n 2独立选自0～3的整数，且n 1、n 2不同时为0； n 1 and n 2 are independently selected from an integer of 0 to 3, and n 1 and n 2 are not 0 at the same time;
    R独立选自通式(2)所示的基团：R is independently selected from the group represented by the general formula (2):
    Figure PCTCN2019125143-appb-100002
    Figure PCTCN2019125143-appb-100002
    其中，*表示与Q连接；Among them, * means connected with Q;
    R 1选自C 1～C 25的烷基或通式(3)所示的基团： R 1 is selected from a C 1 ～C 25 alkyl group or a group represented by the general formula (3):
    Figure PCTCN2019125143-appb-100003
    Figure PCTCN2019125143-appb-100003
    其中，*表示与羰基连接；Wherein, * means it is connected to the carbonyl group;
    m1表示1～3的整数；m1 represents an integer of 1 to 3;
    R 1a选自C 6～C 25的烷基，且m1个R 1a的总碳数不少于8； R 1a is selected from C 6 -C 25 alkyl groups, and the total carbon number of m1 R 1a is not less than 8;
    k a选自0～3的整数； k a is selected from an integer of 0 to 3;
    环B除了具有m1个R 1aO取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基、不含卤素原子取代的C 1～C 5烷氧基的取代基； Ring B except that one R 1a O having m1 substituent groups, but may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ~ C 5 alkoxy group, a halogen atom free of C 1 ~ C 5 alkoxy group substituent;
    通式(2)中，R 2选自氢、C 1～C 25的烷基或通式(4)所示的基团： In the general formula (2), R 2 is selected from hydrogen, a C 1 to C 25 alkyl group or the group represented by the general formula (4):
    Figure PCTCN2019125143-appb-100004
    Figure PCTCN2019125143-appb-100004
    通式(4)中，*表示与N连接；In the general formula (4), * means connected with N;
    m2选自0～3的整数；m2 is selected from an integer of 0 to 3;
    R 2a选自C 6～C 25的烷基； R 2a is selected from C 6 ～C 25 alkyl groups;
    k b选自1～6的整数； k b is selected from an integer of 1 to 6;
    环C除了具有m2个R 2aO取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基、不含卤素原子取代的C 1～C 5烷氧基的取代基； In addition to an outer ring C m2 having a substituent group R 2a O, may also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, a halogen-containing atoms substituted with C 1 ~ C 5 alkoxy group, a halogen atom free of C 1 ~ C 5 alkoxy group substituent;
    L选自含有O、N或S杂原子的C 2～C 15有机链基或不含有O、N或S杂原子的C 2～C 15有机链基，当L选自不含有O、N或S杂原子的C 2～C 15有机链基时，R 2≠H； L is selected from C 2 ～C 15 organic chain groups containing O, N or S heteroatoms or C 2 ～C 15 organic chain groups without O, N or S heteroatoms, when L is selected from C 2 ～C 15 organic chain groups that do not contain O, N or When the C 2 ～C 15 organic chain group of S heteroatom, R 2 ≠H;
    芴环除了具有n 1和n 2个RQ取代基外，还可以含有选自卤素原子、含有卤素原子取代的C 1～C 5烷基、不含卤素原子取代的C 1～C 5烷基、含有卤素原子取代的C 1～C 5烷氧基或不含卤素原子取代的C 1～C 5烷氧基的取代基。 In addition to having a fluorene ring n 1 and n 2 th RQ substituents, they can also contain selected from halogen atoms, substituted with a halogen atom-containing C 1 ~ C 5 alkyl group, a halogen atom free of C 1 ~ C 5 alkyl group, substituent group substituted with halogen atom-containing C 1 ~ C 5 alkoxy group or a halogen atom free of C 1 ~ C 5 alkoxy.
  2. 根据权利要求1所述的含有芴环结构的化合物，其特征在于：所述R选自通式(5)所示的基团：The compound containing a fluorene ring structure according to claim 1, wherein the R is selected from the group represented by the general formula (5):
    Figure PCTCN2019125143-appb-100005
    Figure PCTCN2019125143-appb-100005
    其中，通式(5)中，*表示与Q连接；Among them, in the general formula (5), * means connected with Q;
    k 1选自0～3的整数，并且当k 1＝0时，R 2≠H。 k 1 is selected from an integer of 0 to 3, and when k 1 =0, R 2 ≠H.
  3. 根据权利要求1所述的含有芴环结构的化合物，其特征在于：所述R选自通式(6)所示的基团：The compound containing a fluorene ring structure according to claim 1, wherein the R is selected from the group represented by the general formula (6):
    Figure PCTCN2019125143-appb-100006
    Figure PCTCN2019125143-appb-100006
    其中，通式(6)中，*表示与Q连接；Among them, in the general formula (6), * means connected with Q;
    R 4选自氢、C 1～C 25的烷基或权利要求1中通式(4)所示的基团； R 4 is selected from hydrogen, C 1 to C 25 alkyl group or the group represented by general formula (4) in claim 1;
    k 2选自1～4的整数； k 2 is selected from an integer of 1 to 4;
    k 3选自1～4的整数。 k 3 is selected from an integer of 1-4.
  4. 根据权利要求1所述的含有芴环结构的化合物，其特征在于：所述R选自通式(7)所示的基团：The compound containing a fluorene ring structure according to claim 1, wherein the R is selected from the group represented by the general formula (7):
    Figure PCTCN2019125143-appb-100007
    Figure PCTCN2019125143-appb-100007
    其中，通式(7)中，*表示与Q连接；Among them, in the general formula (7), * means connected with Q;
    R 6选自氢、C 1～C 25的烷基或权利要求1中通式(4)所示的基团； R 6 is selected from hydrogen, C 1 to C 25 alkyl group or the group represented by general formula (4) in claim 1;
    k 4选自0～3的整数； k 4 is selected from an integer from 0 to 3;
    k 5选自0～3的整数； k 5 is selected from an integer from 0 to 3;
    R 5选自氢、天然氨基酸的侧链基、烷基或通式(8)所示的基团： R 5 is selected from hydrogen, a side chain group of a natural amino acid, an alkyl group or a group represented by the general formula (8):
    Figure PCTCN2019125143-appb-100008
    Figure PCTCN2019125143-appb-100008
    其中，*表示连接位点；Among them, * represents the connection site;
    k 6选自1～4的整数； k 6 is selected from an integer of 1 to 4;
    R 1＇选自C 1～C 25的烷基或权利要求1中通式(3)所示的基团； R 1 ′ is selected from C 1 ～C 25 alkyl group or the group represented by general formula (3) in claim 1;
    R 2＇选自氢、C 1～C 25的烷基或权利要求1中通式(4)所示的基团。 R 2 'is selected from hydrogen, C 1 to C 25 alkyl group, or the group represented by the general formula (4) in claim 1.
  5. 根据权利要求1～4任意一项所述的含有芴环结构的化合物，其特征在于：所述Q选自O；优选地，所述R a选自氢、甲基、乙基、丙基、苄基或甲氧苄基；优选地，所述n 1+n 2等于1～3。 The compound containing a fluorene ring structure according to any one of claims 1 to 4, wherein: said Q is selected from O; preferably, the R a is selected from hydrogen, methyl, ethyl, propyl, Benzyl or methoxybenzyl; preferably, n 1 + n 2 is equal to 1-3.
  6. 根据权利要求1～4任意一项所述的含有芴环结构的化合物，其特征在于：所述m1选自2或3，m1个R 1a的总碳数为12～60；优选地，所述R 1a选自C 8～C 22的烷基。 The compound containing a fluorene ring structure according to any one of claims 1 to 4, wherein the m1 is selected from 2 or 3, and the total carbon number of m1 R 1a is 12 to 60; preferably, the R 1a is selected from C 8 to C 22 alkyl groups.
  7. 根据权利要求1～4任意一项所述的含有芴环结构的化合物，其特征在于：所述m2选自2或3，m2个R 2a的总碳数为12～60；优选地，所述R 2a选自C 8～C 22的烷基。 The compound containing a fluorene ring structure according to any one of claims 1 to 4, wherein the m2 is selected from 2 or 3, and the total carbon number of m2 R 2a is 12 to 60; preferably, the R 2a is selected from C 8 to C 22 alkyl groups.
  8. 根据权利要求1～4任意一项所述的含有芴环结构的化合物，其特征在于：所述R 2选自氢、甲基、乙基、丙基、异丙基、异辛基、苄基或4-甲氧苄基。 The compound containing a fluorene ring structure according to any one of claims 1 to 4, wherein the R 2 is selected from hydrogen, methyl, ethyl, propyl, isopropyl, isooctyl, benzyl Or 4-methoxybenzyl.
  9. 根据权利要求1～4任意一项所述的含有芴环结构的化合物，其特征在于：所述含有芴环结构的化合物的结构式选自如下：The compound containing a fluorene ring structure according to any one of claims 1 to 4, wherein the structural formula of the compound containing a fluorene ring structure is selected from the following:
    Figure PCTCN2019125143-appb-100009
    Figure PCTCN2019125143-appb-100009
    Figure PCTCN2019125143-appb-100010
    Figure PCTCN2019125143-appb-100010
    Figure PCTCN2019125143-appb-100011
    Figure PCTCN2019125143-appb-100011
  10. 根据权利要求1所述的含有芴环结构的化合物，其特征在于：所述含有芴环结构的化合物易溶于烃类有机溶剂、芳香烃类有机溶剂、酯类有机溶剂、醚类有机溶剂、水溶性非质子类极性有机溶剂中的至少一种；优选地，所述烃类有机溶剂选自庚烷、己烷、石油醚、环己烷、甲基环己烷中的至少一种；优选地，所述芳香烃类有机溶剂选自甲苯、乙苯、二甲苯中的至少一种；优选地，所述酯类有机溶剂选自乙酸异丙酯、乙酸叔丁酯、 乙酸乙酯中的至少一种；优选地，所述醚类有机溶剂选自***、异丙醚、甲基叔丁基醚、甲基环戊基醚、四氢呋喃中的至少一种；优选地，所述水溶性非质子类极性有机溶剂选自N,N-二甲基甲酰胺、N,N-二乙基甲酰胺、N,N-二甲基乙酰胺、N-甲基-吡咯烷酮、N-乙基-吡咯烷酮、二甲基亚砜、环丁砜、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种。The compound containing a fluorene ring structure according to claim 1, wherein the compound containing a fluorene ring structure is easily soluble in hydrocarbon organic solvents, aromatic hydrocarbon organic solvents, ester organic solvents, ether organic solvents, At least one of the water-soluble aprotic polar organic solvents; preferably, the hydrocarbon organic solvent is selected from at least one of heptane, hexane, petroleum ether, cyclohexane, and methylcyclohexane; Preferably, the aromatic hydrocarbon organic solvent is selected from at least one of toluene, ethylbenzene, and xylene; preferably, the ester organic solvent is selected from isopropyl acetate, tert-butyl acetate, and ethyl acetate. Preferably, the ether organic solvent is selected from at least one of diethyl ether, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, and tetrahydrofuran; preferably, the water-soluble Non-protic polar organic solvent is selected from N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, N-methyl-pyrrolidone, N-ethyl -At least one of pyrrolidone, dimethyl sulfoxide, sulfolane, 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone .
  11. 根据权利要求10所述的含有芴环结构的化合物，其特征在于：25～30℃时，所述含有芴环结构的化合物在N,N-二甲基甲酰胺中的溶解度＞1％。The compound containing a fluorene ring structure according to claim 10, wherein the solubility of the compound containing the fluorene ring structure in N,N-dimethylformamide is >1% at 25-30°C.
  12. 一种氨基酸或肽C端的保护试剂，其特征在于：所述保护试剂包含权利要求1～11任一项所述的含有芴环结构的化合物。A protective reagent for the C-terminus of an amino acid or peptide, characterized in that the protective reagent comprises the compound containing a fluorene ring structure according to any one of claims 1 to 11.
  13. 权利要求1～11任一项所述的含有芴环结构的化合物在均相或非均相溶剂体系中合成肽试剂中的应用。Use of the compound containing a fluorene ring structure according to any one of claims 1 to 11 in the synthesis of peptide reagents in a homogeneous or heterogeneous solvent system.
  14. 一种肽的合成方法，其特征在于：使用权利要求1～11任一项所述的含有芴环结构的化合物；优选地，包括如下步骤：A method for synthesizing peptides, characterized by using the compound containing a fluorene ring structure according to any one of claims 1 to 11; preferably, it comprises the following steps:
    1)载体接入：将权利要求1～11任一项所述的含有芴环结构的化合物作为载体与N-保护氨基酸或N-保护肽化合物通过常规反应进行连接得到含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物；1) Carrier access: The compound containing the fluorene ring structure of any one of claims 1 to 11 is used as a carrier to connect with the N-protected amino acid or N-protected peptide compound through a conventional reaction to obtain the C-terminus of the carrier containing the fluorene ring structure Protected N-protected amino acid or N-protected peptide compound;
    2)N端去保护：将含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶解于溶剂中，加入N端保护的脱保护试剂溶液形成均相或非均相体系进行N端脱保护，加入高极性溶剂进行萃取，得含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液；2) N-terminal deprotection: Dissolve the N-protected amino acid or N-protected peptide compound containing the C-terminal protection of the fluorene ring structure carrier in a solvent, and add the N-terminal protection deprotection reagent solution to form a homogeneous or heterogeneous system. N-terminal deprotection, adding a highly polar solvent for extraction, to obtain N-deprotected amino acid or N-deprotected peptide compound solution containing fluorene ring structure carrier C-terminal protection;
    3)肽链延长：在含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液中，加入N-保护氨基酸或N-保护肽溶液，后加入缩合试剂溶液形成均相或非均相体系进行缩合反应，加入高极性溶剂进行萃取，得到含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶液；3) Peptide chain extension: Add N-protected amino acid or N-protected peptide solution to the C-terminal protected N-deprotected amino acid or N-deprotected peptide compound solution of the fluorene ring structure carrier, and then add the condensation reagent solution to form a homogeneous solution. Condensation reaction in a phase or heterogeneous system, and a highly polar solvent is added for extraction to obtain an N-protected amino acid or N-protected peptide compound solution containing the C-terminal protection of the fluorene ring structure carrier;
    4)重复步骤2)和步骤3)接入下一个氨基酸，直至得到完整的肽链。4) Repeat step 2) and step 3) to insert the next amino acid until a complete peptide chain is obtained.
  15. 根据权利要求14所述的合成方法，其特征在于：步骤2)中的含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物溶液与步骤2)中的N端保护的脱保护试剂溶液成非均相体系；优选地，步骤3)中的含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液与步骤3)中的N-保护氨基酸或N-保护肽、缩合试剂溶液成非均相体系；优选地，步骤2)中溶解含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽 化合物的溶剂和步骤3)中含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液中的溶剂独立选自烃类或烃类与酯类、醚类、卤代烃中的至少一种形成的混合溶剂；优选地，步骤2)溶解含有芴环结构载体C端保护的N-保护氨基酸或N-保护肽化合物的溶剂和步骤3)中含有芴环结构载体C端保护的N-去保护氨基酸或N-去保护肽化合物溶液中的溶剂独立选自己烷、环己烷、甲基环己烷、庚烷、石油醚或己烷、环己烷、甲基环己烷、庚烷、石油醚中至少一种与醋酸异丙酯、乙酸叔丁酯、乙酸乙酯、***、异丙醚、甲基叔丁醚、甲基环戊基醚、二氯甲烷、氯仿中的至少一种形成的混合溶剂中的至少一种；优选地，步骤2)中溶解N端保护的脱保护试剂的溶剂、步骤3)中溶解N-保护氨基酸或N-保护肽、缩合试剂的溶剂独立选自酰胺类溶剂；优选地，所述酰胺类溶剂选自N,N-二甲基甲酰胺、N,N-二乙基甲酰胺、N,N-二甲基乙酰胺、N-甲基-吡咯烷酮、N-乙基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮中的至少一种。The synthesis method according to claim 14, characterized in that: the N-protected amino acid or N-protected peptide compound solution containing the C-terminal protection of the fluorene ring structure carrier in step 2) and the deprotection of the N-terminal protection in step 2) The protective reagent solution forms a heterogeneous system; preferably, the N-deprotected amino acid or N-deprotected peptide compound solution containing the C-terminal protection of the fluorene ring structure carrier in step 3) and the N-protected amino acid in step 3) or The N-protected peptide and the condensation reagent solution form a heterogeneous system; preferably, in step 2), the solvent containing the C-terminal protected N-protected amino acid or N-protected peptide compound of the fluorene ring structure carrier is dissolved and the step 3) contains fluorene The solvent in the solution of the N-deprotected amino acid or N-deprotected peptide compound protected at the C-terminus of the ring structure carrier is independently selected from hydrocarbons or a mixed solvent formed by hydrocarbons and at least one of esters, ethers, and halogenated hydrocarbons Preferably, step 2) dissolve the solvent containing the N-protected amino acid or N-protected peptide compound protected at the C-terminus of the fluorene ring structure carrier and step 3) contains the N-deprotected amino acid or N-deprotected amino acid protected at the C-terminus of the fluorene ring structure carrier -The solvent in the solution of the deprotected peptide compound is independently selected from at least hexane, cyclohexane, methylcyclohexane, heptane, petroleum ether or hexane, cyclohexane, methylcyclohexane, heptane, and petroleum ether. A mixed solvent formed with at least one of isopropyl acetate, tert-butyl acetate, ethyl acetate, diethyl ether, isopropyl ether, methyl tert-butyl ether, methyl cyclopentyl ether, dichloromethane, and chloroform Preferably, the solvent for dissolving the N-terminal protected deprotection reagent in step 2), the solvent for dissolving the N-protected amino acid or N-protected peptide, and the condensation reagent in step 3) are independently selected from amide solvents; Preferably, the amide solvent is selected from N,N-dimethylformamide, N,N-diethylformamide, N,N-dimethylacetamide, N-methyl-pyrrolidone, N-ethyl At least one of pyrrolidone, 1,3-dimethylimidazolinone, and 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone.
  16. 根据权利要求14所述的合成方法，其特征在于：步骤2)和步骤3)所述的高极性溶剂选自水、醇类、腈类、酰胺类、亚砜类、砜类、水溶性醇醚类中的至少一种；优选地，步骤2)和步骤3)所述的高极性溶剂选自水、甲醇、乙腈、N,N-二甲基甲酰胺、N-甲基-吡咯烷酮、1,3-二甲基咪唑啉酮、1,3-二甲基-3,4,5,6-四氢-2-嘧啶酮、二甲基亚砜、环丁砜中的至少一种。The synthesis method according to claim 14, characterized in that: the highly polar solvents in step 2) and step 3) are selected from water, alcohols, nitriles, amides, sulfoxides, sulfones, water-soluble At least one of alcohol ethers; preferably, the highly polar solvent described in step 2) and step 3) is selected from water, methanol, acetonitrile, N,N-dimethylformamide, and N-methyl-pyrrolidone , At least one of 1,3-dimethylimidazolinone, 1,3-dimethyl-3,4,5,6-tetrahydro-2-pyrimidinone, dimethylsulfoxide, and sulfolane.
  17. 根据权利要求14～16任意一项所述的合成方法，其特征在于：所述N-保护氨基酸或N-保护肽的用量为所述含有芴环结构的化合物的0.8～3.0当量，所述缩合试剂的用量为所述含有芴环结构的化合物的0.8～3.0当量；优选地，所述N-保护氨基酸或N-保护肽的用量为所述含有芴环结构的化合物的1～1.1当量，所述缩合试剂的用量为所述含有芴环结构的化合物的1～1.2当量。The synthesis method according to any one of claims 14-16, wherein the amount of the N-protected amino acid or N-protected peptide is 0.8-3.0 equivalents of the compound containing the fluorene ring structure, and the condensation The dosage of the reagent is 0.8-3.0 equivalents of the compound containing the fluorene ring structure; preferably, the dosage of the N-protected amino acid or N-protected peptide is 1-1.1 equivalents of the compound containing the fluorene ring structure. The amount of the condensation reagent is 1 to 1.2 equivalents of the compound containing the fluorene ring structure.
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Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2415744A1 (en) * 2009-03-12 2012-02-08 Ajinomoto Co., Inc. Fluorene compound
CN103080058A (en) * 2010-08-30 2013-05-01 味之素株式会社 Aromatic compound containing specific branch
EP2716650A1 (en) * 2011-05-31 2014-04-09 Ajinomoto Co., Inc. Method for producing peptide
CN107406480A (en) * 2015-03-04 2017-11-28 Jitsubo株式会社 Peptide symthesis method
WO2018034901A1 (en) * 2016-08-16 2018-02-22 Stealth Biotherapeutics N-carboxyanhydride-based-scale synthesis of elamipretide
CN108715605A (en) * 2018-06-06 2018-10-30 安徽工程大学 A kind of chemical synthesis process of antibacterial cyclic peptides Thermoactinoamide A
CN110256277A (en) * 2019-03-19 2019-09-20 广州同隽医药科技有限公司 A kind of compound containing fluorenes ring structure and its application
WO2019198834A1 (en) * 2018-04-13 2019-10-17 Jitsubo株式会社 Method for producing leuprorelin
WO2019198833A1 (en) * 2018-04-13 2019-10-17 Jitsubo株式会社 Peptide synthesis method

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
HUE056740T2 (en) * 2016-06-20 2022-03-28 Sekisui Medical Co Ltd Novel diphenylmethane protective agent
TW201833127A (en) * 2017-02-27 2018-09-16 日商積水醫療股份有限公司 Novel tetracyclic protection agent
CN109678751A (en) * 2019-01-07 2019-04-26 广州同隽医药科技有限公司 A kind of compound containing diphenyl-methane structure

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2415744A1 (en) * 2009-03-12 2012-02-08 Ajinomoto Co., Inc. Fluorene compound
CN103080058A (en) * 2010-08-30 2013-05-01 味之素株式会社 Aromatic compound containing specific branch
EP2716650A1 (en) * 2011-05-31 2014-04-09 Ajinomoto Co., Inc. Method for producing peptide
CN107406480A (en) * 2015-03-04 2017-11-28 Jitsubo株式会社 Peptide symthesis method
WO2018034901A1 (en) * 2016-08-16 2018-02-22 Stealth Biotherapeutics N-carboxyanhydride-based-scale synthesis of elamipretide
WO2019198834A1 (en) * 2018-04-13 2019-10-17 Jitsubo株式会社 Method for producing leuprorelin
WO2019198833A1 (en) * 2018-04-13 2019-10-17 Jitsubo株式会社 Peptide synthesis method
CN108715605A (en) * 2018-06-06 2018-10-30 安徽工程大学 A kind of chemical synthesis process of antibacterial cyclic peptides Thermoactinoamide A
CN110256277A (en) * 2019-03-19 2019-09-20 广州同隽医药科技有限公司 A kind of compound containing fluorenes ring structure and its application

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
TAKAHASHI, DAISUKE ET AL.: "Development of an Efficient Liquid-Phase Peptide Synthesis Protocol Using a Novel Fluorene-Derived Anchor Support Compound with Fmoc Chemistry; AJIPHAS", TETRAHEDRON LETTERS, vol. 53, no. 15, 9 February 2012 (2012-02-09), XP028466059, DOI: 20200310191033 *

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