CN105566431B - A kind of impurity of CDB-2914 and its preparation and detection method - Google Patents
A kind of impurity of CDB-2914 and its preparation and detection method Download PDFInfo
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- CN105566431B CN105566431B CN201610091024.1A CN201610091024A CN105566431B CN 105566431 B CN105566431 B CN 105566431B CN 201610091024 A CN201610091024 A CN 201610091024A CN 105566431 B CN105566431 B CN 105566431B
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- 0 CC1([C@]2(CCC3C4CC5)O)OCC*1C23C=C=C4C(CC1)=C5CC11OCCO1 Chemical compound CC1([C@]2(CCC3C4CC5)O)OCC*1C23C=C=C4C(CC1)=C5CC11OCCO1 0.000 description 3
- NNRHYMTXYVBQML-JCRBPUCRSA-N CC(CC1C(C=C2)=CCC2N(C)C)(C(CC2)C(CC3)C1=C(CC1)C3=CC1=O)[C@H]2C(C)=O Chemical compound CC(CC1C(C=C2)=CCC2N(C)C)(C(CC2)C(CC3)C1=C(CC1)C3=CC1=O)[C@H]2C(C)=O NNRHYMTXYVBQML-JCRBPUCRSA-N 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07J—STEROIDS
- C07J41/00—Normal steroids containing one or more nitrogen atoms not belonging to a hetero ring
- C07J41/0033—Normal steroids containing one or more nitrogen atoms not belonging to a hetero ring not covered by C07J41/0005
- C07J41/005—Normal steroids containing one or more nitrogen atoms not belonging to a hetero ring not covered by C07J41/0005 the 17-beta position being substituted by an uninterrupted chain of only two carbon atoms, e.g. pregnane derivatives
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
Abstract
The present invention provides a kind of impurity compound of CDB-2914 and preparation method thereof, specifically, noval chemical compound impurity F (compound F) and impurity K (compound K) are provided, and the detection method of the impurity and its purposes of the quality analysis for CDB-2914 bulk drug and its preparation are also provided.
Description
Technical field
The present invention provides a kind of impurity compound and its preparation and detection method of CDB-2914, and described is contaminated
Compound is applied to the quality analysis of CDB-2914 bulk drug and its preparation.Belong to analytical chemistry field, more particularly to medicine
Analytical chemistry field.
Background technology
CDB-2914 (Ulipristal Acetate；Compound X；Chemical name：17 α-the β of acetoxyl group-11-[4-
(N, N- dimethylamino) phenyl] -19- norpregna -4,9- diene -3,20- diketone) it is a kind of potent antiprogestin and anti-
Glucocorticoid medicine.Structural formula is as follows：
CDB-2914 has listed in Europe and U.S.'s approval as emergency contraception, for unshielded sexual life or
Used after knowing or suspecting contraceptive failure in 5 days, be a kind of effective and safe emergency contraception.Emergency contraception is defined as preventing
The treatment method of pregnancy caused by unprotect measure sexual behaviour, it is a kind of to prevent from surprisingly cherishing caused by the sexual behaviour of unprotect measure
Pregnant important method.It is as it is accidental, urgent when a standby measure using.
As fibroid curative, in Europe, approval has listed CDB-2914, in the preoperative therapy women of child-bearing age
Spend severe fibroid.Fibroid (also known as leiomyoma of uterus) be it is benign, monoclonal, the uterus of hormone-sensitive is put down
Sliding Muscle neoplasms.It is the most common tumour of the premenopausal genital tract of women, it was reported that Women of childbearing age 20-40% suffers from this tumour.Son
Palace myomata is often asymptomatic, but when there is symptom, and initial symptom is serious uterine hemorrhage, anaemia, and belly is pressurized, belly
Pain, frequent micturition aggravates and infertility.Especially, the reduction of serious blood volume is the symptom of most common fibroid.
Prior art CN102516345B disclose can industrialized production CDB-2914 method, be be easy to get 3,
(ethylenedioxy -19- norpregnas -5 (10), 9 (11)-diene -3,17- diketone (compound II) add 3- with cyanylation agent
Obtain compound III into reaction, then by 17 Alpha-hydroxies protect, obtain formula IV compound, then with lithium methide or methyl form
Formula V compound is obtained through sour water solution after reagent reacting, then the double contractings of 3,20- are obtained through the catalysis such as p-methyl benzenesulfonic acid and glycol reaction
Assimilation compound VI, then epoxidation reaction obtain epoxy material VII, then obtain compound VIII with grignard reagent reacting, then pass through
Hydrolyzed under acidic conditions obtains compound IX, most obtains CDB-2914 (compound X) (Scheme 1) through acetoxylation afterwards.Should
Method synthetic route is short, and totally 8 steps are reacted, and reaction condition is gentle, it is easier to carries out, total recovery about 25-27% or so, and grasps
Work is easier, is suitable for industrialization.
Any material for influenceing pharmaceutical purity is referred to as impurity.Miscellaneous Quality Research is an important content of drug research and development,
It includes selecting suitable analysis method, differentiates exactly and the content of measure impurity and comprehensive pharmacy, toxicity and clinical research
Result determine the reasonable limit of impurity.Whole process of this research through drug research and development.Impurity in medicine is managed by it
Change property and be generally divided into three classes：Organic impurities, inorganic impurity and residual solvent.According to its source, it is miscellaneous that impurity can be divided into technique
Matter (including the complete reactant of unreacted and reagent, intermediate, accessory substance etc. in synthesis), catabolite, from reactant and examination
Mixed impurity etc. in agent.According to its toxicity category, impurity can be divided into toxic impurities and common impurities etc. again.Impurity can also be by it
Classification of chemical structure, such as steroidal, alkaloid, geometric isomer, optical isomer and polymer.Organic impurities is included in technique
Impurity and catabolite of introducing etc., it may be possible to known or unknown, volatile or fixedness.Due to this kind of impurity
Chemical constitution it is typically similar with active component or tool original relationship, therefore generally again can be referred to as relevant material.Analysis method
Selection is directly connected to the specificity and accuracy of impurity determination result, and therefore, when carrying out impurity research, matter of utmost importance is choosing
Select suitable impurity analysis method.
Impurity of the drug Inspection and analysis method should be sensitive, exclusive.The impurity of different structure is entered by suitable analytical technology
Row separation, detection, so as to reach effective control to impurity.It is efficient, quick with separation, the development and renewal of detection technique
Isolation technics be combined with detection means sensitive, stably, accurately, applicable, almost all of organic impurities can be suitable
Under conditions of obtain well separation and detection.In quality standard, the method for detecting impurities generally used at present is mainly height
Effect liquid phase chromatogram method (High Performance Liquid Chromatography；HPLC), thin-layered chromatography (Thin
Layer Chromatography；TLC), gas chromatography (Gas Chromatography；) and capillary electrophoresis GC
(Capillary Electrophoresis；CE).In recent years, mass-spectrometric technique is applied increasingly extensive in terms of impurity of the drug analysis,
Gas-chromatography combination technology, liquid chromatogram GC-MS have become the important means of impurity of the drug analysis.
The content of the invention
In order to better control over the drug quality of CDB-2914 bulk drug and its preparation, the present invention is in bulk drug
Impurity is studied, and obtains related impurities compound, and provide its preparation method and detection method.It is miscellaneous prepared by the present invention
Matter compound, as impurity reference substance, available for the quality analysis of the CDB-2914 bulk drug and its preparation, while also may be used
To provide reference for the selection of process conditions, be advantageous to the control of production process Quality Evaluation of Chinese Medicinal amount.
Technical solution of the present invention is as follows：
The present invention provides a kind of compound or its salt, the structure such as following formula: compound F of the compound or compound K institute
Show：
Compound or its salt described above, wherein the salt is inorganic acid salt or acylate.
As another object of the present invention, also provide a kind of preparation method of compound or its salt described above, it include from
It is prepared by the double ketals (intermediate VI) of 3,20- in the reaction product of CDB-2914 and the chemical combination is prepared through separation
Thing F or the step of compound K.Preferably, it is described that CDB-2914 is prepared by the double ketals (intermediate VI) of 3,20-
Course of reaction is as follows：
Preparation method described above, wherein：
The epoxidation reaction is：
For example, compound VI is dissolved in dichloromethane, under alkalescence condition, adds oxidant and perhalogeno acetone is anti-at room temperature
Should.Preferably, the alkali is selected from pyridine, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, disodium hydrogen phosphate, sodium dihydrogen phosphate etc.；Oxidant
Selected from hydrogen peroxide, metachloroperbenzoic acid etc., preferably hydrogen peroxide；- 10-10 DEG C of reaction temperature.
Described RMgBr reacts：
For example, in THF, stannous chloride catalysis is lower and 4- (dimethylamino of N, N mono-) phenyl-magnesium-bromide RMgBr is carried out
Addition；Preferably, reaction raw materials and RMgBr mol ratio 1:1.5-5, -10-40 DEG C of reaction temperature, reaction time 2-8h.
Described hydrolysis is：
For example, reaction is hydrolyzed under being stirred in dichloromethane and diluted acid, it is preferable that acid is selected from hydrochloric acid, sulfuric acid, sulfuric acid
The inorganic acids such as hydrogen sodium, preferably 0.2-4N HCl solutions, -10-50 DEG C of reaction temperature, reaction time 1-5h.
Described acetylization reaction is：
For example, with carrying out acylation reaction under acylating agent any or two or more in glacial acetic acid, perchloric acid or aceticanhydride,
It is preferred that add glacial acetic acid in perchloric acid and aceticanhydride；- 40-25 DEG C of reaction temperature, preferably -10-25 DEG C；The amount of glacial acetic acid accounts for ice vinegar
The 1-50%V/V, preferably 10-15%V/V for the acetylation reagent volume that acid, perchloric acid, aceticanhydride are formed.
As another embodiment of the present invention, the preparation method of another compound or its salt described above is also provided, its
Including institute is prepared through separation from being prepared by the double ketals (intermediate VI) of 3,20- in the reaction product of CDB-2914
The step of stating compound F or compound K.The reaction of CDB-2914 is prepared in 3,20- double ketals (intermediate VI), such as
May refer to disclosed in prior art CN102516345B can industrialized production CDB-2914 method.
Preparation method described above, wherein described separation is prepared as preparing high pressure liquid chromatography separation；Preferably, institute
The preparation high pressure liquid chromatography condition stated is：Stationary phase is C18 reverse phase fillers, and mobile phase is triethylamine phosphoric acid solution-acetonitrile ladder
Degree elution, Detection wavelength 302nm.
Preferably, preparation method described above, preferably from the recrystallization mother liquor obtained by CDB-2914 recrystallization
Described compound F or compound K is prepared in middle separation.
As another object of the present invention, a kind of compound or its salt described above or compound D or E detection are also provided
Method, wherein：
Wherein methods described is HPLC methods or HPLC-MS methods；
Preferably, the testing conditions of described HPLC methods are：Stationary phase using octadecylsilane chemically bonded silica as filler,
Mobile phase is triethylamine phosphoric acid solution-acetonihile gradient elution, Detection wavelength 302nm；Preferably, mobile phase be pH2.5~
3.5 triethylamine phosphoric acid solution-acetonitrile, volume ratio 30: 70~50: 50；It is highly preferred that mobile phase is pH3.0 triethylamine
Phosphoric acid solution-acetonitrile, volume ratio 40: 60；
Preferably, the testing conditions of described HPLC-MS methods are：HPLC chromatographic condition is stationary phase with octadecyl silicon
Alkane bonded silica gel is filler, and mobile phase is acetonitrile：0.1% formic acid water=50：50, MS condition is to be examined using selective ion
Survey method, detection compound D parent ions [M+H]+M/z 462.2, compound E parent ion [M+H]+M/z 434.0, compound mother F from
Sub [M+H]+M/z 518.2, or compound K parent ion [M+H]+m/z 492.3。
As another object of the present invention, also provide compound or its salt described above in CDB-2914 bulk drug or
Purposes in the relevant material detection of preparation as impurity reference substance.
As another object of the present invention, a kind of quality testing side of CDB-2914 bulk drug or preparation is also provided
Method, it includes the step of measure is wherein about material, and described relevant material is selected from compound F, K, D or E described above,
It is characterized in that described measure is detected using HPLC methods or HPLC-MS methods；Preferably, described HPLC methods or HPLC-
MS methods are as described above；It is highly preferred that described detection is detected using HPLC methods；It is highly preferred that the detection of described HPLC methods
Condition is：Stationary phase is octadecylsilane chemically bonded silica, and mobile phase is pH2.5~3.5 that volume ratio is 30: 70~50: 50
Triethylamine phosphoric acid solution-acetonitrile, Detection wavelength is DAD (PDAD) 302nm, and number of theoretical plate is by acetic acid crow profit
2000 should be not less than by taking charge of his peak and calculating；It is highly preferred that in described relevant material detection, in the chromatogram of need testing solution if any
Impurity peaks, with calculated by peak area, peak corresponding with each impurities phase respectively cannot be greater than 0.15 times of (i.e. impurity of impurity reference substance solution
0.15%) content is no more than.
Purposes described above, wherein described detection is detected using HPLC methods or HPLC-MS methods；Preferably, it is described
Detection using HPLC methods detect；It is highly preferred that described HPLC method testing conditions are：Stationary phase is bonded for octadecylsilane
Silica gel, mobile phase are the triethylamine phosphoric acid solution-acetonitrile for pH2.5~3.5 that volume ratio is 30: 70~50: 50, are more preferably flowed
It is mutually 0.2% triethylamine (phosphoric acid adjusts pH 3.0) buffer solution：Acetonitrile=40:60, Detection wavelength is DAD (Diode Array Detectors
Device) 302nm, number of theoretical plate is calculated by CDB-2914 peak should be not less than 2000；It is highly preferred that described relevant material detection
In, if any impurity peaks in the chromatogram of need testing solution, with calculated by peak area, peak corresponding with each impurities phase respectively cannot be greater than miscellaneous
0.15 times (i.e. impurity content is no more than 0.15%) of matter reference substance solution.Total impurities content must not exceed 1.0%.
As an embodiment of the invention, impurity compound D, E, F or K described above specifically can be by following
Method is prepared.
1st, the preparation process of CDB-2914
By the double ketals (compound VI 1mo1) of 3,20-, dichloromethane, pyridine, three water Hexafluoro acetones, -10-0 is cooled to
DEG C, 50% hydrogen peroxide is added dropwise, reacts 2-3h in -5-5 DEG C, separates organic phase, aqueous phase is extracted twice merging with dichloromethane, uses
10% sodium thiosulfate solution is washed, and organic phase is washed twice, and anhydrous magnesium sulfate is dried, and filtering, is concentrated under reduced pressure into constant weight.
By Mg (1.6mo1), 1,2 one Bromofume and THF add three-necked flask, be added dropwise the bromo- DMAs of 4- and
THF, backflow dropwise addition after initiation reaction is heated, is finished, is obtained RMgBr in 50-60 DEG C of stirring reaction 3h, be cooled to 25 DEG C, add
Enter stannous chloride, after 25 DEG C are stirred 30 minutes, cool down the lower dichloromethane solution that above-mentioned epoxides is added dropwise, keeping temperature 10-
20 DEG C, insulated and stirred 2h is finished, is poured into the saturation NH4Cl aqueous solution of ice, stirs 10 minutes, separates organic phase, aqueous phase is with two
Chloromethanes extracts, and organic phase merges, and adds 2N HCl solutions.2h is stirred at 25 DEG C, separates organic phase, aqueous phase dichloromethane
Extraction, merge, respectively washed once with saturated sodium bicarbonate aqueous solution, water successively, anhydrous magnesium sulfate is dried, and filtering, filtrate is transferred to instead
Answer in bottle, add glacial acetic acid (1.25mol), be cooled to -10-0 DEG C, add 70% perchloric acid (1.18mo1), under then stirring
Aceticanhydride (5.96mo1) is added dropwise in -10-0 DEG C, after 30 minutes, aqueous phase is extracted insulated and stirred with dichloromethane, is merged, successively with full
And sodium bicarbonate aqueous solution, water are respectively washed once, anhydrous magnesium sulfate is dried, filtering, is concentrated under reduced pressure into dry about 110g, through being measured with 10 times
Isopropanol:Ethanol (95:5) Light yellow crystals 244.8g is recrystallized to give, solid is done to obtain in recrystallization mother liquor concentration, stand-by.Product
Isopropanol is used again:Ethanol (95:5) recrystallize, 60 DEG C of drying obtain the product of Light yellow crystals, and recrystallization mother liquor concentration is done
Solid, it is stand-by.
The CDB-2914 of above-mentioned gained is analyzed through LC-MS, containing multiple impurity, including impurity F (i.e. compound
F) and impurity K (i.e. compound K) MS testing results are：Impurity A：[M+H]+397.16, impurity B：[M+H]+415.1 impurity C：
[M+H]+373.1 impurity D：[M+H]+462.2, impurity E：[M+H]+434.0, impurity F：[M+H]+518.2 impurity G：[M+H]+
476.2 impurity H：[M+H]+441.2 impurity I：[M+H]+516.2 impurity J：[M+H]+474.2 impurity K：[M+H]+492.3。
LC-MS conditions used are：
HPLC：Mobile phase is the ︰ 50 of formic acid waters of Yi Jing ︰ 0.1%=50
25~35 DEG C of column temperature
Flow velocity 0.3ml/min
Detection wavelength 302nm
Chromatographic column:Agilent Poroshe11120EC-C18 2.7um 4.6X 50mm
MS：Condition
ESI, fg=25, MS2scan.
2nd, impurity D, E, F, K preparation process
To be analyzed through HPLC, the content of CDB-2914 lmpurities is less than 0.1%, but through to the mother during synthesis
Liquid is analyzed, and the content of wherein impurity is higher, therefore uses preparative separation impurity in the mother liquor treated from building-up process.
By in the preparation process of CDB-2914 gained recrystallization mother liquor be concentrated to dryness to obtain solid, in this solid contain compared with
Impurity at high proportion, this solid be can obtain into high-purity impurity reference substance through the purifying of high pressure preparation liquid phase, processing, concrete operations are such as
Under:
Purification condition:
Gradient elution ratio:
Concrete operation step:
Mother liquor concentrations are obtained into solid to dry, take about 1g, are dissolved with 40% acetonitrile/water, upper prop, gradient elution, according to going out
Peak situation, Fractional Collections eluent, every time purifying after the completion of, purification column with 5% acetonitrile solution rebalancing after, repeat into
Sample purifies.By the multiple batches of purity containing each component for purifying and obtaining>98% eluent merges respectively.Concentration removes most of second
Nitrile, aqueous layer with ethyl acetate extraction, concentration ethyl acetate respectively obtain impurity solid to doing.Impurity A can be obtained per Batch purification about
For 5mg, impurity B is about 10mg, and impurity C is about 5mg, and impurity D is about 15mg, and impurity E is about 5mg, and impurity F is about 5mg, impurity
G is about 5mg, and impurity H is about 15mg, and impurity I is about 5mg, and impurity J is about 10mg, and impurity K is about 5mg.
Analysis testing conditions in above-mentioned purge process are:
Mobile phase is pH3.0 triethylamine phosphate buffers: acetonitrile=40: 60
Detection wavelength 302nm
Column temperature:30℃
Flow velocity 1.0mL/min
As an embodiment of the invention, impurity F, K are additionally provided in CDB-2914 raw material and its preparation
Purposes in product quality detection method.It is preferred that with HPLC and/or HPLC-MS checked for impurities F or K and its content.
It is highly preferred that condition is used by the HPLC,
Mobile phase：Triethylamine phosphoric acid solution-acetonitrile of pH2.5~3.5, volume ratio 30: 70~50: 50；Wherein pH2.5
For~3.5 triethylamine phosphoric acid solution by triethylamine-phosphoric acid buffer to forming, 1000 parts of water add 1~3 part of triethylamine to obtain three second
Amine aqueous solution, using phosphorus acid for adjusting pH as 2.5~3.5；
Flow velocity:0.8~1.2mL/min (preferable flow rate 1.0mL/min)；
Column temperature:25~35 degrees Celsius, preferably 30 DEG C；
Detection wavelength:Using multi-wavelength detection, select respectively each impurity maximum absorption wavelength carry out relevant material detection,
Checking.CDB-2914：303nm；Impurity A：259nm；Impurity B：234nm；Impurity C：241nm；Impurity D：303nm；Impurity
E：304nm；Impurity F：328nm；Impurity G：253nm；Impurity H：262nm；Impurity I：256nm；Impurity J：261nm；Impurity K：
299nm。
Stationary phase：Using octadecylsilane chemically bonded silica as filler
More a step is preferable, and condition is used by the HPLC-MS：
Chromatographic condition is:Mobile phase:0.05%~0.5%HAc/ water:Acetonitrile=(75~80):(25~20)；It is preferred that
0.1%HAc/ water, with acetonitrile ratio preferably 78:22
Flow velocity：0.1~0.5ml/min, preferably 0.3ml/min；Column temperature：20~40 DEG C, preferably 30 DEG C
Mass Spectrometry Conditions are:Using selected ion monitoring ((SIM) method
Dry temperature degree:300~400 DEG C, preferably 350 DEG C
Flow rate of carrier gas:8~12L/min, preferably 10L/min
Atomization gas pressure:30~50psi, preferably 40psi
Capillary voltage:4000V
As a preferred embodiment of the invention, there is provided above-mentioned impurity F or K are as impurity reference substance in acetic acid crow profit
The application taken charge of in the quality analysis of the Related substances separation of he and its preparation, wherein described CDB-2914 and its impurity
Detection method is preferably HPLC methods, and HPLC testing conditions are as follows：
Stationary phase:Using octadecylsilane chemically bonded silica as filler
Mobile phase：PH3.0 triethylamine phosphoric acid solution-acetonitrile, volume ratio 40: 60；Wherein pH3.0 triethylamine phosphorus
Acid solution is by triethylamine-phosphoric acid buffer to forming, and 1000 parts of water add 2 parts of triethylamines, using phosphorus acid for adjusting pH as 3.0；
Flow velocity:0.8~1.2mL/min (preferable flow rate 1.0mL/min)；
Column temperature:25~35 degrees Celsius, preferably 30 DEG C；
Detection wavelength:It is preferred that 302nm.
Number of theoretical plate is calculated by CDB-2914 peak should be not less than 2000
Detection wavelength:UV scanning shows that each impurity and principal component maximum absorption wavelength are as follows：
CDB-2914：303nm；Impurity A：259nm；Impurity B：234nm；Impurity C：241nm；Impurity D：303nm；It is miscellaneous
Matter E：304nm；Impurity F：328nm；Impurity G：253nm；Impurity H：262nm；Impurity I：256nm；Impurity J：261nm；Impurity K：
299nm, consider, using multi-wavelength detection, select the maximum absorption wavelength of each impurity to carry out relevant material detection, test respectively
Card.
Experimental implementation:
CDB-2914 reference substance is weighed respectively and impurity A, B, C, D, E, F, G, H, I, J, K reference substance are each appropriate, is added
Flowing phased soln and dilution are made in every 1ml contains CDB-2914, impurity A, B, C, D, E, F, G, H, I, J, K respectively about respectively
20ug mixed solution, as system suitability solution.Precision measures system suitability solution 10ul injection liquid phases
Chromatograph, chromatogram is recorded, CDB-2914 and the mutual separating degree of each impurity peaks should meet the requirements, and number of theoretical plate presses vinegar
Sour Ulipristal peak, which calculates, should be not less than 2000.
Determination method
Precision weighs this product 10mg, puts in 20ml measuring bottles, with flowing phased soln, and is diluted to scale, shakes up, as trying
Product solution.Precision measures need testing solution 1ml and put in 100ml measuring bottles, is diluted to scale with mobile phase, shakes up, molten as compareing
Liquid.The μ l of contrast solution 10 are measured, inject liquid chromatograph, regulation detection sensitivity makes the peak height of principal component chromatographic peak be full scale
10-20%.Precision measures the μ l of need testing solution 10 again, liquid chromatograph is injected, when record chromatogram to principal component peak retains
Between 3 times.
Each impurity primary study Detection wavelength：
| Wavelength | Impurity |
| 302nm | Impurity D, E, F, K |
| 237nm | Impurity B, C |
| 258nm | Impurity A, G, H, I, J |
If any impurity peaks in the chromatogram of need testing solution, with calculated by peak area, peak corresponding with each impurities phase respectively must not
More than 0.15 times (i.e. impurity content is no more than 0.15%) of contrast solution；Other any single impurity peaks cannot be greater than compareing
0.15 times (i.e. impurity content is no more than 0.15%) of solution, total impurities must not exceed 1.0, and (i.e. total impurities content must not exceed
1.0%), the peak less than contrast solution main peak area 0.05 times (i.e. content is less than 0.05%) is ignored.
The advantages of method of detection of the above-mentioned CDB-2914 about material provided by the invention is that measurement result is accurate
Reliably, specificity is strong, practical, and each impurity can be detected effectively, and reaches good point between CDB-2914 and each impurity
From using the detectable plurality of impurities of the system.
The Related substance of the equally applicable CDB-2914 preparation of the above method.
Brief description of the drawings:
Fig. 1：CDB-2914 and impurity separating degree collection of illustrative plates
Fig. 2：CDB-2914 detects collection of illustrative plates
Fig. 3：Impurity D HPLC measure linear relationship charts
Fig. 4：The HPLC measure linear relationship charts of impurity E
Fig. 5：The HPLC measure linear relationship charts of impurity F
Fig. 6：Impurity K HPLC measure linear relationship charts
Embodiment
Embodiment 1：The preparation of CDB-2914
By the double ketals (100g, 0.25mo1) of 3,20-, dichloromethane (1L), pyridine (5m1), three water Hexafluoro acetones (20m1,
143mmol), -10-0 DEG C are cooled to, 50% hydrogen peroxide (70ml, 1.44mol) is added dropwise, reacting 2-3h raw materials in -5-5 DEG C disappears
Afterwards, organic phase is separated, aqueous phase is extracted twice merging with dichloromethane, and one is washed with 10% sodium thiosulfate solution (10m1)
Secondary, organic phase is washed twice (50m1*2), and anhydrous magnesium sulfate is dried, and filtering, is concentrated under reduced pressure into constant weight about 200m1.
By Mg (9.6g, 0.4mo1), 1,2- Bromofume (1ml) and THF (50m1) add three-necked flask, and it is bromo- that 4- is added dropwise
DMA (71g, 0.35mo1) and THF (200m1) solution, lower dropwise addition is maintained the reflux for after heating initiation reaction, is added
Finish, RMgBr is obtained in 50-60 DEG C of stirring reaction 3h, be cooled to 25 DEG C, add stannous chloride (3g, 30mmol), 25 DEG C are stirred
After mixing 30 minutes, the lower dichloromethane solution that above-mentioned epoxides is added dropwise is cooled down, 10-20 DEG C of keeping temperature, finishes insulated and stirred
2h, pour into the NH of the 500m1 saturations of ice4In the Cl aqueous solution, stir 10 minutes, separate organic phase, aqueous phase is extracted with dichloromethane
400m1*5 times, organic phase merges, and adds 1000m12N HCl solutions.2h is stirred at 25 DEG C, separates organic phase, aqueous phase is with two
Chloromethanes extracts 200m1*3, merges organic phase, is respectively washed once with saturated sodium bicarbonate aqueous solution, water successively, anhydrous magnesium sulfate is done
Dry, filtering, filtrate is transferred in reaction bulb, adds glacial acetic acid (18m1,315mmol), is cooled to -10~0 DEG C, adds 70% high chlorine
Sour (24m1,295mmo1), aceticanhydride (140m1, l.49mo1), insulated and stirred 30 minutes is added dropwise in -10~0 DEG C under then stirring
Afterwards, aqueous phase is extracted twice 200m1*3 with dichloromethane, merges, is respectively washed once with saturated sodium bicarbonate aqueous solution, water successively, nothing
Water magnesium sulfate is dried, filtering, is concentrated under reduced pressure into dry about 110g, through measuring isopropanols with 10 times:Ethanol (95:5) it is recrystallized to give shallow
Yellow crystal 61.2g, solid is done to obtain in recrystallization mother liquor concentration, stand-by.Product uses isopropanol again:Ethanol (95:5) recrystallize,
60 DEG C of drying obtain the product of Light yellow crystals, and solid is done to obtain in recrystallization mother liquor concentration, stand-by.
Embodiment 2：The preparation of impurity compound
By in the preparation process of CDB-2914 gained recrystallization mother liquor be concentrated to dryness to obtain solid, in this solid contain compared with
Impurity at high proportion, this solid be can obtain into high-purity impurity reference substance through the purifying of high pressure preparation liquid phase, processing,
Concrete operations are as follows:
Purification condition:
Gradient elution ratio:
| Time (divides) | A% | B% |
| 0 | 90 | 10 |
| 30 | 40 | 60 |
| 60 | 40 | 60 |
Concrete operation step:
Mother liquor concentrations are obtained into solid to dry, take about 1g, are dissolved with 40% acetonitrile/water, upper prop, gradient elution, according to going out
Peak situation, Fractional Collections eluent, every time purifying after the completion of, purification column with 5% acetonitrile solution rebalancing after, repeat into
Sample purifies.By the multiple batches of purity containing each component for purifying and obtaining>98% eluent merges respectively.Concentration removes most of second
Nitrile, aqueous layer with ethyl acetate extraction, concentration ethyl acetate respectively obtain impurity solid to doing.Impurity A can be obtained per Batch purification about
For 5mg, impurity B is about 10mg, and impurity C is about 5mg, and impurity D is about 15mg, and impurity E is about 5mg, and impurity F is about 5mg, impurity
G is about 5mg, and impurity H is about 15mg, and impurity I is about 5mg, and impurity J is about 10mg, and impurity K is about 5mg.Wherein impurity D and E are
Known compound.Wherein, gained noval chemical compound impurity F (i.e. compound F), K (i.e. compound K) structure and known compound
Impurity D and E structure, it is respectively：
The impurity F, molecular formula：C32H39NO5
Molecular weight：517.67
Chemical name：3,17 α-the β of diacetoxy-11-(4-N, N- dimethylaminophenyl)-19- norpregnas-3,5
(10), 9 (11)-triolefin -20- ketone
MS：518.27 [M+H]+, 540.26 [M+Na]+
INSTRUMENT MODEL：BRUKER AV-500 type NMRs
It is as shown in the table that 1H-NMR spectrums, 13C-NMR spectrums, DEPT spectrums, COSY spectrums, hsqc spectrum and the HMBC of impurity F compose ownership
The impurity K, molecular formula：C30H37NO5
Molecular weight：491.63
Chemical name：17 α-the β of acetoxyl group-11-(11)-two of (4-N, N- dimethylaminophenyl)-19- norpregnas-4,9
Alkene -3,20- diketone-N- oxides
MS：475.30 [M+H-OH]+, 492.30 [M+H]+, 514.27 [M+Na]+
INSTRUMENT MODEL：BRUKER AV-500 type NMRs
It is as shown in the table that impurity K 1H-NMR spectrums, 13C-NMR spectrums, DEPT spectrums, COSY spectrums, hsqc spectrum and HMBC composes ownership
Embodiment 3：The detection method of CDB-2914 and its impurity
The method of detection of the CDB-2914 about material uses liquid chromatographic detection, and HPLC testing conditions are as follows:
Stationary phase：Using octadecylsilane chemically bonded silica as filler
Mobile phase：0.2% triethylamine (phosphoric acid adjusts pH 3.0) buffer solution：Acetonitrile=40:60
Detection wavelength：DAD 302nm、258nm、237nm
Flow velocity：1.0ml/min；
Column temperature：25℃
Experimental implementation：
CDB-2914 reference substance is weighed respectively and each impurity reference substance is each appropriate, adds flowing phased soln and dilution is made
Per the mixed solution containing CDB-2914, each about 20u g of each impurity respectively in 1ml, as system suitability solution.Essence
It is close to measure system suitability solution 10ul injection liquid chromatographs, record chromatogram (as shown in Figure 1), CDB-2914
And the mutual separating degree of each impurity peaks should meet the requirements, number of theoretical plate is calculated by CDB-2914 peak should be not less than 2000.
Determination method:
Precision weighs CDB-2914 10mg, puts in 20ml measuring bottles, with flowing phased soln, and is diluted to scale, shakes up,
As need testing solution.Precision measures need testing solution 1ml and put in 100ml measuring bottles, is diluted to scale with mobile phase, shakes up, and makees
For contrast solution.The μ l of contrast solution 10 are measured, inject liquid chromatograph, regulation detection sensitivity makes the peak height of principal component chromatographic peak
For the 10-20% of full scale.Precision measures the μ l of need testing solution 10 again, injects liquid chromatograph, record chromatogram to principal component
3 times of peak retention time, chromatogram is as shown in Figure 2.
Each impurity primary study Detection wavelength：
| Wavelength | Impurity |
| 302nm | Impurity D, E, F, K |
| 237nm | Impurity B, C |
| 258nm | Impurity A, G, H, I, J |
If any impurity peaks in the chromatogram of need testing solution, with calculated by peak area, peak corresponding with each impurities phase respectively must not
More than 0.15 times (i.e. impurity content is no more than 0.15%) of contrast solution；Other any single impurity peaks cannot be greater than compareing
0.15 times (i.e. impurity content is no more than 0.15%) of solution, total impurities must not exceed 1.0 times, and (i.e. total impurities content is no more than
1.0%), the peak less than contrast solution main peak area 0.05 times (i.e. content is less than 0.05%) is ignored.
Method validation：
Impurity determination Method validation result is as follows：
Claims (4)
1. a kind of detection method of compound, the compound is compound F：
Compound F：
It is characterized in that methods described is HPLC methods or HPLC-MS methods, the testing conditions of described HPLC methods are：Stationary phase with
Octadecylsilane chemically bonded silica is filler, and mobile phase is triethylamine phosphoric acid solution-acetonitrile of pH2.5~3.5, and volume ratio is
30: 70~50: 50, Detection wavelength 302nm；The testing conditions of described HPLC-MS methods are：HPLC chromatographic condition is fixation
Mutually using octadecylsilane chemically bonded silica as filler, mobile phase is acetonitrile：0.1% formic acid water=50：50, MS condition is to adopt
With selective HPLC-ION, detection compound F parent ions [M+H]+m/z518.2。
2. detection method according to claim 1, wherein described mobile phase is pH3.0 triethylamine phosphoric acid solution-second
Nitrile, volume ratio 40: 60.
3. the quality determining method of a kind of CDB-2914 bulk drug or preparation, it is included according to described in claim 1 or 2
The step of method measure is wherein about material.
4. quality determining method according to claim 3, wherein in described relevant material detection, the color of need testing solution
If any impurity peaks in spectrogram, with calculated by peak area, peak corresponding with each impurities phase respectively cannot be greater than impurity reference substance solution
0.15 times, i.e. impurity content is no more than 0.15%.
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Application publication date: 20160511 Assignee: Pharmaceutical Co., Ltd., Changzhou Pharmaceutical Factory No.4 Assignor: Changzhou City No.4 Pharmaceutical Factory Co., Ltd. Contract record no.: 2018320000315 Denomination of invention: Ulipristal acetate impurities and preparation and detection method thereof Granted publication date: 20171124 License type: Exclusive License Record date: 20181114 |
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