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CN-122017052-A - Isolation and measurement of enzam Lu An Z3And methods for genotoxic impurities thereof

CN122017052ACN 122017052 ACN122017052 ACN 122017052ACN-122017052-A

Abstract

The invention belongs to the technical field of medicine analysis, and particularly relates to a method for separating and measuring enzate Lu An Z 3 and genotoxic impurities thereof. The impurities include one or more of impurity X d , impurity X g , impurity X h . The method comprises the steps of adopting octadecylsilane chemically bonded silica as a chromatographic column filling agent, adopting trifluoroacetic acid solution as a mobile phase A and acetonitrile as a mobile phase B, performing linear gradient elution of high performance liquid chromatography, and entering a detector for detection. And then judging whether the content of each impurity in the sample is qualified or not by adopting a limit method according to the chromatogram. The method has the characteristics of good separation degree, good durability, high sensitivity and good reproducibility.

Inventors

  • TAN ZHIWEN
  • YANG YUFENG
  • ZHOU CHUNYAN

Assignees

  • 重庆华邦制药有限公司

Dates

Publication Date
20260512
Application Date
20241111

Claims (10)

  1. 1. The method for separating enzate Lu An Z 3 and genotoxic impurities thereof based on high performance liquid chromatography is characterized in that enzate Lu An Z 3 and the genotoxic impurities thereof form a composition, the genotoxic impurities comprise any one or more of impurity Xd, impurity Xg and impurity X h , the method comprises the steps of adopting octadecylsilane bonded silica gel as a chromatographic column filling agent, adopting trifluoroacetic acid solution as a mobile phase A and acetonitrile as a mobile phase B, separating enzate Lu An Z 3 and the genotoxic impurities thereof through linear gradient elution, and each component in the composition has the following structural formula:
  2. 2. the method according to claim 1, characterized in that the procedure of the linear gradient elution is as follows: Time-minutes Mobile phase a- (parts by volume) Mobile phase B- (parts by volume) 0 60±10 40±10 10±0.5 60±10 40±10 30±0.5 20±10 80±10 35±0.5 20±10 80±10 36±0.5 60±10 40±10 45±0.5 60±10 40±10 。
  3. 3. The method according to claim 1, characterized in that the procedure of the linear gradient elution is as follows: Time-minutes Mobile phase a- (parts by volume) Mobile phase B- (parts by volume) 0 60±2 40±2 10 60 40±2 30 20 80 35 20 80 36 60±2 40±2 45 60±2 40±2 。
  4. 4. The method according to claim 1, wherein the flow rate is 0.8-1.2mL/min and the column temperature is 20-30 ℃.
  5. 5. The method of claim 1, wherein the method is used to isolate genotoxic impurities from enzane Lu An Z 3 and related substances thereof, wherein the genotoxic impurities comprise any one or more of impurity X d , impurity X g and impurity X h , and the related substances comprise impurity X, Impurity SM 3 , impurity Z 3a , impurity Z 3c , impurity Z 3g , impurity Z 3k , Impurity Z 3m , impurity Z 3b , impurity Z 3l , impurity X a , impurity Z 2 , Impurity Z 3h , impurity Z 3q , impurity Z 3r , impurity Z 3s , impurity Z 3p , Any one or more of the impurities Z 3j , wherein the structural formula of the related substances is as follows:
  6. 6. A method for identifying the enzate Lu An Z 3 and the genotoxic impurities thereof, which is characterized in that the method is adopted to separate the composition and detect the composition by a detector with the detection wavelength of 275+/-10 nm, so as to obtain a chromatogram, and the chromatographic characteristics of the detected product and a reference product are compared to determine whether the detected product contains the enzate Lu An Z 3 and the genotoxic impurities thereof.
  7. 7. The method of claim 56, wherein the components of the composition are identified in ascending order of retention time, impurity X g , impurity X h , and impurity X d .
  8. 8. The method of claim 6, wherein the retention time is 7.9.+ -. 0.5min, determined as impurity X g , the retention time is 17.7.+ -. 0.5min, determined as impurity X h , and the retention time is 22.8.+ -. 0.5min, determined as impurity X d .
  9. 9. The method for detecting whether the content of the genotoxic impurities in the enzate Lu An Z 3 is qualified is characterized by comprising the following steps of: (1) Separating and identifying enza Lu An Z 3 and genotoxic impurities thereof by the method of any one of claims 6-8 to obtain a chromatogram; (2) Judging whether the impurity content in the detected product is qualified or not according to the chromatogram obtained in the step (1); If the peak areas of the impurities X g , X h and/or X d in the detected product are not larger than the peak areas of the corresponding impurities in the reference product solution, the impurity content is qualified, otherwise, if the peak areas of the impurities X g , X h and/or X d in the detected product are larger than the peak areas of the corresponding impurities in the reference product solution, the impurity content is unqualified.
  10. 10. The method of claim 9, wherein the sample formulation solvent is tetrahydrofuran.

Description

Method for separating and measuring enza Lu An Z 3 and genotoxic impurities thereof Technical Field The invention belongs to the technical field of medicine analysis, and particularly relates to a method for separating and measuring enzate Lu An Z 3 and genotoxic impurities thereof. Background Enza Lu An, also known as enzalutamide, is an androgen receptor inhibitor, developed by An Si tay in combination with company Medivation in the united states. Enzam Lu An is used primarily for the treatment of metastatic castration-resistant prostate cancer that is asymptomatic or has mild symptoms after failure of androgen deprivation therapy and has not received chemotherapy. It inhibits proliferation and induces death of prostate cancer cells by competitively inhibiting androgen binding to androgen receptor. Enza3835Z 3 is a key intermediate in the synthesis process of enzas Lu An, and the structural formula of the enzas Lu An Z 3 is shown in formula 1. It has been found that three genotoxic impurities with a warning structure, impurity X d, impurity X g and impurity X h, may exist in the enzate Lu An Z 3. In order to ensure the quality of the crude drug and the preparation products, the three genotoxic impurities need to be controlled. However, the analytical methods of the impurity X d, the impurity X g and the impurity X h are not carried out in pharmacopoeias of various countries, and are not reported in literature. In the prior art, CN106153772B discloses a method for detecting related substances of enzalutamide by adopting high performance liquid chromatography. The patent uses pentafluorophenyl bonded silica gel as a stationary phase, adopts an acid solution-organic phase as a mobile phase, and performs gradient elution, wherein the mobile phase is trifluoroacetic acid solution and acetonitrile. However, the method cannot separate and detect the impurity X d, the impurity X g and the impurity X h in the enzate Lu An Z 3. Therefore, the establishment of the separation and measurement method for the impurities X d, X g and X h in the enzal Lu An Z 3 has important significance for realizing the quality control of the enzal Lu An intermediate Z 3 and the enzal Lu An finished products. Disclosure of Invention Accordingly, it is an object of the present invention to provide a method for separating enzate Lu An Z 3 and its genotoxic impurities based on high performance liquid chromatography, which can accomplish the separation of various substances in a short time. In order to achieve the above purpose, the technical scheme of the invention is as follows: A method for separating enzate Lu An Z 3 and genotoxic impurities thereof based on a high performance liquid chromatography, wherein the enzate Lu An Z 3 and the genotoxic impurities jointly form a composition, the genotoxic impurities comprise any one or more of an impurity X d, an impurity X g and an impurity X h, the method comprises the steps of adopting octadecylsilane bonded silica gel as a chromatographic column filling agent, adopting trifluoroacetic acid solution as a mobile phase A and acetonitrile as a mobile phase B, separating the enzate Lu An Z 3 and the genotoxic impurities thereof through linear gradient elution, and each component in the composition has the following structural formula: the separated material can be used for the next production. The above impurities may be arranged and combined in various ways. For example, combination of impurity X d and impurity X h. As another example, impurity X g and impurity X h are combined in manner 2. For another example, combination 3 is impurity X d, impurity X g, and impurity X h. Again, this is not exhaustive of all permutations and combinations. In theory, when the upper limit of the substances for separation, identification and/or content detection in the method is n (the number of substances), 1-n substances can be naturally detected. Preferably, the mobile phase A is a trifluoroacetic acid solution with a concentration of 0.05%. As a preferable scheme, the preparation method of the trifluoroacetic acid solution in the mobile phase comprises the steps of weighing 1000ml of water, adding 0.5ml of trifluoroacetic acid, and uniformly mixing. As a preferred embodiment, the procedure for the linear gradient elution is as follows: Time-minutes Mobile phase a- (parts by volume)Mobile phase B- (parts by volume)060±1040±1010±0.560±1040±1030±0.520±1080±1035±0.520±1080±1036±0.560±1040±1045±0.560±1040±10 。 As a preferred method, the procedure for the linear gradient elution is as follows: Time-minutes Mobile phase a- (parts by volume)Mobile phase B- (parts by volume)060±240±2106040±23020803520803660±240±24560±240±2 。 For example, 58 parts of the mobile phase A, 42 parts of the mobile phase B (FIG. 6), 59 parts of the mobile phase A, 41 parts of the mobile phase B, 60 parts of the mobile phase A, 40 parts of the mobile phase B (FIG. 3), 61 parts of the mobile phase A, 39 parts of the mobile phase B, 62 parts of