Separation and detection method of cis-isomer in olotinib

Abstract

The invention provides a separation and detection method of a cis-isomer in olotinib. According to the method, chiral chromatographic column high performance liquid chromatography is adopted, cellulose derivative bonded silica gel is taken as a stationary phase, and n-hexane and ethanol are taken as a mobile phase. The method provided by the invention can effectively separate and measure the cis-isomer in the olotinib, and has the advantages of high efficiency, simplicity, convenience, high sensitivity, good separation effect and separation degree of 5.42. The invention provides a detection method for developing and marketing the olatinib and accurately detecting the cis-isomer in the olatinib, and ensures the product quality.

Description

Technical Field

[0001] The present invention belongs to the field of drug analysis, and in particular relates to a method for separating and detecting cis isomers in oclacitinib. Background Art

[0002] Oclacitinib was approved by the US FDA in 2013 for the control of pruritus and atopic dermatitis caused by canine allergic dermatitis. Oclacitinib is a very effective JAK1 inhibitor, and also inhibits the function of JAK1-dependent cytokines in some anti-allergic, inflammatory and pruritus responses. It has little effect on cytokines that are not involved in activating JAK1. In early 2019, Oclacitinib maleate tablets produced by Zoetis in the United States were reviewed and approved for registration in my country, and the "Imported Veterinary Drug Registration Certificate" was issued. Since Zoetis launched Apoquel in 2013, it has benefited tens of millions of dogs suffering from itchy diseases. Apoquel ( Generic name: Oclacitinib Maleate Tablets) has become Zoetis' largest single product worldwide. The currently available Oclacitinib has an effective configuration of trans, and the cis isomer is reported to be inactive and has certain toxic side effects.

[0003]

[0004] At present, there are no reports on the determination method of cis-isomer in oclacitinib through searching relevant literature, enterprise standards and pharmacopoeia. It is very necessary to establish a separation and detection method for cis-isomer in oclacitinib, which provides a detection method for the development and marketing of oclacitinib generic drugs and the accurate detection of cis-isomer in oclacitinib, thus ensuring product quality. Summary of the invention

[0005] The object of the present invention is to provide a method for effectively separating and determining the cis isomer in oclacitinib, which is efficient, simple, highly sensitive and has a good separation effect. The specific technical scheme is as follows:

[0006] A method for separating and detecting cis isomers in oclacitinib, characterized in that the method is carried out by high performance liquid chromatography using a chiral chromatographic column, using a cellulose derivative bonded silica gel as the stationary phase, and using n-hexane and ethanol as the mobile phase.

[0007] A method for separating and detecting cis isomers in oclacitinib, wherein the volume ratio of n-hexane to ethanol is (75-85):(15-25). Preferably, the volume ratio of n-hexane to ethanol is 80:20.

[0008] The flow rate of the mobile phase is 0.6-1.2 ml / min. Preferably, the flow rate of the mobile phase is 0.8 ml / min.

[0009] The detection wavelength of the liquid chromatography is 200nm-380nm. Preferably, the detection wavelength of the liquid chromatography is 220nm.

[0010] Chiral chromatographic column model: ChiralGel BH / BH-R, Chiral MZ(2), Ultimate Cellu-J, Chiral MD(2), Ultimate Cellu-DR, Hypersil TM Chiral HPLC columns, AstecCellulose DMP, ChiralGel CH / CH-R, Any one of .

[0011] Preferably, the chiral chromatographic column model is ChiralGel BH / BH-R, Chiral MD(2), Ultimate Cellu-DR, Hypersil TM Chiral HPLC columns, AstecCellulose DMP, Any one of .

[0012] A method for separating and detecting cis isomers in oclacitinib, characterized in that:

[0013] (1) Take appropriate amounts of oclacitinib and cis-isomer, dissolve in ethanol and dilute to make a sample solution containing approximately 1 mg per 1 ml;

[0014] (2) Using cellulose derivative bonded silica gel as filler; using a mixture of n-hexane and ethanol in a volume ratio of 80:20 as mobile phase; setting the flow rate to 0.8 ml per minute; the detection wavelength to 220 nm; and the column temperature to room temperature;

[0015] (3) Take 5-20 μl of the sample solution of step (1) and inject it into a high performance liquid chromatograph (4.6 mm×250 mm, 5 μm) to complete the separation and detection of the cis isomers in oclacitinib.

[0016] The beneficial effects of the present invention are:

[0017] 1) The method provided by the present invention can effectively separate and determine the cis isomers in oclacitinib. The method is efficient, simple, highly sensitive, and has a good separation effect, with a separation degree of 5.42. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 HPLC detection spectrum of Oclacitinib + cis isomer

[0020] Figure 2 HPLC detection spectrum of Oclacitinib

[0021] Figure 3 HPLC detection spectrum of oclacitinib cis-isomer DETAILED DESCRIPTION

[0022] The present invention is described in detail below through specific examples. The following examples are used to explain the present invention rather than to limit the present invention.

[0023] Example 1

[0024] 1. Instruments and Reagents

[0025]

[0026] 2. Methods and Results

[0027] (1) Take appropriate amounts of oclacitinib and cis-isomer, dissolve in ethanol and dilute to make a sample solution containing approximately 1 mg per 1 ml;

[0028] (2) Cellulose derivative bonded silica gel was used as filler; a mixture of n-hexane and ethanol in a volume ratio of 80:20 was used as mobile phase; the flow rate was set to 0.8 ml per minute; the detection wavelength was 220 nm; and the column temperature was 25°C;

[0029] (3) Take 10 μl of the sample solution in step (1) and inject it into a high performance liquid chromatograph (4.6 mm×250 mm, 5 μm) to complete the separation and detection of the cis isomer in oclacitinib.

[0030] Test results (such as Figure 1 ):

[0031] The peak time of oclacitinib is 21.998min, and the peak time of the cis isomer is 14.505min.

[0032] Systematic review:

[0033] Peak Peak name Retention time Half peak width Theoretical plates Separation Tailing Factor Asymmetry 1 Cis isomer 14.505 0.740 4925.544 3.526 1.201 1.340 2 Oclacitinib 21.998 0.740 4925.544 3.526 1.201 1.340

[0034] Example 2

[0035] Detection of cis-isomer of oclacitinib:

[0036] Test results (such as Figure 3 ):

[0037] The purity of the cis-isomer of oclacitinib is 99.86%, and the peak time is 14.490 min.

[0038] Example 3

[0039] Oclacitinib optical purity test:

[0040] Test results (such as Figure 2 ):

[0041] The purity of oclacitinib is 99.86%, and the cis-isomer of oclacitinib is 0.14%. The peak time of oclacitinib is 21.908min, and the peak time of oclacitinib cis-isomer is 14.502min.

[0042] Systematic review:

[0043] Peak Peak name Retention time Half peak width Theoretical plates Separation Tailing Factor Asymmetry 1 Cis isomer 14.502 0.467 5773.438 5.418 0.953 1.129 2 Oclacitinib 21.908 0.810 4080.007 5.418 1.251 1.436

[0044] It can be seen that the separation and detection method of the present invention can separate the cis isomer in oclacitinib, and this method provides a detection method for the subsequent development and marketing of oclacitinib and the accurate detection of the cis isomer in oclacitinib.

[0045] The present invention is not limited to the above embodiments. Any simple or equivalent changes or modifications made to the above embodiments based on the technical essence of the present invention are within the technical scope of the present invention.

Claims

1. A method for separating and detecting cis isomers in oclacitinib, characterized in that The method adopts chiral chromatographic column high performance liquid chromatography, takes cellulose derivative bonded silica gel as stationary phase, and takes normal hexane and ethanol as mobile phase.

2. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: Furthermore, the volume ratio of n-hexane to ethanol is (75-85):(15-25).

3. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The volume ratio of n-hexane to ethanol is 80:

20.

4. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The flow rate of the mobile phase is 0.6-1.2 ml / min.

5. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The flow rate of the mobile phase is 0.8 ml / min.

6. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The detection wavelength of the liquid chromatography is 200nm-380nm.

7. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The detection wavelength of the liquid chromatography is 220 nm.

8. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The chiral chromatographic column model is IC, IK IM, ChiralGel BH / BH-R, Chiral MZ(2), Ultimate Cellu-J, Chiral MD(2), OD-H, Ultimate Cellu-DR, Hypersil TM Chiral HPLC columns, Astec Cellulose DMP, ChiralGelCH / CH-R, Any one of IB N-5.

9. The method for separating and detecting cis isomers in oclacitinib according to claim 1, characterized in that: The chiral chromatographic columns are ChiralGel BH / BH-R, Chiral MD(2), OD-H, Ultimate Cellu-DR, Hypersil TM Chiral HPLC columns, Astec Cellulose DMP, Any one of IB N-5.

10. A method for separating and detecting cis isomers in oclacitinib according to any one of claims 1 to 9, characterized in that: (1) Take appropriate amounts of oclacitinib and cis-isomer, dissolve in ethanol and dilute to make a sample solution containing approximately 1 mg per 1 ml; (2) using cellulose derivative bonded silica gel as filler; using a mixture of n-hexane and ethanol in a volume ratio of 80:20 as mobile phase; setting the flow rate to 0.8 ml per minute; the detection wavelength to 220 nm; and the column temperature to room temperature; (3) Take 5-20 μl of the sample solution of step (1) and inject it into a high performance liquid chromatograph (4.6 mm×250 mm, 5 μm) to complete the separation and detection of the cis isomers in oclacitinib.

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