阅读说明：本技术 一种盐酸贝尼地平原料药中的杂质检测方法 (Method for detecting impurities in benidipine hydrochloride bulk drug ) 是由 王熙红 林丽丽 杨淑萍 宋晓娜 于 2019-10-14 设计创作，主要内容包括：本发明涉及一种盐酸贝尼地平原料药中的杂质检测方法,提供了一种高检出能力的盐酸贝尼地平杂质检测方法,并提供了一种盐酸贝尼地平原料药中β异构体的检测方法,解决了现有检测方法中盐酸贝尼地平原料药中β异构体杂质较难分离检测的技术问题。本发明提供一种盐酸贝尼地平原料药中的杂质检测方法,该方法为采用液相色谱法进行检测,且检测限低、灵敏度高、准确度高。本发明广泛应用于盐酸贝尼地平原料药中的杂质检测方法。(The invention relates to a method for detecting impurities in a benidipine hydrochloride raw material drug, provides a method for detecting benidipine hydrochloride impurities with high detection capability, provides a method for detecting beta isomers in the benidipine hydrochloride raw material drug, and solves the technical problem that the beta isomer impurities in the benidipine hydrochloride raw material drug are difficult to separate and detect in the existing detection method. The invention provides a method for detecting impurities in a benidipine hydrochloride raw material medicine, which adopts liquid chromatography for detection and has the advantages of low detection limit, high sensitivity and high accuracy. The method is widely applied to the impurity detection method in the benidipine hydrochloride bulk drug.)

1. A method for detecting impurities in a benidipine hydrochloride raw material medicine is characterized by adopting a liquid chromatography method for detection, and comprises the following steps:

(1) preparing a test solution: weighing 25mg of the product, accurately weighing, placing in a 50ml measuring flask, adding methanol-water (1: 1) for dissolving, diluting to scale, and shaking to obtain sample solution;

(2) preparing a reference substance solution: precisely weighing a proper amount of benidipine hydrochloride reference substance, adding methanol-water (1: 1) for dissolving, and quantitatively diluting to prepare a solution containing about 1.0 mu g of benidipine hydrochloride in each 1ml as a reference substance solution;

(3) preparation of system applicability solution: taking a proper amount of raw materials of an impurity A reference substance, an impurity B reference substance, an impurity C reference substance and benidipine hydrochloride, precisely weighing, adding methanol-water (1: 1) to dissolve and dilute to prepare a solution containing 1.0 mu g of the impurity A, the impurity B, the impurity C and 0.5mg of the benidipine hydrochloride in each 1ml of the solution, and taking the solution as a system applicability solution;

(4) detecting the liquid chromatography condition:

measuring by high performance liquid chromatography, and separating with chromatographic column using octadecylsilane chemically bonded silica as filler; using 0.05mol/L potassium dihydrogen phosphate solution (pH is adjusted to 3.0 by phosphoric acid) -methanol-tetrahydrofuran (65: 27: 8) as a mobile phase; the column temperature was 25 ℃; the detection wavelength is 237 nm; injecting 10 mul of system applicability solution into a liquid chromatograph, adjusting the flow rate to ensure that the retention time of the benidipine hydrochloride peak is about 20 minutes, the separation degree between the benidipine hydrochloride peak and the impurity C peak is more than 1.5, and the peak emergence sequence is as follows in sequence: impurity A, impurity B, benidipine and impurity C; precisely measuring 10 μ l of each of the blank solvent, the sample solution and the reference solution, respectively injecting into a liquid chromatograph, and recording chromatogram;

continuously feeding 5 probes of the reference substance solution, wherein the relative standard deviation of the peak area of benidipine is less than 3.5%; the retention time of the chromatogram of the test solution to the main component peak is 2 times, if an impurity peak exists in the chromatogram of the test solution except the solvent peak, the peak area of the impurity A, the peak area of the impurity B (which are multiplied by 1.6 for correction), the peak area of the impurity C and the peak areas of other single impurities are not more than 0.5 times (0.1%) of the main peak area of the reference solution, and the sum of the peak areas of the impurities is not more than 0.2% of the main peak area of the reference solution.

2. The method for detecting impurities in a benidipine hydrochloride raw material drug according to claim 1, wherein the chromatographic column is an Agilent Pursuit column, 4.6mm x 100mm, 3 μm or equivalent performance chromatographic column.

Technical Field

The invention relates to the field of detection methods, and particularly relates to a method for detecting impurities in a benidipine hydrochloride raw material medicine.

Background

Benidipinehydrochloride hydrochloride is hydrochloride of (+/-) - (R) -1, 4-dihydro-2, 6-dimethyl-4- (m-nitrophenyl) -3, 5-methyl picolinate [ (R) -1-benzyl-3-piperidinol ester ], belongs to dihydropyridine calcium ion antagonists, can be efficiently and specifically combined with a dihydropyridine receptor combining part, and has efficient and specific inhibition effect on 2+2+ Ca channels. Benidipine has an inhibitory effect not only on muscle type (L type) Ca channels, but also on voltage-dependent type N and T type Ca channels, and is the only calcium ion antagonist currently having an effect on all three channels. In addition, benidipine has high affinity effect on cell membranes, blood vessel selectivity effect and kidney protection effect, and is an ideal, safe and effective medicament for resisting hypertension and treating diseases such as renal parenchymal hypertension, angina pectoris and the like.

The benidipine hydrochloride has two chiral atoms in the molecule, which are respectively positioned at the 4-position of a dihydropyridine ring and the 3' -position of a side-chain piperidine ring, so that the benidipine hydrochloride has four optical isomers of (S) - (S) - (+) -alpha, (R) - (R) - (-) -alpha, (R) - (S) - (+) -beta and (S) - (R) - (-) -beta, and the medicinal active ingredients are a mixture of (S) - (S) - (+) -alpha and (R) - (R) - (-) -alpha. Therefore, the separation of alpha isomer from beta isomer must be carried out in the post-treatment stage of synthesizing and preparing benidipine hydrochloride.

According to the synthesis sequence of the dihydropyridine main ring, the synthesis of benidipine hydrochloride is mainly divided into two types of five routes. Wherein the primary ring is synthesized by: 1) after the dihydropyridine main ring is subjected to acyl chlorination, the dihydropyridine main ring is connected with a side chain to directly synthesize benidipine hydrochloride; 2) after the acyl chlorination of the main ring of dihydropyridine, the dihydropyridine is firstly connected with 3-piperidinol and then connected with benzyl. The postsynthetic main ring is: 1) synthesis of the main ring by beta-aminocrotonate; 2) synthesis of the main ring by acetoacetate; 3) a one-pot method using 3-nitrobenzaldehyde with beta-aminocrotonate and acetoacetate, and the like.

EP0063365a1, EP0161877a2, JP57-171968A, EP0106275a2, etc. disclose one or more routes for synthesizing benidipine hydrochloride and analogs thereof, wherein EP0106275a2 summarizes the synthetic routes for benidipine hydrochloride, and in the above documents, it is suggested that the synthetically prepared benidipine hydrochloride is subjected to resolution of α isomer from β isomer by means of column chromatography separation, thereby obtaining pharmaceutically acceptable (±) - α -benidipine hydrochloride.

Therefore, the detection of the content of the beta isomer in the benidipine hydrochloride raw material medicine has great influence on the drug effect of the benidipine hydrochloride tablet.

At present, no specific method for detecting the beta isomer of benidipine hydrochloride exists in Chinese and Japanese pharmacopoeias, so that a method needs to be developed for quantitative detection, the developed method can ensure that the dehydrogenized derivative (impurity B) and the process impurity (impurity A) of the benidipine hydrochloride can be effectively detected, and other impurities can be detected to the maximum extent.

Disclosure of Invention

The invention aims to overcome the defects of the prior art, and provides the method for detecting the impurities in the benidipine hydrochloride raw material drug, which can effectively detect the beta isomer impurities in the benidipine hydrochloride raw material drug, and has the advantages of low detection limit, high sensitivity and high accuracy according to the structural characteristics of the benidipine hydrochloride chiral molecules.

The technical scheme adopted by the invention for solving the technical problem is as follows:

a method for detecting impurities in a benidipine hydrochloride raw material medicine adopts liquid chromatography for detection, and comprises the following steps:

(1) preparing a test solution: weighing 25mg of the product, accurately weighing, placing in a 50ml measuring flask, adding methanol-water (1: 1) for dissolving, diluting to scale, and shaking to obtain sample solution;

(2) preparing a reference substance solution: precisely weighing a proper amount of benidipine hydrochloride reference substance, adding methanol-water (1: 1) for dissolving, and quantitatively diluting to prepare a solution containing about 1.0 mu g of benidipine hydrochloride in each 1ml as a reference substance solution;

(3) preparation of system applicability solution: taking a proper amount of raw materials of an impurity A reference substance, an impurity B reference substance, an impurity C reference substance and benidipine hydrochloride, precisely weighing, adding methanol-water (1: 1) to dissolve and dilute to prepare a solution containing 1.0 mu g of the impurity A, the impurity B, the impurity C and 0.5mg of the benidipine hydrochloride in each 1ml of the solution, and taking the solution as a system applicability solution;

(4) detecting the liquid chromatography condition:

measuring by high performance liquid chromatography, and separating with chromatographic column using octadecylsilane chemically bonded silica as filler; using 0.05mol/L potassium dihydrogen phosphate solution (pH is adjusted to 3.0 by phosphoric acid) -methanol-tetrahydrofuran (65: 27: 8) as a mobile phase; the column temperature was 25 ℃; the detection wavelength is 237 nm; injecting 10 mul of system applicability solution into a liquid chromatograph, adjusting the flow rate to ensure that the retention time of the benidipine hydrochloride peak is about 20 minutes, the separation degree between the benidipine hydrochloride peak and the impurity C peak is more than 1.5, and the peak emergence sequence is as follows in sequence: impurity A, impurity B, benidipine and impurity C; precisely measuring 10 μ l of each of the blank solvent, the sample solution and the reference solution, respectively injecting into a liquid chromatograph, and recording chromatogram;

continuously feeding 5 probes of the reference substance solution, wherein the relative standard deviation of the peak area of benidipine is less than 3.5%; the retention time of the chromatogram of the test solution to the main component peak is 2 times, if an impurity peak exists in the chromatogram of the test solution except the solvent peak, the peak area of the impurity A, the peak area of the impurity B (which are multiplied by 1.6 for correction), the peak area of the impurity C and the peak areas of other single impurities are not more than 0.5 times (0.1%) of the main peak area of the reference solution, and the sum of the peak areas of the impurities is not more than 0.2% of the main peak area of the reference solution.

Preferably, the column is an Agilent Pursuit column, 4.6mm by 100mm, 3 μm or equivalent performance column.

The invention has the beneficial effects that:

the method for detecting impurities in the benidipine hydrochloride raw material medicine can effectively detect the beta isomer impurities in the benidipine hydrochloride raw material medicine, and has the advantages of low detection limit, high sensitivity and high accuracy.

Detailed Description

The present invention will be further described with reference to specific examples to assist understanding of the invention. The method used in the invention is a conventional production method if no special provisions are made; the starting materials used, unless otherwise specified, are conventional commercial products.

Examples

A method for detecting impurities in a benidipine hydrochloride raw material medicine adopts liquid chromatography for detection, and comprises the following steps:

(1) preparing a test solution: weighing 25mg of the product, accurately weighing, placing in a 50ml measuring flask, adding methanol-water (1: 1) for dissolving, diluting to scale, and shaking to obtain sample solution;

(2) preparing a reference substance solution: precisely weighing a proper amount of benidipine hydrochloride reference substance, adding methanol-water (1: 1) for dissolving, and quantitatively diluting to prepare a solution containing about 1.0 mu g of benidipine hydrochloride in each 1ml as a reference substance solution;

(3) preparation of system applicability solution: taking a proper amount of raw materials of an impurity A reference substance, an impurity B reference substance, an impurity C reference substance and benidipine hydrochloride, precisely weighing, adding methanol-water (1: 1) to dissolve and dilute to prepare a solution containing 1.0 mu g of the impurity A, the impurity B, the impurity C and 0.5mg of the benidipine hydrochloride in each 1ml of the solution, and taking the solution as a system applicability solution;

(4) detecting the gas chromatography conditions:

measuring by high performance liquid chromatography, using octadecyl silane bonded silica gel as filler, Agilent Pursuit column, 4.6mm × 100mm, 3 μm or equivalent chromatographic column; using 0.05mol/L potassium dihydrogen phosphate solution (pH is adjusted to 3.0 by phosphoric acid) -methanol-tetrahydrofuran (65: 27: 8) as a mobile phase; the column temperature was 25 ℃; the detection wavelength is 237 nm; injecting 10 mul of system applicability solution into a liquid chromatograph, adjusting the flow rate to ensure that the retention time of the benidipine hydrochloride peak is about 20 minutes, the separation degree between the benidipine hydrochloride peak and the impurity C peak is more than 1.5, and the peak emergence sequence is as follows in sequence: impurity A, impurity B, benidipine and impurity C; precisely measuring 10 μ l of each of the blank solvent, the sample solution and the reference solution, respectively injecting into a liquid chromatograph, and recording chromatogram;

continuously feeding 5 probes of the reference substance solution, wherein the relative standard deviation of the peak area of benidipine is less than 3.5%; the retention time of the chromatogram of the test solution to the main component peak is 2 times, if an impurity peak exists in the chromatogram of the test solution except the solvent peak, the peak area of the impurity A, the peak area of the impurity B (which are multiplied by 1.6 for correction), the peak area of the impurity C and the peak areas of other single impurities are not more than 0.5 times (0.1%) of the main peak area of the reference solution, and the sum of the peak areas of the impurities is not more than 0.2% of the main peak area of the reference solution.

In conclusion, the method for detecting the impurities in the benidipine hydrochloride raw material medicine can effectively detect the beta isomer impurities in the benidipine hydrochloride raw material medicine, and has the advantages of low detection limit, high sensitivity and high accuracy.

However, the above description is only an embodiment of the present invention, and the scope of the present invention should not be limited by this, and all equivalent changes and modifications made in the claims of the present invention should be covered by the present invention.