阅读说明：本技术 与洛铂有关物质的检测 (Detection of lobaplatin-related substances ) 是由 窦啟玲 汪立冬 常新亮 于 2019-03-19 设计创作，主要内容包括：本发明涉及与洛铂有关物质的检测。本发明提供了与洛铂有关物质的检测方法,所述与洛铂有关物质为化合物J,其结构式为<Image he="194" wi="678" file="DDA0001999851310000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>所述检测方法为HPLC-MS法或者HPLC法,所述HPLC-MS法的检测条件为：用十八烷基硅烷键合硅胶为填充剂,以18-22mmol/L的甲酸铵为流动相A,甲醇：乙腈的体积比例＝1:(0.8-1.2)为流动相B,进行梯度洗脱。该检测方法的灵敏度高,专属性强,重复性好,准确度高。(The invention relates to detection of substances related to lobaplatin. The invention provides a method for detecting a lobaplatin related substance, wherein the lobaplatin related substance is a compound J, and the structural formula of the lobaplatin related substance is shown in the specification The detection method is an HPLC-MS method or an HPLC method, and the detection conditions of the HPLC-MS method are as follows: octadecylsilane chemically bonded silica is used as a filling agent, 18-22mmol/L ammonium formate is used as a mobile phase A, and methanol: acetonitrile with volume ratio of 1 (0.8-1.2) as mobile phase B, and gradient elution is carried out. The detection method has the advantages of high sensitivity, strong specificity, good repeatability and high accuracy.)

1. A method for detecting a lobaplatin-related substance, wherein the lobaplatin-related substance is a compound J, and the structure of the compound J is shown in the specification

2. The detection method according to claim 1, wherein the compound J is prepared from a compound (2) or a compound (3) or a compound (4) of the following structural formula

3. The detection method according to claim 1 or 2, wherein the compound J is a compound (1) as a starting material

4. The detection method according to any one of claims 1 to 3, wherein the compound J is produced by a method comprising:

wherein, in the reaction for preparing the compound 2 from the compound 1, chloroplatinic acid salt or chloroplatinic acid salt, alkali metal iodide and hydroxide are reacted to prepare a compound 2; preferably, the chloroplatinic acid salt is selected from potassium or sodium chloroplatinic acid, preferably potassium chloroplatinic acid; preferably, the alkali metal iodide is selected from potassium iodide or sodium iodide, preferably potassium iodide; preferably, the hydroxide is selected from potassium hydroxide or sodium hydroxide, preferably potassium hydroxide;

and/or in the reaction of preparing the compound 3 by the compound 2, adding the compound 2 into water and a ketone solvent to obtain a material A, then adding a silver nitrate solution into the material A for reaction, and filtering to obtain a solution of the compound 3;

and/or in the reaction for preparing the compound 4 by the compound 3, mixing and stirring the solution of the compound 3 and resin, and then filtering to obtain a compound 4 solution;

and/or, in the reaction for preparing the product by the compound 4, the obtained solution of the compound 4 is adjusted to be alkaline by an acidic regulator, and then the final product is obtained by freeze-drying.

5. The detection method according to any one of claims 1 to 4, wherein the detection method is an HPLC-MS method or an HPLC method.

6. The detection method according to claim 5, wherein the HPLC-MS method comprises the following HPLC detection conditions: octadecylsilane chemically bonded silica is used as a filling agent, 18-22mmol/L ammonium formate is used as a mobile phase A, and methanol: the volume ratio of acetonitrile is 1, (0.8-1.2) is used as a mobile phase B, and gradient elution is carried out; preferably, the mobile phase A is 20mmol/L ammonium formate solution, and the mobile phase B is methanol: the volume ratio of acetonitrile is 1:1.

7. The detection method according to claim 6, wherein the gradient elution pattern in the HPLC-MS method is as follows:

0-3 minutes: 97 vol% mobile phase a: 3 volume% mobile phase B;

3-10 minutes: mobile phase a decreased from 97 vol% to 92 vol%, mobile phase B increased from 3 vol% to 8 vol%;

10-18 minutes: mobile phase a decreased from 92% to 87% by volume and mobile phase B increased from 8% to 13% by volume;

18-25 minutes: mobile phase a decreased from 87 vol% to 10 vol%, mobile phase B increased from 13 vol% to 90 vol%;

25-26 minutes: mobile phase a increased from 10 vol% to 97 vol%, and mobile phase B decreased from 90 vol% to 3 vol%;

26-34 minutes: 97 vol% mobile phase a: 3 volume% mobile phase B;

wherein, each time range of the gradient elution can be increased by 1-2 minutes or the time range of the gradient elution from 3-10 minutes can be decreased by 1-2 minutes.

8. The method of claim 7, wherein the MS conditions in HPLC-MS are electrospray ion source and m/z of the compound detected by positive ion detection is 326.

9. The test method according to any one of claims 1 to 8, wherein the flow rate is 0.8-1.2ml, preferably 1.0ml per minute; the column temperature is 39-41 deg.C, preferably 40 deg.C.

10. The detection method according to any one of claims 1 to 9, wherein a separation degree of a peak of the compound J from its neighboring related substance in a chromatogram of a system suitability test solution is not less than 1.5.

11. The detection method according to any one of claims 1 to 10, wherein the peak area of the chromatogram of the test solution is not larger than the peak area of compound J of the control solution if compound J is present.

12. The detection method according to any one of claims 1 to 11, wherein said lobaplatin comprises either one or both of lobaplatin diastereomer i and lobaplatin diastereomer ii.

Technical Field

The invention relates to the field of medicines, in particular to a method for detecting substances related to lobaplatin, belonging to the technical field of medicine analysis quality control.

Background

Lobaplatin (Lobaplatin, D19466), also known as Lobaplatin, is a third-generation platinum-based antitumor drug following cisplatin and carboplatin, and its chemical name is: cis- [ trans-1, 2-cyclobutanAlkane bis (methylamine) -N, N']- [ (2S) -lactic acid-O1, O2]-platinum (II), formula C₉H₁₈N₂O₃Pt has a molecular weight of 397.34 and a chemical structural formula shown in the following formula (a):

lobaplatin has alkylating effect, belongs to alkylating agent (broad sense), and has good antitumor effect, such as inhibiting in vitro AH 135-tumor, B16-melanoma, colon cancer 115, and in vivo mouse P338 leukemia. Lobaplatin is characterized by strong anticancer activity, low toxicity, no accumulative toxicity and renal toxicity and less toxicity to bone marrow, and currently marketed lobaplatin for injection is mainly used for treating breast cancer, small cell lung cancer and chronic myelogenous leukemia.

Disclosure of Invention

In order to ensure the safety, effectiveness and controllable quality of the medicine, the research on related substances and detection methods of the related substances is very important. For the drug, due to the existence of three chiral carbons and related substances generated in the preparation process, confirming the structure of the related substances and finding a suitable detection method for controlling the product quality of the drug become technical problems to be solved urgently in the field.

The technical problem to be solved by the invention is to provide a new detection method to establish the detection of related substances in the lobaplatin so as to carry out quality control on the lobaplatin compound.

One skilled in the art will recognize that any substance that affects the purity of a drug is collectively referred to as a related substance. Research on related substances is an important part of drug development, and includes selecting appropriate analytical methods, accurately distinguishing the content of the related substances to be measured, and determining the reasonable limits of the related substances by combining the results of pharmaceutical, toxicological and clinical studies. This study is throughout the entire process of drug development.

Specifically, the present invention is realized by the following technical means.

The invention provides a method for detecting a lobaplatin related substance, wherein the lobaplatin related substance is a compound J, and the structure of the compound J is

Preferably, in the detection method, the compound J is prepared by a compound (2) or a compound (3) or a compound (4) of the following structural formula

Preferably, in the detection method, the compound J is a compound (1) as a starting material

Preferably, in the detection method, the compound J is prepared by the following method:

wherein, in the reaction for preparing the compound 2 from the compound 1, chloroplatinic acid salt or chloroplatinic acid salt, alkali metal iodide and hydroxide are reacted to prepare a compound 2; preferably, the chloroplatinic acid salt is selected from potassium or sodium chloroplatinic acid, preferably potassium chloroplatinic acid; preferably, the alkali metal iodide is selected from potassium iodide or sodium iodide, preferably potassium iodide; preferably, the hydroxide is selected from potassium hydroxide or sodium hydroxide, preferably potassium hydroxide;

and/or in the reaction of preparing the compound 3 by the compound 2, adding the compound 2 into water and a ketone solvent to obtain a material A, then adding a silver nitrate solution into the material A for reaction, and filtering to obtain a solution of the compound 3;

and/or in the reaction for preparing the compound 4 by the compound 3, mixing and stirring the solution of the compound 3 and resin, and then filtering to obtain a compound 4 solution;

and/or, in the reaction for preparing the product by the compound 4, the obtained solution of the compound 4 is adjusted to be alkaline by an acidic regulator, and then the final product is obtained by freeze-drying.

Preferably, in the detection method described above, in the reaction step of preparing the compound 2 from the compound 1, the compound 1 and an aqueous solution of potassium hydroxide are mixed to obtain a solution F, an aqueous solution of potassium chloroplatinite and potassium iodide are mixed to obtain a solution E, and the solution F and the solution E are mixed and reacted to obtain the compound 2; preferably, the reaction temperature is 25-35 ℃, preferably 30 ℃; preferably, the reaction time is 1 to 3 hours, preferably 2 hours; preferably, the molar ratio of compound 1 to chloroplatinic acid salt is 1:0.5-2, preferably 1: 0.85;

preferably, in the reaction step of preparing the compound 3 from the compound 2, the molar ratio of the compound 2 to the silver nitrate is 1 (1-2), preferably 1: 1.7; preferably, the reaction temperature is 25-35 ℃; more preferably, the reaction temperature is 30 ℃; preferably, the reaction time is 15 to 20 hours in a dark place; preferably, the reaction time is 18 hours in the absence of light; preferably, the ketone solvent is selected from one of acetone, methyl butanone, methyl ethyl ketone or methyl isobutyl ketone, and acetone is more preferred;

preferably, in the reaction for preparing the compound 4 by the compound 3, the temperature for mixing and stirring the compound 3 solution and the resin is 25-35 ℃; preferably, the stirring time is 0.5-2 hours, preferably the stirring time is 1 hour, and more preferably the stirring temperature is 30 ℃; preferably, the resin is a resin that has been treated; preferably, the resin which has been treated is a resin which has been treated with an aqueous solution of sodium hydroxide, preferably having a concentration of 1 to 2mol/L, more preferably 1.5 mol/L;

preferably, in the reaction for preparing the product by the compound 4, the acidity regulator is selected from one of p-toluenesulfonic acid or methanesulfonic acid, and is preferably p-toluenesulfonic acid; preferably, the concentration of the aqueous solution of p-toluenesulfonic acid is 5 to 15 wt%, preferably 10 wt%; the pH value of the alkalescence is 7-8; preferably, after adjusting to be slightly alkaline, reacting for 15-25 hours at the temperature of 20-30 ℃; preferably, the reaction is carried out at 25 ℃ for 20 hours.

Preferably, the detection method is an HPLC-MS method or an HPLC method.

Preferably, in the detection method, the HPLC detection conditions of the HPLC-MS method are: octadecylsilane chemically bonded silica is used as a filling agent, 18-22mmol/L ammonium formate is used as a mobile phase A, and methanol: the volume ratio of acetonitrile is 1, (0.8-1.2) is used as a mobile phase B, and gradient elution is carried out; preferably, the mobile phase A is 20mmol/L ammonium formate solution, and the mobile phase B is methanol: the volume ratio of acetonitrile is 1:1.

Preferably, in the detection method, the gradient elution pattern in the HPLC-MS method is as follows:

0-3 minutes: 97 vol% mobile phase a: 3 volume% mobile phase B;

3-10 minutes: mobile phase a decreased from 97 vol% to 92 vol%, mobile phase B increased from 3 vol% to 8 vol%;

10-18 minutes: mobile phase a decreased from 92% to 87% by volume and mobile phase B increased from 8% to 13% by volume;

18-25 minutes: mobile phase a decreased from 87 vol% to 10 vol%, mobile phase B increased from 13 vol% to 90 vol%;

25-26 minutes: mobile phase a increased from 10 vol% to 97 vol%, and mobile phase B decreased from 90 vol% to 3 vol%;

26-34 minutes: 97 vol% mobile phase a: 3 volume% mobile phase B;

wherein, each time range of the gradient elution can be increased by 1-2 minutes or the time range of the gradient elution from 3-10 minutes can be decreased by 1-2 minutes.

For example, the time range corresponding to gradient elution may be 0 to 4 minutes (or 0 to 5 minutes), 4 to 11 minutes (or 5 to 12 minutes), 11 to 19 minutes (or 12 to 20 minutes), 19 to 26 minutes (or 20 to 27 minutes), 26 to 27 minutes (or 27 to 28 minutes), 27 to 35 minutes (or 28 to 36 minutes); the time may be 0 to 3 minutes, 3 to 9 minutes (or 3 to 8 minutes), 9 to 17 minutes (or 8 to 16 minutes), 17 to 24 minutes (or 16 to 23 minutes), 24 to 25 minutes (or 23 to 24 minutes), or 25 to 33 minutes (or 24 to 32 minutes).

Preferably, in the detection method, the MS condition in the HPLC-MS is that an electrospray ion source is adopted, and m/z of the compound detected by positive ion detection is 326.

Preferably, for the detection method described above, wherein the flow rate is 0.8-1.2ml per minute, preferably 1.0 ml; the column temperature is 39-41 deg.C, preferably 40 deg.C.

Preferably, in the detection method described above, in the chromatogram of the system suitability test solution, the separation degree of the peak of compound J from the related substance adjacent thereto in the lobaplatin is not less than 1.5.

In the detection method, if compound J exists in the chromatogram of the sample solution, the peak area of compound J in lobaplatin should not be larger than that of compound J in the control solution.

With respect to the detection method described above, wherein said lobaplatin comprises either one or both of lobaplatin diastereomer i and lobaplatin diastereomer ii.

The invention provides a pharmaceutical composition containing related substances, which is a pharmaceutical preparation; preferably, the pharmaceutical composition is a pharmaceutical preparation for injection.

Preferably, the pharmaceutical composition comprises an adjuvant, and preferably, the adjuvant is one or more selected from the group consisting of a filler, a disintegrant, a lubricant, a suspending agent, a binder, a sweetener, a flavoring agent, a preservative, an antioxidant, and a matrix; preferably, the auxiliary material is a filler and/or an antioxidant.

The invention provides an application of the related substance or the pharmaceutical composition in preparing an anti-tumor medicament.

Preferably, for the above-mentioned use, wherein the tumor is a lung cancer, ovarian cancer, leukemia and/or renal cancer cell; further preferably, the tumor is a leukemia cell.

Preferably, for the above-mentioned application, the related substance or the above-mentioned pharmaceutical composition is used for preparing anti-THP-1 tumor drugs.

To enable formulation in the compositions of the present invention, pharmaceutically acceptable excipients may be added in the preparation of these formulations, such as: fillers, disintegrants, lubricants, suspending agents, binders, sweeteners, flavoring agents, preservatives, antioxidants, bases, and the like. The filler comprises: starch, pregelatinized starch, lactose, mannitol, chitin, microcrystalline cellulose, sucrose, etc.; the disintegrating agent comprises: starch, pregelatinized starch, microcrystalline cellulose, sodium carboxymethyl starch, crospolyvinylpyrrolidone, low-substituted hydroxypropylcellulose, croscarmellose sodium, etc.; the lubricant comprises: magnesium stearate, sodium lauryl sulfate, talc, silica, and the like; the suspending agent comprises: polyvinylpyrrolidone, microcrystalline cellulose, sucrose, agar, hydroxypropyl methylcellulose, and the like; the adhesive comprises: starch slurry, polyvinylpyrrolidone, hydroxypropylmethylcellulose, and the like; the sweetener comprises: saccharin sodium, aspartame, sucrose, sodium cyclamate, glycyrrhetinic acid, and the like; the flavoring agent comprises: sweeteners and various essences; the preservative comprises: parabens, benzoic acid, sodium benzoate, sorbic acid and its salts, benzalkonium bromide, chloroacetidine acetate, eucalyptus oil, etc.; the antioxidant includes: sodium sulfite, sodium metabisulfite, dibutyl phenol, sodium bisulfite, sodium thiosulfate, tert-butyl p-hydroxyanisole, thiourea, vitamin c, propyl gallate, alpha-tocopherol, ascorbyl palmitate; the matrix comprises: PEG6000, PEG4000, insect wax, etc. Therefore, it is within the scope of the present invention to add any other substance that contributes to the formation of a stable drug effect to the compound of the present invention.

The invention has the following beneficial effects:

the invention establishes a method for detecting the platinum compound with the structural formula (J) as a related substance in the lobaplatin quality standard so as to establish a lobaplatin quality detection system.

Drawings

FIG. 1-1A is an HPLC chromatogram (215nm) of the compound prepared in example 1-1 in an HPLC-MS structure confirmation assay;

FIG. 1-1B is an HPLC chromatogram (210nm) of the compound prepared in example 1-1 in an HPLC-MS structure confirmation assay;

FIG. 1-2A is an MS spectrum of the compound prepared in example 1-1 at an HPLC-MS structure confirmation detection time of 0.13 min;

FIG. 1-2B is an MS spectrum of the compound prepared in example 1-1, at an HPLC-MS structure confirmation detection time of 0.249 min;

FIG. 2 shows the preparation of the compound obtained in example 1-1¹An H-NMR spectrum;

FIG. 3 shows the preparation of the compound of example 1-1¹³A C-NMR spectrum;

FIG. 4 is a QNMR spectrum of the compound prepared in example 1-1;

FIG. 5 is a UV spectrum of the compound prepared in example 1-1; wherein, in the figure, the wavelength of peak 1 is 256.5nm, the absorbance is 0.7763, the wavelength of peak 2 is 226.5nm, the absorbance is 0.3026, the wavelength of peak 3 is 218.5nm, the absorbance is 0.2872, the wavelength of peak 4 is 382.0nm, the absorbance is 0.0038, the wavelength of peak 5 is 229.0nm, the absorbance is 0.2932, and the wavelength of peak 6 is 220.5nm, the absorbance is 0.2746;

FIG. 6 is an IR spectrum of the compound prepared in example 1-1;

FIG. 7 is a DSC spectrum of the compound prepared in example 1-1;

FIG. 8 is a typical spectrum of the compound prepared in example 1-1 of example 2;

FIG. 9-1A is a total ion chromatogram of an empty solvent in a methodological validation specificity experiment;

FIG. 9-1B is a MS spectrum of an empty solvent in a methodological validation specificity experiment;

FIG. 9-2A is a total ion chromatogram of the J-localization Solution (STD) of the substance of interest in lobaplatin in a methodological validation experiment;

FIG. 9-2B is a MS spectrum of J-site Solution (STD) of related substance in lobaplatin at t-16.174 min in a methodology validation specificity experiment;

FIG. 9-3A is an HPLC chromatogram (235nm) of test solution lobaplatin diastereomer I and lobaplatin diastereomer II in a methodological validation specificity experiment;

FIG. 9-3B is a total ion chromatogram of a test sample solution in a methodological validation specificity experiment;

FIG. 9-3C is a MS spectrum of the test solution at t-16.329 min in a methodology validation specificity experiment;

FIG. 9-4A is a total ion chromatogram of a sample spiked solution in a methodology validation specificity experiment;

FIG. 9-4B is a MS spectrum of a sample solution at t 16.174min for methodology validation in a specificity experiment;

FIG. 10 is a graph showing the linear relationship of related substance J in lobaplatin in methodological validation;

FIG. 11-1 is a graph showing the inhibitory activity of the compound prepared in example 1-1 of example 3 on lung cancer cell NCI-H460;

FIG. 11-2 is a graph showing the inhibitory activity of the positive control drug on lung cancer cell NCI-H460 in example 3;

FIG. 12-1 is a graph showing the inhibitory activity of the compound prepared in example 1-1 of example 3 on ovarian cancer cells SK-OV-3;

FIG. 12-2 is a graph showing the inhibitory activity of the positive control drug on the ovarian cancer cells SK-OV-3 in example 3;

FIG. 13-1 is a graph showing the inhibitory activity of the compound prepared in example 1-1 of example 3 on leukemia cells Jurkat clone 6-1;

FIG. 13-2 is a graph showing the inhibitory activity of the positive control drug on leukemia cells Jurkat Clone E6-1 in example 3;

FIG. 14-1 is a graph showing the inhibitory activity of the compound prepared in example 1-1 of example 3 on THP-1 of leukemia cells;

FIG. 14-2 is a graph showing the inhibitory activity of the positive control drug on THP-1 leukemia cells in example 3;

FIG. 15-1 is a graph showing the inhibitory activity of the compound prepared in example 1-1 of example 3 on SK-NEP-1, renal cancer cells;

FIG. 15-2 is a graph showing the inhibitory activity of the positive control drug on the renal cancer cell SK-NEP-1 in example 3.

Detailed Description

The invention provides a method for detecting substances related to lobaplatin. The following will describe, as specific examples, the preparation of the compound, the confirmation of the structure of the compound, the measurement of the antitumor activity of the compound, and the measurement of the amount of lobaplatin.

Herein, in the present invention, any substance affecting the purity of the drug is collectively referred to as "related substance affecting the quality of lobaplatin" or "related substance affecting the quality", and is simply referred to as "related substance", for example, a peak of related substance affecting the quality of lobaplatin appearing in an HPLC chromatogram peak for detecting the quality of lobaplatin, is simply referred to as "related substance peak"; the "related substance" in the present invention is sometimes an "impurity" known to those skilled in the art to affect the purity of the drug, however, the "related substance" in the present invention is not limited to the category of "impurity" but also includes substances having a certain anticancer activity even higher than that of lobaplatin, which belong to the category of substances related to lobaplatin with respect to the active molecule "lobaplatin", and the principles of their anticancer activity or other positive effects and functions in developing new drugs have not been fully studied.

The invention provides a method for detecting a lobaplatin related substance, wherein the lobaplatin related substance is a compound J, and the structure of the compound J is

Preferably, in the detection method, the compound J is prepared by a compound (2) or a compound (3) or a compound (4) of the following structural formula

Preferably, the compound of formula (J) is prepared from compound 4 of the following structural formula

Preferably, in the reaction for preparing the product by the compound 4, the obtained solution of the compound 4 is adjusted to be alkaline by an acidic regulator, and then is freeze-dried to obtain the final product; preferably, the acidity regulator is selected from one of p-toluenesulfonic acid or methanesulfonic acid, preferably p-toluenesulfonic acid; the concentration of the preferred aqueous solution of p-toluenesulfonic acid is 5 to 15 wt%, preferably 10 wt%; the pH value of the alkalescence is 7-8; preferably, after adjusting to be slightly alkaline, reacting for 15-25 hours at the temperature of 20-30 ℃; preferably, the reaction is carried out at 25 ℃ for 20 hours;

preferably, the compound 4 is prepared from a compound 3 with the following structural formula

Preferably, the solution of the compound 3 and the resin are mixed and stirred, and then the mixture is filtered to obtain a compound 4 solution; preferably, the mixing and stirring temperature of the compound 3 solution and the resin is 25-35 ℃; preferably, the stirring time is 0.5-2 hours; the stirring time is preferably 1 hour, and the stirring temperature is more preferably 30 ℃; more preferably, the resin is a resin that has been treated; preferably, the resin that has been treated is a resin treated with an aqueous solution of sodium hydroxide, preferably having a concentration of 1 to 2mol/L, preferably 1.5 mol/L.

Preferably, the compound 3 is prepared from a compound 2 with the following structural formula

Preferably, the compound 2 is added into water and a ketone solvent to obtain a material A, then a silver nitrate solution is added into the material A for reaction, and a solution of a compound 3 is obtained by filtration; preferably, the molar ratio of compound 2 to silver nitrate is 1 (1-2), preferably 1: 1.7; preferably, the reaction temperature is 25-35 ℃; more preferably, the reaction temperature is 30 ℃; preferably, the reaction time is 15 to 20 hours in a dark place; preferably, the reaction time is 18 hours in the absence of light; preferably, the ketone solvent is selected from one of acetone, methyl butanone, methyl ethyl ketone, or methyl isobutyl ketone, and acetone is more preferred.

In a preferred embodiment of the present invention, the compound J is derived from the compound (1)

In a preferred embodiment of the present invention, wherein the detection method is HPLC-MS method or HPLC method; preferably, the HPLC detection conditions of the HPLC-MS method are as follows: octadecylsilane chemically bonded silica is used as a filling agent, ammonium formate solution with the concentration of 18-22mmol/L is used as a mobile phase A, and methanol: acetonitrile in a volume ratio of 1 (0.8-1.2) as mobile phase B, preferably methanol: the acetonitrile is used as a mobile phase B with the volume ratio of 1:1, and gradient elution is carried out; preferably, the flow rate is 0.8-1.2ml, preferably 1.0ml per minute; the column temperature is 39-41 deg.C, preferably 40 deg.C.

Preferably, in the chromatogram of the system suitability test, the separation degree of the peak of the compound J from the peak of the adjacent related substance is not less than 1.5; preferably, the system suitability test solution is continuously fed for 6 times, and the relative standard deviation of the peak area of the compound J is not more than 10%.

Preferably, if compound J exists in the chromatogram of the test solution, the peak area of the compound J should not be larger than the peak area (0.5%) of compound J in the control solution, and 0.5% refers to the concentration of compound J in the control solution being 0.5% of that in the test solution.

Lobaplatin is known to have 2 isomers, lobaplatin diastereomer I and lobaplatin diastereomer II, and their structural formulas are shown below:

lobaplatin diastereomer I (RRS for short):

lobaplatin diastereomer II (SSS for short):