阅读说明：本技术 洛铂中有关物质的检测 (Detection of related substances in lobaplatin ) 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：本发明涉及一种洛铂中有关物质的检测。本发明提供了一种洛铂中有关物质的检测方法,所述有关物质选自化合物K、化合物I、化合物F或/和铂类物质M的一种或两种以上,其中：所述化合物F的结构为<Image he="179" wi="331" file="DDA0001999837410000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>化合物K的结构为<Image he="197" wi="428" file="DDA0001999837410000012.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>化合物I的结构为<Image he="216" wi="464" file="DDA0001999837410000013.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>铂类物质M为<Image he="205" wi="434" file="DDA0001999837410000014.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>或<Image he="208" wi="431" file="DDA0001999837410000015.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>和<Image he="218" wi="474" file="DDA0001999837410000016.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>或<Image he="213" wi="457" file="DDA0001999837410000017.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>的混合物,所述检测方法为HPLC法或者HPLC-MS法,所述HPLC法的检测条件为：用十八烷基硅烷键合硅胶为填充剂,以8-12mol/L的醋酸铵溶液为流动相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,所述检测方法的灵敏度高,专属性强,重复性好,准确度高,在洛铂药物的质量可控上具有重大的技术进步。(The invention relates to detection of related substances in lobaplatin. The invention provides a method for detecting related substances in lobaplatin, wherein the related substances are selected from one or more than two of a compound K, a compound I, a compound F or/and a platinum substance M, and the method comprises the following steps: the compound F has the structure The structure of the compound K is The structure of the compound I is The platinum compound M is Or And or The detection method is an HPLC method or an HPLC-MS method, and the detection conditions of the HPLC method are as follows: octadecylsilane chemically bonded silica is used as a filling agent, 8-12mol/L ammonium acetate solution 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, gradient elution is carried out, and the detection method has the advantages of high sensitivity, strong specificity, good repeatability and high accuracy, and has great technical progress in the quality control of the lobaplatin medicament.)

1. A method for detecting related substances in lobaplatin is characterized in that the related substances are selected from one or more than two of a compound K, a compound I, a compound F and/or a platinum substance M, wherein:

the compound F has the structure

The structure of the compound K isThe structure of the compound I is

The platinum compound M is A mixture of (a).

2. The assay of claim 1 wherein said compound I is via an intermediatePreparation of compound K byIs prepared by

3. The detection method according to claim 1 or 2, wherein the detection method is an HPLC method or an HPLC-MS method.

4. The detection method according to claim 3, wherein the detection conditions of the HPLC method are: octadecylsilane chemically bonded silica is used as a filling agent, 8-12mol/L ammonium acetate solution 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 gradient elution 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;

preferably, the detection wavelength of the compound F is 219-221nm, and the detection wavelengths of the compound I, the compound K and the platinum-based substance M are 234-236 nm; the flow rate is 0.5-1.5ml per minute, and the column temperature is 38-42 ℃.

5. The detection method according to claim 4, wherein the column temperature is 39-41 ℃, preferably 40 ℃; preferably, the concentration of the ammonium acetate solution is 9-11mmol, preferably 10 mmol; preferably, the detection wavelength of the compound F is 220nm, the detection wavelength of the compound I, the compound K and the platinum-based substance M is 235nm, the flow rate is 1ml per minute, the methanol: the volume ratio of acetonitrile is 1:1.

6. The detection method according to any one of claims 3 to 5, wherein the peak areas of the compound F, the compound I and the compound K in the sample solution do not exceed the peak area of the main component in the control solution as calculated by the main component self-control method with a correction factor added thereto; preferably, the peak area of the platinum-containing substance M in the sample solution should not exceed 10 times the peak area of the main component in the control solution, as calculated by the peak area of the main component self-control method without adding a correction factor.

7. The detection method according to any one of claims 1 to 6, wherein, if a peak of the substance of interest is present in the chromatogram of the test solution, the peak of the chromatogram of the substance of interest is identified as: the relative retention time of the compound F is 1.02-1.10, the relative retention time of the compound I is 1.10-1.20, the relative retention time of the compound K is 0.20-0.25, and the relative retention time of the platinum-based substance M is 0.85-0.95; preferably, the relative retention time of compound F is 1.06, the relative retention time of compound I is 1.15, the relative retention time of compound K is 0.23, and the relative retention time of platinum species M is 0.89.

8. The assay of any one of claims 1-7 wherein the correction factor for compound F is from 1.8 to 1.9, the correction factor for compound I is from 0.8 to 0.9, and the correction factor for compound K is from 0.4 to 0.5; preferably, the correction factor for compound F is 1.86, the correction factor for compound I is 0.88, and the correction factor for compound K is 0.45.

9. The detection method according to any one of claims 1 to 8, wherein the degree of separation of the peak of the substance of interest from the peaks of adjacent substances of interest is not less than 1.5.

10. The detection method according to any one of claims 1 to 9, 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 related substances in 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-cyclobutanebis (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 invention aims to provide a novel detection method which can simultaneously detect a plurality of related substances in lobaplatin.

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 comprises selecting a proper analysis method, accurately distinguishing and determining the content of the related substances, and determining the reasonable limit of the related substances by combining the results of pharmaceutical, toxicological and clinical researches. 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 related substances in lobaplatin, wherein the related substances are selected from one or more than two of a compound K, a compound I, a compound F and/or a platinum substance M, wherein:

the compound F has the structure

The structure of the compound K is

The structure of the compound I is

The platinum compound M is

A mixture of (a).

Preferably, forThe detection method as described above, wherein the compound I is passed through an intermediate

Preparation of compound K byIs prepared byAnd (4) preparing.

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

Preferably, in the detection method, the detection conditions of the HPLC method are: octadecylsilane chemically bonded silica is used as a filling agent, 8-12mol/L ammonium acetate solution 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 gradient elution 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); or 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), 25 to 33 minutes (or 24 to 32 minutes);

preferably, the detection wavelength of the compound F is 219-221nm, and the detection wavelengths of the compound I, the compound K and the platinum-based substance M are 234-236 nm; the flow rate is 0.5-1.5ml per minute, and the column temperature is 38-42 ℃.

Preferably, in the detection method, the column temperature is 39-41 ℃, preferably 40 ℃; preferably, the concentration of the ammonium acetate solution is 9-11mmol, preferably 10 mmol; preferably, the detection wavelength of the compound F is 220nm, the detection wavelength of the compound I, the compound K and the platinum-based substance M is 235nm, the flow rate is 1ml per minute, the methanol: the volume ratio of acetonitrile is 1:1.

Preferably, in the detection method described above, the peak areas of the compound F, the compound I and the compound K in the sample solution are not more than the peak area of the main component in the control solution, as calculated by the main component self-control method with the correction factor added, in the peak areas; preferably, the peak area of the platinum-containing substance M in the sample solution should not exceed 10 times the peak area of the main component in the control solution, as calculated by the peak area of the main component self-control method without adding a correction factor.

Preferably, in the detection method, if a peak of the relevant substance exists in the chromatogram of the sample solution, the peak of the chromatogram in the chromatogram is identified by the relevant substance, and the peak is located: the relative retention time of the compound F is 1.02-1.10, the relative retention time of the compound I is 1.10-1.20, the relative retention time of the compound K is 0.20-0.25, and the relative retention time of the platinum-based substance M is 0.85-0.95; preferably, the relative retention time of compound F is 1.06, the relative retention time of compound I is 1.15, the relative retention time of compound K is 0.23, and the relative retention time of platinum species M is 0.89.

Preferably, the detection method described above, wherein the correction factor for compound F is 1.8 to 1.9, the correction factor for compound I is 0.8 to 0.9, and the correction factor for compound K is 0.4 to 0.5; preferably, the correction factor for compound F is 1.86, the correction factor for compound I is 0.88, and the correction factor for compound K is 0.45.

Preferably, in the detection method described above, the degree of separation between the peak of the substance of interest and the peak of the adjacent substance of interest is not less than 1.5.

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

The invention has the following beneficial effects:

the invention confirms the structures of the compound I, the compound K, the compound F and the compound G and the compound L in the platinum substance M, confirms the related substances in the lobaplatin, establishes a complete lobaplatin quality detection system, and can simultaneously detect a plurality of related substances in the lobaplatin. The method has the advantages of high sensitivity, strong specificity, good repeatability and high accuracy, and has great technical progress on the quality control of the lobaplatin medicament.

Drawings

FIG. 1-1A is an HPLC-MS spectrum (wavelength 215nm) in a structure confirmation assay of Compound I in example 1;

FIGS. 1-1B are HPLC-MS spectra (wavelength 210nm) in the structure confirmation assay of Compound I in example 1

FIGS. 1-2 are MS spectra in a structure confirmation assay of compound I in example 1 using HPLC-MS;

FIG. 2 is a drawing of Compound I of example 1¹An H-NMR spectrum;

FIG. 3 is a drawing of Compound I of example 1¹³A C-NMR spectrum;

FIG. 4 is a UV absorption spectrum of Compound I of example 1, wherein the wavelength at 1 is 220.5nm, the wavelength at 2 is 196.5nm, and the wavelength at 3 is 205.5 nm;

FIG. 5 is an infrared absorption spectrum of Compound I in example 1;

FIG. 6 is a DSC plot of Compound I of example 1;

FIG. 6A is an HPLC chromatogram of Compound I of example 1;

FIG. 7 is an HPLC chromatogram of the reaction mixture in example 2;

FIG. 8-1A is an HPLC chromatogram (215nm) of Compound K of example 2 in HPLC-MS structure confirmation;

FIG. 8-1B is an HPLC chromatogram (210nm) of Compound K of example 2 in HPLC-MS structure confirmation;

FIG. 8-2 is an MS spectrum of Compound K of example 2 in HPLC-MS structure confirmation;

FIG. 9 is of Compound K of example 2¹H NMR spectrum;

FIG. 10 is of Compound K of example 2¹³A C-NMR spectrum;

FIG. 11 is a Q NMR spectrum of compound K of example 2;

FIG. 12 is an HPLC chromatogram of Compound K of example 2;

FIG. 13 is a UV spectrum of Compound K of example 2;

FIG. 14 is an IR spectrum of Compound K of example 2;

FIG. 15 is a DSC plot of Compound K of example 2;

FIG. 16-1A is an HPLC-MS spectrum (wavelength 215nm) in a structure confirmation assay of Compound G1 in example 4;

FIG. 16-1B is an HPLC-MS spectrum (wavelength 210nm) in a structure confirmation assay of Compound G1 in example 4;

FIG. 16-2 is an MS spectrum in a structure confirmation assay combined with HPLC-MS of Compound G1 in example 4;

FIG. 17 is a drawing of compound G1 from example 4¹An H-NMR spectrum;

FIG. 18 is a drawing of Compound G1 in example 4¹³A C-NMR spectrum;

FIG. 19 is a UV spectrum of Compound G1 of example 4;

FIG. 20 is an IR spectrum of compound G1 in example 4;

FIG. 21 is a DSC spectrum of compound G1 in example 4;

FIG. 22 is a Q-NMR spectrum of compound G1 in example 4;

FIG. 23 is an HPLC chromatogram of Compound G1 in example 4;

FIG. 24-1A is an HPLC chromatogram (wavelength 215nm) in the structure confirmation assay for the compound G2 in example 4 used in combination with HPLC-MS

FIG. 24-1B is an HPLC chromatogram (wavelength 210nm) in the structure confirmation assay of the compound G2 in example 4 in combination with HPLC-MS

FIG. 24-2 is an MS spectrum in a structure confirmation assay combined with HPLC-MS of Compound G2 in example 4;

FIG. 25 is a drawing of Compound G2 in example 4¹An H-NMR spectrum;

FIG. 26 is a drawing of Compound G2 of example 4¹³A C-NMR spectrum;

FIG. 27 is a UV spectrum of Compound G2 of example 4;

FIG. 28 is an IR spectrum of compound G2 in example 4;

FIG. 29 is a DSC spectrum of compound G2 in example 4;

FIG. 30 is a Q-NMR spectrum of compound G2 in example 4;

FIG. 31 is an HPLC chromatogram of Compound G2 in example 4;

FIG. 32-1A is an HPLC-MS spectrum (wavelength 215nm) in a structure confirmation assay of Compound L1 in example 6;

FIG. 32-1B is an HPLC-MS spectrum (wavelength 210nm) in a structure confirmation assay of Compound L1 in example 6;

FIG. 32-2 is an MS spectrum in a structure confirmation assay of HPLC-MS combination of Compound L1 in example 6;

FIG. 33 is a 1H-NMR spectrum of compound L1 in example 6;

FIG. 34 is a drawing of Compound L1 of example 6¹³A C-NMR spectrum;

FIG. 35 is a UV spectrum of Compound L1 in example 6;

FIG. 36 is an IR spectrum of Compound L1 in example 6;

FIG. 37 is a DSC spectrum of compound L1 of example 6;

FIG. 38 is a QNMR spectrum of compound L1 in example 6;

FIG. 39 is an HPLC chromatogram of Compound L1 in example 6;

FIG. 40-1A is an HPLC-MS spectrum (wavelength 215nm) in a structure confirmation assay of Compound L2 in example 6;

FIG. 40-1B is an HPLC-MS spectrum (wavelength 210nm) in a structure confirmation assay of Compound L2 in example 6;

FIG. 40-2 is an MS spectrum in an HPLC-MS combined structure confirmation assay of Compound L2 in example 6;

FIG. 41 is a drawing of Compound L2 in example 6¹An H-NMR spectrum;

FIG. 42 is a drawing of compound L2 of example 6¹³A C-NMR spectrum;

FIG. 43 is a UV spectrum of Compound L2 of example 6;

FIG. 44 is an IR spectrum of Compound L2 in example 6;

FIG. 45 is a DSC spectrum of compound L2 in example 6;

FIG. 46 is a QNMR spectrum of compound L2 in example 6;

FIG. 47 is an HPLC chromatogram of Compound L2 in example 6;

FIG. 48 is an HPLC chromatogram of the filter cake obtained in the second stage of example 7;

FIG. 49 is an HPLC chromatogram of Compound F of example 8;

FIG. 50 is an MS spectrum from HPLC-MS structure confirmation of Compound F in example 8;

FIG. 51 is a photograph of Compound F of example 8¹H NMR spectrum;

FIG. 52 is a photograph of Compound F of example 8¹³A C-NMR spectrum;

FIG. 53 is the Q NMR spectrum of compound F in example 8;

FIG. 54-1 is a typical map of the identification of Compound F in example 9-1;

FIG. 54-2 is a typical map of Compound I and platinum-based substance M in example 9-1;

FIG. 54-3 is a typical spectrum of Compound K, platinum-based substance M and Compound I in example 9-1;

FIG. 55-1 is a schematic diagram showing the specificity of a blank solution having a wavelength of 220nm in example 10;

FIG. 55-2 is a schematic diagram showing the specificity of a blank solution having a wavelength of 235nm in example 10;

FIG. 55-3 is a schematic diagram showing the specificity of the resolution solution (RS-1) of example 10 having a wavelength of 220 nm;

FIG. 55-4 is a schematic diagram showing the specificity of the resolution solution (RS-1) of example 10 having a wavelength of 235 nm;

FIG. 55-5 is a schematic diagram showing the specificity of the resolution solution (RS-2) of example 10 having a wavelength of 235 nm;

FIG. 56-1 is a graph showing the linear relationship between the self-control of lobaplatin at 235nm wavelength in example 10;

FIG. 56-2 is a graph showing the linear relationship between the self-control of lobaplatin at a wavelength of 220nm in example 10;

FIG. 56-3 is a schematic linear relationship of Compound F in example 10;

FIG. 56-4 is a schematic linear relationship of Compound I of example 10;

FIGS. 56-5 are graphs showing the linear relationship of Compound K in example 10;

FIG. 57-1 is a graph showing the inhibitory activity of Compound I on NCI-H460 in example 11;

FIG. 57-2 is a graph showing the inhibitory activity of STSP on NCI-H460 in example 11;

FIG. 58-1 is a graph showing the inhibitory activity of Compound I on 95-D in example 11;

FIG. 58-2 is a graph showing the inhibitory activity of STSP on 95-D in example 11;

FIG. 59-1 is a graph showing the AGS inhibitory activity of Compound I in example 11;

FIG. 59-2 is a graph showing the AGS inhibitory activity of STSP in example 11;

FIG. 60-1 is a schematic representation of the inhibitory activity of compound I on OVCAR-3 in example 11;

FIG. 60-2 is a schematic representation of the inhibitory activity of STSP on OVCAR-3 in example 11;

FIG. 61-1 is a graph showing the inhibitory activity of Compound I in example 11 on HL-60;

FIG. 61-2 is a graph showing the inhibitory activity of STSP on HL-60 in example 11;

FIG. 62-1 is a graph showing the inhibitory activity of Compound I in example 11 on THP-1;

FIG. 62-2 is a graph showing the THP-1 inhibitory activity of STSP in example 11;

FIG. 63-1 is a graph showing the inhibitory activity of Compound I of example 11 on Jurkat Clone E6-1;

FIG. 63-2 is a graph showing the inhibitory activity of STSP on Jurkat Clone E6-1 in example 11;

FIG. 64-1 is a graph showing the inhibitory activity of Compound I in example 11 on DU 145;

FIG. 64-2 is a graph showing the inhibitory activity of STSP on DU 145 in example 11;

FIG. 65-1 is a graphic representation of the inhibitory activity of compound I on SK-NEP-1 in example 11;

FIG. 65-2 is a schematic diagram showing the inhibitory activity of STSP on SK-NEP-1 in example 11;

Detailed Description

The invention provides a method for detecting a plurality of related substances in lobaplatin. The preparation of the lobaplatin-related substances I, F, K and M and the like found in the present invention, the confirmation of the structure of these novel substances, and the antitumor activity of the compounds of these novel substances will be described below by way of examples; the method for detecting related substances in lobaplatin of the present invention will be further described in detail by examples.

Wherein, the platinum substance M is a mixture of a compound L1 or L2 and a compound G1 or G2, L1 is one of the following structures, when L1 is one of the structures, the other structure is L2, and the specific structure is L2

Or

Since the single crystal cultivation failed and there was no single crystal diffraction check data, the absolute configuration of compounds L1 and L2 could not be confirmed, but two chiral enantiomers could be confirmed, and the other check confirmation data except for single crystal diffraction could confirm only the related substances as two compounds, but could not finally confirm the specific compounds.

G1 is one of the following structures, that is, when G1 is one of the structures, the other structure is G2, specifically:

or

Since the single crystal cultivation failed and there was no single crystal diffraction check data, the absolute configuration of compounds G1 and G2 could not be confirmed, but two chiral enantiomers could be confirmed, and the other check confirmation data except for single crystal diffraction could confirm only the related substances as two compounds, but could not finally confirm the specific compounds.

The invention provides a method for detecting related substances in lobaplatin, wherein the related substances are selected from one or more than two of a compound K, a compound I, a compound F and/or a platinum substance M, wherein:

the compound F has the structure

The structure of the compound K is

The structure of the compound I is

The platinum compound M is

A mixture of (a).

In a preferred embodiment of the present invention, wherein the detection method is HPLC method or HPLC-MS method; preferably, the detection conditions of the HPLC method are as follows: octadecylsilane chemically bonded silica is used as a filling agent, 8-12mol/L ammonium acetate solution 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 gradient elution 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); or 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), 25 to 33 minutes (or 24 to 32 minutes);

preferably, the detection wavelength of the compound F is 219-221nm, and the detection wavelengths of the compound I, the compound K and the platinum-based substance M are 234-236 nm; the flow rate is 0.5-1.5ml per minute, and the column temperature is 38-42 ℃.

In a preferred embodiment of the present invention, wherein the peak areas of the compound F, the compound I and the compound K in the test solution do not exceed the peak area of the main component in the control solution, as calculated by the main component self-control method with the addition of the correction factor; preferably, the peak area of the platinum-containing substance M in the sample solution should not exceed 10 times the peak area of the main component in the control solution, as calculated by the peak area of the main component self-control method without adding a correction factor.

Preferably, if a peak of a related substance exists in the chromatogram of the test solution, the peak of the chromatogram in the chromatogram is identified by the related substance for positioning: the relative retention time of the compound F is 1.02-1.10, the relative retention time of the compound I is 1.10-1.20, the relative retention time of the compound K is 0.20-0.25, and the relative retention time of the platinum-based substance M is 0.85-0.95; preferably, the relative retention time of compound F is 1.06, the relative retention time of compound I is 1.15, the relative retention time of compound K is 0.23, and the relative retention time of platinum species M is 0.89.

In particular with respect to the retention time of diastereomer II of lobaplatin. Specifically, as the lobaplatin compound, 2 isomers, lobaplatin diastereomer I and lobaplatin diastereomer II, which are represented by the following structural formulae, are known:

lobaplatin diastereomer I (RRS for short):

lobaplatin diastereomer II (SSS for short):