米格列奈钙3个主要降解杂质的研究

Study on Three Principal Degraded Impurities of Mitiglinide Calcium

目的：分离纯化米格列奈钙3个主要降解杂质（杂质A、B、C）,并进行结构鉴定,同时建立3个杂质的高效液相色谱含量测定方法。方法：以米格列奈钙为原料,经酸破坏后采用制备型高效液相色谱仪分离制备3个杂质单体,并采用红外光谱、质谱、^1H核磁共振谱、^13C核磁共振谱、电子轰击质谱、液相电喷雾电离质谱联用、旋光光谱对3个杂质进行结构鉴定;对3批米格列奈钙原料药进行3个杂质的含量测定,色谱柱为Agilent Extend C18,流动相为0.01 mol/L醋酸钠溶液-乙腈-三乙胺（60∶40∶0.1,p H 3.0）,流速为1.0 ml/min,检测波长为210 nm,进样量为20μl;另进行3个杂质及米格列奈钙的响应值试验。结果：经酸破坏后得到3个杂质A、B、C,经分离纯化后纯度分别为99.05%、98.87%、99.98%;经结构鉴定后分别确证为S-苄基丁二酸、S-苄基丁二酸-4-甲酯、米格列奈甲酯;建立的杂质含量测定方法学考察均符合相关要求,检测质量浓度线性范围分别为0.387 5~3.875、0.395~3.95、0.392 5~3.925μg/ml（r均为1.000 0）;3个杂质及米格列奈钙的响应值分别为2.316 1、2.636 1、2.617 8、2.620 4。结论：鉴定并确证了米格列奈钙的3个主要降解杂质,此3个杂质可以通过高效液相色谱法中的主成分自身对照法对其进行定量分析。

OBJECTIVE：To isolate and purify three principal degraded impurities of mitiglinide calcium（impurity A,B,C）and identify their structures,establish HPLC method for content determination of impurity A,B,C. METHODS：Mitiglinide calcium was used as raw material and reacted with acid;3 impurities were then separated by HPLC and their structures were elucidated by IR,MS,^1H NMR,^13 C NMR,LC-ESI-MS and ORD. 3 impurities of 3 batches of mitiglinide calcium were determined,and the determination was performed on Agilent Extend-C_（18） column with mobile phase consisted of 0.01 mol/L sodium acetate solution-acetonitrile-triethylamine（60 ∶ 40 ∶ 0.1,p H=3.0）at the flow rate of 1.0 ml/min. The detection wavelength was set at 210 nm and sample size was 20 μl. The response tests of 3 impurities and mitiglinide calcium were conducted. RESULTS：After treated with acid,impurity A,B,C had been obtained,and their purity were 99.05%,98.87%,99.98%,respectively after isolation and purification; after identifying the structure, 3 impurities were S-2-bezylsuccinic acid, S-2-bezylsuccinic acid-4-methyl ester, methyl（2S）-2-benzyl-3-（cis-hexahydroisoindolin-2-ylcarbonyl） propionate;methodological study of content determination of impurities were all up to the requirement. The linear range of impurity A,B,C were 0.387 5-3.875,0.395-3.95 and 0.392 5-3.925 μg/ml（all r were 1.000 0）. The response value of impurity A,B,C and mitiglinide calcium were 2.316 1,2.636 1,2.617 8 and 2.620 4,respectively. CONCLUSIONS：The structures of 3 principal degraded impurities of mitiglinide calcium have been identified and confirmed;the content of them can be determined by HPLC main component self-comparison method.