【摘要】 建立简单、灵敏、快速的高效液相色谱法,对甘草酸单铵盐原料药中主成分异构体进行分离以及同时与有关物质的分离检测,并对各待测物结构进行鉴定。参考欧洲药典7.0版(EP7.0)、英国药典2012版(BP2012)、中国国家药品标准(WS1-XG-2002)及国内外相关文献,对流动相的组成进行选择,通过优化流动相中盐的浓度、水系溶液的pH值、有机相的加入比例、柱温及流速等参数,确定了最佳的色谱条件:采用Durashell C18色谱柱(250 mm×4.6 mm,5μm),以0.01 mol.mL 1高氯酸铵(氨水调至pH 8.2)甲醇(48∶52)为流动相,流速0.8 mL.min 1,检测波长254 nm,柱温50℃,进样量10μL。利用质谱、核磁共振谱、紫外光谱和液相色谱等实验技术对获得的18α-甘草酸、18β-甘草酸、有关物质A、有关物质B对照品结构进行确证,以保证其结构的准确无误;采用高效液相色谱法对法定机构提供的甘草酸单铵盐对照品进行定位,并采用二级管阵列检测器分析测定各待测物色谱峰的纯度。在优化的色谱条件下,18α-甘草酸、18β-甘草酸、有关物质A和B在0.50～100μg.mL 1内线性关系良好(r=0.999 9),检出限分别为0.15、0.10、0.10和0.15μg.mL 1。本方法检测灵敏、重现性好、结果准确可靠,可用于甘草酸单铵盐原料药主成分异构体及其与有关物质的同时分离检测。在EP7.0、BP2012收载的甘草酸单铵盐质量标准中,对主成分异构体的分离以及对色谱图中相对保留时间约为1.2的物质的结构归属有待商榷。本方法能够为甘草酸原料药、制剂以及中药材的质量标准的制定、质量控制及结构鉴定提供依据。更多还原

【Abstract】 A simple,fast and sensitive analytical method for the simultaneous separation and detection of 18α-glycyrrhizinic acid,18β-glycyrrhizinic acid,related substance A and related substance B by RP-HPLC and drug quality standard was established.The structures of principal component isomer and related substances of raw material drug of ammonium glycyrrhizinate have been confirmed.Reference European Pharmacopoeia EP7.0 version,British Pharmacopoeia 2012 version,National Drug Standards of China(WS 1-XG-2002),domestic and international interrelated literature were referred to select the composition of mobile phase.The experimental parameters including salt concentration,pH,addition quantities of organic solvent,column temperature and flow rate were optimized.Finally,the assay was conducted on a Durashell-C 18 column(250 mm × 4.6 mm,5 μm) with 0.01 mol.mL 1 ammonium perchlorate(add ammonia to adjust the pH value to 8.2) methanol(48∶52) as mobile phase at the flow rate of 0.8 mL.min 1,and the detection wavelength was set at 254 nm.The column temperature was 50 ℃ and the injection volume was 10 μL.The MS,NMR,UV and RP-HPLC were used to confirm the structures of principal component isomer and related substances of raw material drug of ammonium glycyrrhizinate.Under the optimized separation conditions,the calibration curves of 18α-glycyrrhizinic acid,18βglycyrrhizinic acid,related substance A and related substance B showed good linearity within the concentration of 0.50 100 μg.mL 1(r = 0.999 9).The detection limits for 18α-glycyrrhizinic acid,18β-glycyrrhizinic acid,related substance A and related substance B were 0.15,0.10,0.10,0.15 μg.mL 1 respectively.The method is sensitive,reproducible and the results are accurate and reliable.It can be used for chiral resolution of 18α-glycyrrhizinic acid,18β-glycyrrhizinic acid,and detection content of principal component and related substances of raw material drug of ammonium glycyrrhizinate.It is concluded that the separation of principal component isomer of raw material drug of ammonium glycyrrhizinate and the validity of the substance’s structure assignments of retention time being 1.2 in the European pharmacopoeia EP7.0 version,British pharmacopoeia 2012 version remains open to question.It may be of practical value for the quality control of raw material drug,preparation,and Chinese herbal medicine of ammonium glycyrrhizinate.更多还原