逍遥片原料药材及中间提取物赭曲霉毒素AUPLC-FLD检测方法的建立  

Establishment of UPLC-FLD method for determination of ochratoxin A in raw materials and intermediate extracts of Xiaoyao Tablets

作　　者：程实 王萍 孙巍 张磊 章顺楠 叶正良[4] CHENG Shi;WANG Ping;SUN Wei;ZHANG Lei;ZHANG Shunnan;YE Zhengliang(Graduate School,Tianjin University of Traditional Chinese Medicine,Tianjin 301608,China;Tasly Pharmaceutical Group Company Limited,International Department,Tianjin 300402,China;Tasly Pharmaceutical Group Company Limited,National&Local United Engineering Laboratory of TCM Advanced Manufacturing Technology,Tianjin 300402,China;Tasly Holding Group Company Limited,Tianjin 300402,China)

机构地区：[1]天津中医药大学研究生院,天津301608 [2]天士力医药集团股份有限公司国际产业中心,天津300402 [3]天士力医药集团股份有限公司中药先进制造技术国家地方联合工程实验室,天津300402 [4]天士力控股集团有限公司,天津300402

出　　处：《药物评价研究》2021年第3期512-516,共5页Drug Evaluation Research

基　　金：国家科技部“重大新药创制”科技重大专项(2018ZX09303024)。

摘　　要：目的建立免疫亲和柱前处理、超高效液相色谱串联荧光检测器(UPLC-FLD)分析,同时用于检测逍遥片原料药材柴胡、当归、炙甘草、炒白术、白芍及中间提取物1和2中赭曲霉毒素A(ochratoxin A,OTA)的方法。方法样品用80%甲醇-水溶液(体积比)超声提取,离心,免疫亲和柱纯化,ACQUITY UPLC BEH C18色谱柱分离,流动相A为0.5%冰醋酸溶液(pH值为2.97),流动相B为乙腈,梯度洗脱。激发波长310 nm,发射波长465 nm,柱温30℃,体积流量0.3 mL/min,进样体积2μL,荧光检测器检测。结果OTA在1.5~37.5 ng/mL显示良好的线性关系,R2≥0.9991。OTA加标回收率为92.36%~119.89%,RSD为2.50%~9.12%。样品在48 h内稳定。共检测42批次样品,33批呈阳性,柴胡和炙甘草被污染。结论建立的免疫亲和柱前处理、UPLC检测的方法可用于中药和中间提取物中OTA的检测,排除假阳性结果,适合痕量分析,结果准确可靠。Objective To establish an immunoaffinity pre-column processing,high performance liquid chromatography tandem fluorescence detector analysis,and to detect the raw materials of Xiaoyao Tablets Bupleuri Radix,Glycyrrhizae Radix Et Rhizoma,Radix Paeoniae Alba,Rhizoma Atractylodis Macrocephalae,Radix Angelicae Sinensis and Intermediate Extract 1,Extract 2 Method of Ochratoxin A.Method The sample was extracted with 80%methanol-water solution(volume ratio)ultrasonically,centrifuged,and purified by immunoaffinity column,separated by ACQUITY UPLC BEH C18 column,mobile phase A:0.5%glacial acetic acidwater solution(pH=2.97),mobile phase B:Acetonitrile elution,excitation wavelength 310 nm,emission wavelength 465 nm,column temperature 30℃,flow rate 0.3 mL/min,injection volume 2μL,fluorescence detector detection.Results Ochratoxin A showed a good linear relationship in the mass concentration range of 1.5 to 37.5μg/mL,and R2 was greater than 0.9991.The recovery rate of ochratoxin A was 92.36%—119.89%,and the relative standard deviation was 2.50%—9.12%.The sample is stable within 48 hours.A total of 42 batches of samples were detected and 33 batches were positive.Bupleuri Radix and Glycyrrhizae Radix Et Rhizoma were contaminated.Conclusion The established immunoaffinity column pretreatment and ultra-high performance liquid chromatography detection method is suitable for the detection method of OTA in traditional Chinese medicine and intermediate extracts,eliminating false positive results,suitable for trace analysis,conforming to international reporting standards,and the results are accurate and reliable.

关 键 词：逍遥片 赭曲霉毒素A 免疫亲和柱 限度 超高效液相色谱串联荧光检测器 

分 类 号：R917[医药卫生—药物分析学]