利用乌头类双酯型生物碱水解规律测定醇胺型乌头碱的含量
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摘要
目的:研究乌头类双酯型生物碱水解转化为醇胺型生物碱的转化率,水解乌头类双酯型生物碱对照品,利用水解转化率计算出水解原液中醇胺型生物碱的量,以此量作为醇胺型生物碱对照品的量,建立测定制川乌中乌头原碱、次乌头原碱、新乌头原碱含量的方法。方法:通过控制乌头碱、次乌头碱、新乌头碱3种双酯型生物碱的水解条件,得到乌头原碱、次乌头原碱、新乌头原碱。采用高效液相色谱四级杆飞行时间串联质谱(HPLC-QTOF-MS)法,使用Agilent ZORBAX Extend-C18RRHT(2.1 mm×50 mm,1.8μm)色谱柱,流速为0.21 ml·min~(-1),柱温为30℃,流动相为甲醇-水(含0.1%甲酸和2.5 mmol·L~(-1)醋酸铵)系统梯度洗脱,采用电喷雾离子(ESI)源,正离子方式检测。结果:在本文的水解条件下,乌头碱对于乌头原碱的转化率为99.64%;次乌头碱对于次乌头原碱的转化率为99.94%;新乌头碱对于新乌头原碱的转化率为99.57%。HPLC-QTOF-MS方法学表明,乌头原碱、次乌头原碱、新乌头原碱在一定范围内具有较好的线性关系,r值均大于0.999 1,精密度、重复性和稳定性的RSD都小于5%,加样回收率均在99.43%~100.10%之间。结论:本方法操作简便、专属性和重复性较好,在缺乏对照品的情况下,可以为乌头类中药材质量控制提供依据。
Objective: To investigate the hydrolysis conversion rate of alcohol amine-diterpene alkaloids from aconitum alkaloids,hydrolyze aconitum alkaloids reference substance,calculate the amount of alcohol amine-diterpene alkaloids in the hydrolysis solution by the hydrolysis conversion rate,which is used as the amount of alcohol amine-diterpene alkaloids reference substance,and establish a content determination method for aconine,hypaconitine and aconine in Aconiti radix cocta. Methods: Through controlling the hydrolysis conditions of aconitine,hypaconitine and mesaconitine,aconine,hypaconitine and aconine were obtained. The determination was performed on an Agilent ZORBAX Extend-C18RRHT( 2. 1 mm × 50 mm,1. 8 μm) column with the mobile phase consisting of methanol( A)-water( B containing 0. 1% formic acid and 2. 5 mmol·L~(-1)ammonium acetate) with gradient elution by HPLC-QTOF-MS. The flow rate was 0. 21 ml·min~(-1). The column temperature was 30 ℃. MS instrument was equipped with an ESI + ion source. Results:Under the hydrolysis conditions of this study,the conversion rate of aconine from aconitine was 99. 64%; the conversion rate of hypaconitine from hypaconine was 99. 94%; the conversion rate of mesaconitine from mesaconine was 99. 57%. The HPLC-QTOF-MS methodological investigation showed the 3 kinds of alcohol amine-diterpene alkaloids were with good linearity( r > 0. 999 1). The RSD of the precision,repeatability and stability tests were less than 5%. The average recoveries were within the range of 99. 43%~(-1)00. 10%. Conclusion: The validated method is simple,specific,reliable and reproducible. In the absence of reference substance,it can be used for the quality control of the herbs of Aconitum L. species.
Objective: To investigate the hydrolysis conversion rate of alcohol amine-diterpene alkaloids from aconitum alkaloids,hydrolyze aconitum alkaloids reference substance,calculate the amount of alcohol amine-diterpene alkaloids in the hydrolysis solution by the hydrolysis conversion rate,which is used as the amount of alcohol amine-diterpene alkaloids reference substance,and establish a content determination method for aconine,hypaconitine and aconine in Aconiti radix cocta. Methods: Through controlling the hydrolysis conditions of aconitine,hypaconitine and mesaconitine,aconine,hypaconitine and aconine were obtained. The determination was performed on an Agilent ZORBAX Extend-C18RRHT( 2. 1 mm × 50 mm,1. 8 μm) column with the mobile phase consisting of methanol( A)-water( B containing 0. 1% formic acid and 2. 5 mmol·L~(-1)ammonium acetate) with gradient elution by HPLC-QTOF-MS. The flow rate was 0. 21 ml·min~(-1). The column temperature was 30 ℃. MS instrument was equipped with an ESI + ion source. Results:Under the hydrolysis conditions of this study,the conversion rate of aconine from aconitine was 99. 64%; the conversion rate of hypaconitine from hypaconine was 99. 94%; the conversion rate of mesaconitine from mesaconine was 99. 57%. The HPLC-QTOF-MS methodological investigation showed the 3 kinds of alcohol amine-diterpene alkaloids were with good linearity( r > 0. 999 1). The RSD of the precision,repeatability and stability tests were less than 5%. The average recoveries were within the range of 99. 43%~(-1)00. 10%. Conclusion: The validated method is simple,specific,reliable and reproducible. In the absence of reference substance,it can be used for the quality control of the herbs of Aconitum L. species.
引文
1李谦,过立农,郑健,等.乌头属药用植物的研究进展[J].药物分析杂志,2016,36(7):1129-1149
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5王瑞,刘芳,孙毅坤,等.不同附子炮制品中乌头碱、新乌头碱、次乌头碱含量的HPLC测定[J].药物分析杂志,2006,26(10):1361-1363
6陆燕萍,巩晓宇,陈秋燕,等.乌头类碱类生物碱的含量测定概述[J].中国药业,2016,25(7):1-3
7吴成凤,李国卫,王樟根,等.LC-MS/MS法同时测定乌头汤中乌头碱型生物碱含量[J].中药新药与临床药理,2015,26(6):833-837
8程丽丽,许妍妍,Nyirimigabo E,等.一测多评测定乌头类药材中生物碱[J].现代药物与临床,2013,28(4):528-534
9张毅,翁代群,王海军,等.乌头碱的水解动力学研究[J].中草药,2009,40:120-122
10包懿,宋凤瑞,刘志强,等.乌头碱类双酯型二萜生物碱水解反应的电喷雾质谱分析[J].质谱学报,2009,30(1):1-5
11马鸿雁,李楠,杨明.乌头碱水解实验和热力学研究[J].成都中医药大学学报,2005,28(3):57-59
12张小龙,吴先富,王昆,等.HPLC-QTOF-MS法测定风湿骨痛胶囊中9个乌头类生物碱的含量[J].药物分析杂志,2015,35(3):414-419
13洪波,仇永清.附子中双酯型乌头碱类成分水解减毒机理的密度泛函理论研究[J].分子科学学报,2008,24(3):216-219
14刘文龙,刘凤强,宋凤瑞,等.乌头类双酯型生物碱组分转化为单酯水解性及脂型生物碱组分的研究[J].高等学校化学学报,2011,32(3):717-720
2杨武斌,王平.乌头碱药理作用及毒性研究进展[J].时珍国医国药,2014,25(2):427-429
3蔡超群,杨春华,梁敬钰,等.乌头属二萜生物碱成分构效关系研究进展[J].海峡药学,2013,25(3):1-5
4郑琴,陆浩伟,郝伟伟.乌头类双酯型生物碱水解转化规律及含量计算方法研究[J].中国药学杂志,2011,46(9):652-656
5王瑞,刘芳,孙毅坤,等.不同附子炮制品中乌头碱、新乌头碱、次乌头碱含量的HPLC测定[J].药物分析杂志,2006,26(10):1361-1363
6陆燕萍,巩晓宇,陈秋燕,等.乌头类碱类生物碱的含量测定概述[J].中国药业,2016,25(7):1-3
7吴成凤,李国卫,王樟根,等.LC-MS/MS法同时测定乌头汤中乌头碱型生物碱含量[J].中药新药与临床药理,2015,26(6):833-837
8程丽丽,许妍妍,Nyirimigabo E,等.一测多评测定乌头类药材中生物碱[J].现代药物与临床,2013,28(4):528-534
9张毅,翁代群,王海军,等.乌头碱的水解动力学研究[J].中草药,2009,40:120-122
10包懿,宋凤瑞,刘志强,等.乌头碱类双酯型二萜生物碱水解反应的电喷雾质谱分析[J].质谱学报,2009,30(1):1-5
11马鸿雁,李楠,杨明.乌头碱水解实验和热力学研究[J].成都中医药大学学报,2005,28(3):57-59
12张小龙,吴先富,王昆,等.HPLC-QTOF-MS法测定风湿骨痛胶囊中9个乌头类生物碱的含量[J].药物分析杂志,2015,35(3):414-419
13洪波,仇永清.附子中双酯型乌头碱类成分水解减毒机理的密度泛函理论研究[J].分子科学学报,2008,24(3):216-219
14刘文龙,刘凤强,宋凤瑞,等.乌头类双酯型生物碱组分转化为单酯水解性及脂型生物碱组分的研究[J].高等学校化学学报,2011,32(3):717-720