生附子煎煮过程中生物碱含量变化及水解机理
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摘要
目的观察生附子煎煮过程中生物碱成分含量的变化和水解,探索生附子煎煮减毒的机理。方法采用高效液相色谱(HPLC)法测定生附片煎煮前后7种成分的含量,比较其含量变化;采用新乌头碱、次乌头碱、乌头碱、苯甲酰新乌头原碱、苯甲酰次乌头原碱、苯甲酰乌头原碱对照品模拟煎煮过程,考察不同温度和水解时间对6种单、双酯型生物碱含量的影响,观察酯型生物碱煎煮前后水解程度,分析其降解率和稳定性,并确定其水解产物。结果生附片煎煮前后含量测定HPLC图中确定8个色谱峰,生附片中3种双酯型生物碱总量为1.756 mg/g,3种单酯型生物碱总量为0.724 mg/g,煎煮1 h后,其含量分别为0.026 mg/g和1.961 mg/g;对照品水解试验HPLC图中确定10个色谱峰,新乌头碱、次乌头碱、乌头碱对照品50℃水解30 min,其降解率分别为18.72%,4.85%,28.91%,50℃水解1 h,其降解率分别为33.3%,6.99%,40.14%,100℃水解30 min以上,其降解率均为100.00%;苯甲酰新乌头原碱、苯甲酰次乌头原碱、苯甲酰乌头原碱对照品50℃水解1 h后的含量基本不变,100℃水解1 h,其降解率分别为63.31%,42.36%,64.15%,100℃水解2 h以上,其降解率均为100.00%;根据对照品水解过程中的降解率大小,确定其稳定性大小依次为苯甲酰次乌头原碱>苯甲酰新乌头原碱>苯甲酰乌头原碱>次乌头碱>新乌头碱>乌头碱。结论附子中双酯型生物碱在50℃下即发生水解,100℃下30 min可水解完全,单酯型生物碱在50℃下较稳定,100℃下2 h即可水解完全。生附片煎煮过程是一个溶出与降解的动态过程,煎煮1 h后双酯型生物碱基本消失。
Objective To observe the content of alkaloid of Radix Aconite Lateralis and the raw of hydrolysis,and toxicity reducing mechanism during decocting process. Methods HPLC method was used to determine the content of seven components before and after decocting and compare the change of content; the boiling process was simulated,the degradation rate and stability of mesaconitine,hypaconitine,aconine,benzoylmesaconine,benzoylaconine,ben- zoylhypaconine were observed in different temperature and time,the degree of hydrolysis of ester- type alkaloids before and after decocting was observed,and the hydrolysate was determined. Results Eight peaks were identified in the HPLC chromatogram of Radix Aconite Lateralis before and after decocting; the total content of three digester alkaloids was 1. 756 mg / g,and the total content of three kinds of monoester alkaloids was 0. 724 mg / g,after boiling 1 h,those content were 0. 026 mg / g and 1. 961 mg / g; ten peaks were identified in the HPLC chromatogram of hydrolysis test of reference product,the degradation rate of mesaconitine,hypaconitine,aconitine when hydrolyzing 30 min at 50 ℃ were 18. 72%,4. 85% and 28. 91% respectively,the degradation rate of them when hydrolyzing 1 h in 50 ℃ respectively,the degradation rate was 100. 00% for 30 min or more in 100 ℃; the content of benzoylmesaconine,benzoylhypaconine,benzoylaconine when hydrolyzing 1 h in 50 ℃ were the same basically,the degradation rate of them when hydrolyzing 1 h in 100 ℃ were 63. 31%,42. 36%,64. 15% respectively,the degradation rate was 100. 00% when hydrolyzing 2 h or more in 100 ℃; according to the degradation rate of the reference substance,the results were evaluated as follows benzoylhypaconine > benzoylmesaconine > benzoylaconine > hypaconine > mesaconine > aconine. Conclusion The diester- type alkaloids could be hydrolyzed at 50 ℃ and be hydrolyzable at 100 ℃ for 30 min,the monoester- type alkaloid is stable at 50 ℃ and could be hydrolyzed at 100 ℃ for 2 h; the decocting process was a dynamic process of dissolution and degradation,and the content of the digester- type alkaloid disappears after decocting for 1 h.
Objective To observe the content of alkaloid of Radix Aconite Lateralis and the raw of hydrolysis,and toxicity reducing mechanism during decocting process. Methods HPLC method was used to determine the content of seven components before and after decocting and compare the change of content; the boiling process was simulated,the degradation rate and stability of mesaconitine,hypaconitine,aconine,benzoylmesaconine,benzoylaconine,ben- zoylhypaconine were observed in different temperature and time,the degree of hydrolysis of ester- type alkaloids before and after decocting was observed,and the hydrolysate was determined. Results Eight peaks were identified in the HPLC chromatogram of Radix Aconite Lateralis before and after decocting; the total content of three digester alkaloids was 1. 756 mg / g,and the total content of three kinds of monoester alkaloids was 0. 724 mg / g,after boiling 1 h,those content were 0. 026 mg / g and 1. 961 mg / g; ten peaks were identified in the HPLC chromatogram of hydrolysis test of reference product,the degradation rate of mesaconitine,hypaconitine,aconitine when hydrolyzing 30 min at 50 ℃ were 18. 72%,4. 85% and 28. 91% respectively,the degradation rate of them when hydrolyzing 1 h in 50 ℃ respectively,the degradation rate was 100. 00% for 30 min or more in 100 ℃; the content of benzoylmesaconine,benzoylhypaconine,benzoylaconine when hydrolyzing 1 h in 50 ℃ were the same basically,the degradation rate of them when hydrolyzing 1 h in 100 ℃ were 63. 31%,42. 36%,64. 15% respectively,the degradation rate was 100. 00% when hydrolyzing 2 h or more in 100 ℃; according to the degradation rate of the reference substance,the results were evaluated as follows benzoylhypaconine > benzoylmesaconine > benzoylaconine > hypaconine > mesaconine > aconine. Conclusion The diester- type alkaloids could be hydrolyzed at 50 ℃ and be hydrolyzable at 100 ℃ for 30 min,the monoester- type alkaloid is stable at 50 ℃ and could be hydrolyzed at 100 ℃ for 2 h; the decocting process was a dynamic process of dissolution and degradation,and the content of the digester- type alkaloid disappears after decocting for 1 h.
引文
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[13]吴克红,唐力英,王祝举,等.附子的化学和生物活性研究进展[J].中国实验方剂学杂志,2014,20(2):212-220.
[14]匡海学.中药化学[M].北京:中国中医药出版社,2003:1106-1108.
[2]陈务华,潘宗奇,陈秀玲,等.浅议《伤寒论》中生附子应用及临床体会[J].新中医,2014,46(6):240-241.
[3]郭丽丽.《伤寒论》《金匾要略方论》中附子临床应用的规律研究[D].广州:广州中医药大学,2009.
[4]洪俐,游国均.附子炮制诸法利弊与临床应用[J].时珍国医国药,2007,18(11):2836.
[5]周林,李飞,任玉珍,等.附子中生物碱含量与在胆巴液中浸泡时间变化规律的研究[J].中国实验方剂学杂志,2014,20(10):44.
[6]黄志芳,唐小龙,罗恒,等.HPLC-Q-TOF-MS分析附子的化学成分及煎煮过程中的变化规律[J].中国实验方剂学杂志,2015,21(1):57-63.
[7]张意涵,杨昌林,黄志芳,等.附子煎煮过程中13种生物碱含量的动态变化规律研究[J].药物分析杂志,2015,35(1):16-23.
[8]陈平,陈益民,陈佳,等.液相色谱-质谱法研究附子生物碱的煎煮减毒机理[J].色谱,2013,31(11):1087-1092.
[9]周思思,马增春,梁乾德,等.基于UPLC/Q-TOF-MS分析附子煎煮过程中化学成分的变化[J].中西医结合学报,2012,10(8):894-900.
[10]林华,沈玉巧,邓广海,等.附子炮制品及其煎煮液6种生物碱的含量测定[J].今日药学,2014,24(10):704-707.
[11]孙婷婷,栾立标.HPLC分离测定四逆汤中3种双酯型生物碱及其6种水解产物[J].中国中药杂志,2006,31(19):1634.
[12]陈东安,易进海,黄志芳,等.附子煎煮过程中酯型生物碱含量的动态变化[J].中国实验方剂学杂志,2011,17(3):64-68.
[13]吴克红,唐力英,王祝举,等.附子的化学和生物活性研究进展[J].中国实验方剂学杂志,2014,20(2):212-220.
[14]匡海学.中药化学[M].北京:中国中医药出版社,2003:1106-1108.
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