多种现代仪器分析方法在益母草注射液质量标准提高研究中的应用
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摘要
近几年中药注射剂不良反应等问题事件,已成为公众的焦点和热点。为了扭转局面,建立中药注射液指纹图谱及完善中药注射液质量标准等各种科研任务接踵而来。
益母草注射液为国家药典委员会首批要求建立中药注射液指纹图项目及首批列入中药注射液标准提高行动计划项目品种。本研究以此为契机,建立了益母草注射液、中间体及药材的HPLC指纹图谱评价方法,并完善了益母草注射液质量标准。
首先,利用多种色谱分析方法对益母草注射液进行较为深入的研究(1)液相色谱和飞行时间质谱联用(HPLC-TOF)对益母草注射液主要成分进行精确分子量定性,较深入的从物质基础上研究规范工艺下益母草注射液所含成分。(2)筛选不同的毛细管柱,不同的缓冲盐,检测波长。建立一个信息量大的毛细管电泳色谱条件,为过渡到适用性强的液相色谱,奠定色谱基础。(3)利用薄层色谱比较不同展开条件下益母草注射液的展开状况。(4)液相二极管检测器联用(HPLC-DAD)建立控制pH的反相色谱法、双离子对色谱法及离子交换色谱法的色谱条件。(5)为了合理筛选色谱方法,建立色谱评价标准。以重复性、耐用性、分离度、信息量、可操作性与实验成本共六项指标来衡量。对各项指标按实验结果进行计分,量化评定。最终筛选出控制pH的反相色谱法作为益母草注射液指纹图谱的色谱条件。
再次,在上述现代仪器分析方法了解的物质基础之上,利用筛选出的指纹色谱条件完善整个益母草注射液指纹图谱标准。(1)对临床上长期使用具有代表性的样品提取共有峰。(2)调查原生产工艺,对工艺参数进行优化。(3)合成标准指纹图谱和建立益母草注射液随行对照模型。(4)建立中间体及药材的标准指纹图谱方法。
最终,通过对益母草注射液现行标准的分析,增加有关物质检查,指纹图谱方法及盐酸水苏碱液相含量测定方法,完善整个益母草注射液的质量标准,基本保证该注射液质量的稳定性,为下一步成分与功效研究奠定基础。
In recent years reports about Adverse Reaction (ADR) caused by Traditional Chinese Medicine Injection (TCMI) were gradually increased and TCMI ADR has come to be the focus of society. To change this kind of situation, various scientific researches and tasks, such as establishing fingerprint chromatography and improving quality standards of TCMI, were carried out one after another.
Establishing fingerprint of Leonurus injection and enhancing the quality standards was one of the first action plan that was approved by China Pharmacopoeia Committee. The research established appraisal methods of HPLC fingerprint of Leonurus injection, intermediate and medicinal materials and perfected the quality standards.
Firstly, many kinds of methods of chromatography were used to analyze Leonurus injection. (1)The combination of HPLC with Time-of-Flight mass spectrometer technology(HPLC-TOF) was used to measure molecular weight accurately for analyzing the main components of Leonurus injection and researched deeply on main components in the injection produced by standard technology.(2) A chromatographic condition of capillary zone electrophoresis was established after screening different capillary column, buffer salt and determined wavelength. Much information was acquired and good foundation was set up for the purpose of building a much stronger applicable liquid Chromatogram. (3) Thin layer chromatography with different condition was carried out to analyze Leonurus injection. (4) The analytical condition of reversed phase chromatography, binary ion pair chromatography and ion - exchange chromatography were built by the combination of HPLC with Photodiode Array Detector (HPLC-DAD) technology. (5) Establishing the criterion for evaluation different chromatographic methods. The criterion had six-assessment indexes including repeatability, durability, resolution, information, operability and experiment cost. According to these indexes, the results of experiments were estimated quantitatively and the optimal fingerprint chromatography of Leonurus injection was selected. The best one was reversed phase chromatography.
Secondly, using the selected fingerprint chromatography improved the whole quality standards of Leonurus injection. (1) Extracted common chromatographic peaks from clinical long-term applicative and representative samples. (2) Surveyed former production technology and optimized technological parameter. (3) Composed the standard fingerprint and builted standard reference material of Leonurus injection. (4) Established standard fingerprint of intermediate and medicinal materials.
Thirdly, added 3 items after analyzing the current quality standards of Leonurus injection. The added items were related substance test, fingerprint chromatography and quantitative determination of stachydrine hydrochloride by HPLC method. The quality standard of Leonurus injection was improved and the stability of this injection had guaranteed. Good foundation was set up for researching component in this injection and efficacy of it in the future.
益母草注射液为国家药典委员会首批要求建立中药注射液指纹图项目及首批列入中药注射液标准提高行动计划项目品种。本研究以此为契机,建立了益母草注射液、中间体及药材的HPLC指纹图谱评价方法,并完善了益母草注射液质量标准。
首先,利用多种色谱分析方法对益母草注射液进行较为深入的研究(1)液相色谱和飞行时间质谱联用(HPLC-TOF)对益母草注射液主要成分进行精确分子量定性,较深入的从物质基础上研究规范工艺下益母草注射液所含成分。(2)筛选不同的毛细管柱,不同的缓冲盐,检测波长。建立一个信息量大的毛细管电泳色谱条件,为过渡到适用性强的液相色谱,奠定色谱基础。(3)利用薄层色谱比较不同展开条件下益母草注射液的展开状况。(4)液相二极管检测器联用(HPLC-DAD)建立控制pH的反相色谱法、双离子对色谱法及离子交换色谱法的色谱条件。(5)为了合理筛选色谱方法,建立色谱评价标准。以重复性、耐用性、分离度、信息量、可操作性与实验成本共六项指标来衡量。对各项指标按实验结果进行计分,量化评定。最终筛选出控制pH的反相色谱法作为益母草注射液指纹图谱的色谱条件。
再次,在上述现代仪器分析方法了解的物质基础之上,利用筛选出的指纹色谱条件完善整个益母草注射液指纹图谱标准。(1)对临床上长期使用具有代表性的样品提取共有峰。(2)调查原生产工艺,对工艺参数进行优化。(3)合成标准指纹图谱和建立益母草注射液随行对照模型。(4)建立中间体及药材的标准指纹图谱方法。
最终,通过对益母草注射液现行标准的分析,增加有关物质检查,指纹图谱方法及盐酸水苏碱液相含量测定方法,完善整个益母草注射液的质量标准,基本保证该注射液质量的稳定性,为下一步成分与功效研究奠定基础。
In recent years reports about Adverse Reaction (ADR) caused by Traditional Chinese Medicine Injection (TCMI) were gradually increased and TCMI ADR has come to be the focus of society. To change this kind of situation, various scientific researches and tasks, such as establishing fingerprint chromatography and improving quality standards of TCMI, were carried out one after another.
Establishing fingerprint of Leonurus injection and enhancing the quality standards was one of the first action plan that was approved by China Pharmacopoeia Committee. The research established appraisal methods of HPLC fingerprint of Leonurus injection, intermediate and medicinal materials and perfected the quality standards.
Firstly, many kinds of methods of chromatography were used to analyze Leonurus injection. (1)The combination of HPLC with Time-of-Flight mass spectrometer technology(HPLC-TOF) was used to measure molecular weight accurately for analyzing the main components of Leonurus injection and researched deeply on main components in the injection produced by standard technology.(2) A chromatographic condition of capillary zone electrophoresis was established after screening different capillary column, buffer salt and determined wavelength. Much information was acquired and good foundation was set up for the purpose of building a much stronger applicable liquid Chromatogram. (3) Thin layer chromatography with different condition was carried out to analyze Leonurus injection. (4) The analytical condition of reversed phase chromatography, binary ion pair chromatography and ion - exchange chromatography were built by the combination of HPLC with Photodiode Array Detector (HPLC-DAD) technology. (5) Establishing the criterion for evaluation different chromatographic methods. The criterion had six-assessment indexes including repeatability, durability, resolution, information, operability and experiment cost. According to these indexes, the results of experiments were estimated quantitatively and the optimal fingerprint chromatography of Leonurus injection was selected. The best one was reversed phase chromatography.
Secondly, using the selected fingerprint chromatography improved the whole quality standards of Leonurus injection. (1) Extracted common chromatographic peaks from clinical long-term applicative and representative samples. (2) Surveyed former production technology and optimized technological parameter. (3) Composed the standard fingerprint and builted standard reference material of Leonurus injection. (4) Established standard fingerprint of intermediate and medicinal materials.
Thirdly, added 3 items after analyzing the current quality standards of Leonurus injection. The added items were related substance test, fingerprint chromatography and quantitative determination of stachydrine hydrochloride by HPLC method. The quality standard of Leonurus injection was improved and the stability of this injection had guaranteed. Good foundation was set up for researching component in this injection and efficacy of it in the future.
引文
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10 从悦,王金辉等.益母草化学成分的分离与鉴定Ⅱ.中国药物化学杂志,2002,13(6):349
11 姜华等.益母草注射液对大鼠肠系膜淋巴微循环的影响 四川中医 2004.09.15;22(9):7-9
12 毕儒刚.益母草注射液的制备及其降血粘作用的试验研究.中国医院药学杂志 1989,9(4):172-173
13 郑鸿翱等.益母草注射液治疗兔心肌缺血再灌注损伤的机制探讨 中国现代医学杂志1997.07.15;7(7):19-22
14 陈少如等.益母草治疗心肌缺血或再灌注损伤及其机制研究 微循环学杂志 2001.12.15;11(4):16-19
15 陈少如等.益母草注射液对冠心病患者及大鼠心肌缺血再灌注心律失常的治疗作用 临床心血管病杂志 2002.10.25;18(10):490-491
16 陈穗等.益母草注射液对大鼠心肌缺血再灌注时心律失常的保护作用 汕头大学医学院学报 1999.09.15;12(3):9-10
17 尹俊等.益母草对心肌缺血大鼠血液流变学及血栓形成的影响,血栓与止血学.2001.02.15;7(1):13-15
18 姜华等.益母草注射液对失血性休克大鼠淋巴微循环的作用 陕西中医 2004.08.05;25(8):759-760
19 宋晶萍.董树彪.益母草颗粒中盐酸水苏碱的HPLC分析.广西医学,2002,24(8):1289
20 王文彤等.不同溶剂提取对益母草中生物碱含量的影响.天津药学,2003,15(3):5
21 姜舜尧.益母草药材中水苏碱成分的高效液相色谱法分析.药物分析杂志,2001,21(4):243.
22 罗淑荣,麦路,朱兆仪.益母草中生物碱的分离及含量测定.中药通报,1985,10(1):32.
23 唐盈,黄晓强等.四种含益母草中成药中水苏碱的含量测定.药物分析杂志,1989,9(1):22.
24 姜舜尧,田颂九,陈碧莲.几种中药生物碱的毛细管区带电泳分析.现代药物分析论坛.世界医药出版社,1999.128.
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2 医学科学院药物研究所等编.中药志,第二册,北京:人民卫生出版社,1993:481
3 江苏新医学院编.中药大辞典,下册,上海科学技术出版社,1991:1832
4 中国科学院四川分院中医中药研究所.四川中药志,第二册,四川人民出版社,1960:1441
5 阮金兰,杜俊蓉,曾庆忠.益母草的化学、药理和临床研究进展[J],中药学,2003,34(11),15-19.
6 张娴,彭国平.益母草化学成分研究.天然药物研究与开发,2004,16(2):104
7 晁志,周秀佳.类中药的研究概况和进展.中草药,1998,29(6):414
8 阮金兰,杜俊蓉,曾庆忠等.益母草的,药理和临床研究进展.中草药,2003,34(11):15
9 王金辉,从悦等.益母草化学成分的分离与鉴定.中国药物化学杂志,2002,12(3):146
10 从悦,王金辉等.益母草化学成分的分离与鉴定Ⅱ.中国药物化学杂志,2002,13(6):349
11 姜华等.益母草注射液对大鼠肠系膜淋巴微循环的影响 四川中医 2004.09.15;22(9):7-9
12 毕儒刚.益母草注射液的制备及其降血粘作用的试验研究.中国医院药学杂志 1989,9(4):172-173
13 郑鸿翱等.益母草注射液治疗兔心肌缺血再灌注损伤的机制探讨 中国现代医学杂志1997.07.15;7(7):19-22
14 陈少如等.益母草治疗心肌缺血或再灌注损伤及其机制研究 微循环学杂志 2001.12.15;11(4):16-19
15 陈少如等.益母草注射液对冠心病患者及大鼠心肌缺血再灌注心律失常的治疗作用 临床心血管病杂志 2002.10.25;18(10):490-491
16 陈穗等.益母草注射液对大鼠心肌缺血再灌注时心律失常的保护作用 汕头大学医学院学报 1999.09.15;12(3):9-10
17 尹俊等.益母草对心肌缺血大鼠血液流变学及血栓形成的影响,血栓与止血学.2001.02.15;7(1):13-15
18 姜华等.益母草注射液对失血性休克大鼠淋巴微循环的作用 陕西中医 2004.08.05;25(8):759-760
19 宋晶萍.董树彪.益母草颗粒中盐酸水苏碱的HPLC分析.广西医学,2002,24(8):1289
20 王文彤等.不同溶剂提取对益母草中生物碱含量的影响.天津药学,2003,15(3):5
21 姜舜尧.益母草药材中水苏碱成分的高效液相色谱法分析.药物分析杂志,2001,21(4):243.
22 罗淑荣,麦路,朱兆仪.益母草中生物碱的分离及含量测定.中药通报,1985,10(1):32.
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