平衡透析法测定羊耳菊提取物中9个成分的血浆蛋白结合率
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摘要
为了测定民族药羊耳菊提取物中9个成分的血浆蛋白结合率。首先采用平衡透析法测定提取物中9个成分血浆蛋白结合率,生物样本用甲醇沉淀蛋白法进行处理,再以葛根素为内标,利用超高效液相色谱-质谱仪串联测定血浆及缓冲溶液中的绿原酸、新绿原酸、隐绿原酸、东莨菪苷、1,3-O-二咖啡酰基奎宁酸、3,4-O-二咖啡酰基奎宁酸、3,5-O-二咖啡酰基奎宁酸和4,5-O-二咖啡酰基奎宁酸等9个成分的浓度。结果发现在所研究浓度范围内,9个成分呈现良好的线性关系(r≥0.999),准确度、精密度、提取回收率和稳定性等均符合生物样品测定要求,且并无内源性杂质干扰,表明该文所建立的方法稳定可靠,可用于羊耳菊药材中9种成分在人和大鼠血浆中的血浆蛋白结合率测定。9种成分在人和大鼠血浆中的平均蛋白结合率分别在(41.07±0.046)%～(94.95±0.008)%,(37.66±0.043)%～(97.46±0.013)%。结果表明羊耳菊提取物中9个成分在人和大鼠中的血浆蛋白结合率存在差异。
To determine the plasma protein binding rate of the nine compounds in Inula cappa extraction by the method of equilibrium dialysis. The proteins in plasma samples were precipitated by methanol, and the ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was developed for determination of the concentrations of the nine active compounds, namely chlorogenic acid, scopolin, neochlorogenic acid, cryptochlorogenic acid, 1,3-O-dicaffeoylquinic acid, galuteolin, 3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, with the internal standard of puerarin. We found that all components have a good linearity(r≥0.999), and accuracy, precision, extraction recovery and stability conformed to the requirements of determination, without endogenous compounds disturbing within the range of optimum concentration. This suggested that the method was stable and reliable, and could be used for the determination of the plasma protein binding rates of the nine active compounds in rat and human plasma of I. cappa. The plasma protein binding rates of the nine active compounds in rat and human plasma respectively were(41.07±0.046)%-(94.95±0.008)%, and(37.66±0.043)%-(97.46±0.013)%. According to the results, there were differences in the plasma protein binding rates of the nine compounds in I. cappa extraction between rat and human.
To determine the plasma protein binding rate of the nine compounds in Inula cappa extraction by the method of equilibrium dialysis. The proteins in plasma samples were precipitated by methanol, and the ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was developed for determination of the concentrations of the nine active compounds, namely chlorogenic acid, scopolin, neochlorogenic acid, cryptochlorogenic acid, 1,3-O-dicaffeoylquinic acid, galuteolin, 3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, with the internal standard of puerarin. We found that all components have a good linearity(r≥0.999), and accuracy, precision, extraction recovery and stability conformed to the requirements of determination, without endogenous compounds disturbing within the range of optimum concentration. This suggested that the method was stable and reliable, and could be used for the determination of the plasma protein binding rates of the nine active compounds in rat and human plasma of I. cappa. The plasma protein binding rates of the nine active compounds in rat and human plasma respectively were(41.07±0.046)%-(94.95±0.008)%, and(37.66±0.043)%-(97.46±0.013)%. According to the results, there were differences in the plasma protein binding rates of the nine compounds in I. cappa extraction between rat and human.
引文
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[16] 巩仔鹏, 吴林霖, 伍萍, 等. 羊耳菊提取物在大鼠粪便中的代谢产物分析[J]. 中国实验方剂学杂志, 2017, 23(24): 100.
[17] 周玲,居文政,刘子修,等. 灯盏细辛注射液多成分血浆蛋白结合率测定[J]. 中国药理学通报,2011, 27(5):719.
[18] 冯志强,韩坚,谢智勇,等. 黄芩提取物中黄酮类成分血浆蛋白结合率的测定[J]. 中国药理学通报,2012, 28(2): 286.
[19] 黄勇,陈慧,郑林,等. 液质联用超滤法测定注射用复方荭草中原儿茶酸、异荭草素及野黄芩苷的血浆蛋白结合率[J]. 中国药学杂志,2011, 46(15): 1200.
[20] 孙佳慧, 赵海峰, 郭兴蕾, 等. 中药有效成分血浆蛋白结合率的研究进展与相关规律探讨[J]. 实用药物与临床, 2017, 20(1): 91.
[2] 国家中医药管理局中华本草编委会. 中华本草·苗药卷[M].贵阳:贵州科技出版社, 2005: 273.
[3] 贵州中医研究所.苗族医药学[M].贵阳:贵州民族出版社,1992:506.
[4] 贵州省药品监督管理局.贵州省中药材、民族药材质量标准[S].贵阳:贵州科技出版社,2003.
[5] 中国民族药志编委会. 中国民族药志[M]. 成都:四川民族出版社, 2007:257.
[6] Wang Y L, Li Y J, Wang A M, et al. Two new phenolic glycosides from Inula cappa[J]. J Asian Nat Prod Res, 2010, 12(9):765.
[7] 关焕玉,兰燕宇,廖尚高,等. 羊耳菊中咖啡酰基奎宁酸类化学成分研究[J].天然产物研究与开发,2014(26):1948.
[8] 周雯,王霞,付思红,等.羊耳菊的化学成分研究[J].中国药学杂志,2017,52(1):25.
[9] Amit Singh, Khima Nand Balodi, Suneet Naithani, et al. Vascular plant diversity with special reference to invasion of alien species on the Doon University Campus, Dehardun, India[J]. Int J Biodivers Conserv, 2017,9(3):56.
[10] Shikha Mohan, Damodar Gupta.Phytochemical analysis and differential in vitro cytotoxicity assessment of root extracts of Inula racemosa[J]. Biomed Pharmacother, 2017,5:781.
[11] Jiraporn N, Puttinan M, Yingmanee T. Inhibition of Inula cappa (Ham. ex D. Don) DC. extracts on herpes simplex virus infection in vitro[J]. Afr J Microbiol Res, 2011, 5(24): 4049.
[12] 巩仔鹏, 李梅, 侯靖宇, 等.外翻肠囊法研究羊耳菊提取物在大鼠肠内的吸收[J]. 中国中药杂志, 2018, 43(3): 609.
[13] 伍萍, 李梅, 巩仔鹏, 等. 基于在体循环肠灌流模型分析羊耳菊提取物的肠吸收特性[J]. 中国实验方剂学杂志, 2018, 24(2): 1.
[14] 巩仔鹏, 侯靖宇, 李梅, 等.大鼠肠道菌群对羊耳菊提取物的代谢作用研究[J]. 中国中药杂志, 2017, 42(18): 3584.
[15] 巩仔鹏, 陈亭亭, 侯靖宇, 等. 基于UHPLC/Q-TOF/MS分析羊耳菊有效组分的入血成分[J].中国药理学通报, 2017, 33(11): 1605.
[16] 巩仔鹏, 吴林霖, 伍萍, 等. 羊耳菊提取物在大鼠粪便中的代谢产物分析[J]. 中国实验方剂学杂志, 2017, 23(24): 100.
[17] 周玲,居文政,刘子修,等. 灯盏细辛注射液多成分血浆蛋白结合率测定[J]. 中国药理学通报,2011, 27(5):719.
[18] 冯志强,韩坚,谢智勇,等. 黄芩提取物中黄酮类成分血浆蛋白结合率的测定[J]. 中国药理学通报,2012, 28(2): 286.
[19] 黄勇,陈慧,郑林,等. 液质联用超滤法测定注射用复方荭草中原儿茶酸、异荭草素及野黄芩苷的血浆蛋白结合率[J]. 中国药学杂志,2011, 46(15): 1200.
[20] 孙佳慧, 赵海峰, 郭兴蕾, 等. 中药有效成分血浆蛋白结合率的研究进展与相关规律探讨[J]. 实用药物与临床, 2017, 20(1): 91.