基于多指标成分含量测定及HPLC指纹图谱的多茎重楼品质评价
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摘要
目的 HPLC法测定不同产地多茎重楼Paris polyphylla Smith var. yunnanensis根茎中重楼皂苷Ⅶ、H、Ⅵ、Ⅱ、Ⅲ、Ⅰ、Ⅴ的含量,建立其HPLC指纹图谱,并与滇重楼根茎对照药材HPLC指纹图谱进行比较分析。方法采用Thermore C18色谱柱(250 mm×4.6 mm,5μm);流动相为水-乙腈,梯度洗脱,检测波长203 nm;柱温30℃;体积流量1 mL/min;进样量10μL,外标法与一测多评法同时对不同产地多茎重楼根茎中重楼皂苷Ⅶ、H、Ⅵ、Ⅱ、Ⅲ、Ⅰ、Ⅴ的含量进行测定,采用单因素方差分析法对含量差异进行分析;建立多茎重楼根茎HPLC指纹图谱,并与滇重楼根茎对照药材HPLC指纹图谱进行比较分析。结果不同产地多茎重楼根茎重楼皂苷Ⅶ、H、Ⅵ、Ⅱ、Ⅲ、Ⅰ、Ⅴ的含量测定结果表明,在线性范围内线性关系良好(r>0.9997),平均加样回收率为98.34%～99.34%,RSD≤1.00%;不同产地多茎重楼根茎中重楼皂苷Ⅶ、H、Ⅵ、Ⅱ、Ⅲ、Ⅰ、Ⅴ的含量存在显著(P<0.05)或极显著(P<0.01)差异,重楼皂苷(Ⅰ+Ⅱ+Ⅵ+Ⅶ)总量在1.239%～6.236%,显著高于《中国药典》规定的重楼药材质量控制标准的0.60%,且外标法与一测多评法测定结果无显著性差异。HPLC指纹图谱相似度评价结果表明,不同产地多茎重楼和滇重楼对照药材根茎中共有12个共有指纹特征峰,二者HPLC指纹图谱相似度较高。结论该方法操作简单、准确、重复性好,可用于多茎重楼根茎中重楼皂苷Ⅶ、H、Ⅵ、Ⅱ、Ⅲ、Ⅰ、Ⅴ的含量测定;不同产地多茎重楼根茎中重楼皂苷(Ⅰ+Ⅱ+Ⅵ+Ⅶ)总含量较高,不同产地多茎重楼与滇重楼对照药材根茎HPLC指纹图谱相似度较高。
Objective HPLC was used to determine the content of the polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ in the rhizomes of multiple stems Paris polyphylla var. yunnanenesis from different places of origin, establishing HPLC fingerprint and comparative analysis HPLC fingerprint between multiple stems P. polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis. Methods HPLC was performed on the column of Thermore C18 with mobile phase of acetonitrile-water gradient at the flow rate of 1 mL/min;The detection wave-length was 203 nm, column temperature was 30 ℃, and volume was 10 μL; Content of multiple stems P.polyphylla var. yunnanenesis. was measured by external standard and quantitative analysis of multi-components(QAMS), and one-way ANOVA was used to explore its content difference. The HPLC fingerprint of rhizome from multiple stems P. polyphylla var.yunnanenesis was established and compared with that of rhizome from P. polyphylla var. yunnanensis. Results The determination results of 10 batches of rhizome of multiple stems P. polyphylla var. yunnanenesIs showed a good linear relationship in the linear range(r > 0.999 7), with an average recovery of 98.34%—99.34% and RSD ≦ 1.00%. There was a significant difference(P < 0.05)or highly significant difference(P < 0.01) about content of polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ among different places of origin in the rhizomes of multiple stems P. polyphylla var. yunnanenesis, the total content of polyphyllins(Ⅰ+Ⅱ+Ⅵ+Ⅶ)% ranging from1.239%—6.236%, which was significantly higher than 0.60% of the quality control standard of Paridis Rhizoma prescribed by Chinese Pharmacopoeia. No significant difference was observed between the results of external standard and quantitative analysis of multi-components methods. The HPLC fingerprint similarity evaluation results showed that there were 12 common fingerprint peaks between multiple stems P. polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis, and with the high similarity of HPLC fingerprints. Conclusion The method was simple, accurate and reproducible, and can be used for the determination of polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ in multiple stems of P. polyphylla var. yunnanenesis, total content of polyphyllins(I + II + VI + VII)% in multiple stems of P. polyphylla var. yunnanenesis was relatively higher and HPLC fingerprints of rhizome of multiple stems P.polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis from different places of origin also have high similarity.
Objective HPLC was used to determine the content of the polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ in the rhizomes of multiple stems Paris polyphylla var. yunnanenesis from different places of origin, establishing HPLC fingerprint and comparative analysis HPLC fingerprint between multiple stems P. polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis. Methods HPLC was performed on the column of Thermore C18 with mobile phase of acetonitrile-water gradient at the flow rate of 1 mL/min;The detection wave-length was 203 nm, column temperature was 30 ℃, and volume was 10 μL; Content of multiple stems P.polyphylla var. yunnanenesis. was measured by external standard and quantitative analysis of multi-components(QAMS), and one-way ANOVA was used to explore its content difference. The HPLC fingerprint of rhizome from multiple stems P. polyphylla var.yunnanenesis was established and compared with that of rhizome from P. polyphylla var. yunnanensis. Results The determination results of 10 batches of rhizome of multiple stems P. polyphylla var. yunnanenesIs showed a good linear relationship in the linear range(r > 0.999 7), with an average recovery of 98.34%—99.34% and RSD ≦ 1.00%. There was a significant difference(P < 0.05)or highly significant difference(P < 0.01) about content of polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ among different places of origin in the rhizomes of multiple stems P. polyphylla var. yunnanenesis, the total content of polyphyllins(Ⅰ+Ⅱ+Ⅵ+Ⅶ)% ranging from1.239%—6.236%, which was significantly higher than 0.60% of the quality control standard of Paridis Rhizoma prescribed by Chinese Pharmacopoeia. No significant difference was observed between the results of external standard and quantitative analysis of multi-components methods. The HPLC fingerprint similarity evaluation results showed that there were 12 common fingerprint peaks between multiple stems P. polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis, and with the high similarity of HPLC fingerprints. Conclusion The method was simple, accurate and reproducible, and can be used for the determination of polyphyllins Ⅶ,H,Ⅵ,Ⅱ,Ⅲ,Ⅰ and Ⅴ in multiple stems of P. polyphylla var. yunnanenesis, total content of polyphyllins(I + II + VI + VII)% in multiple stems of P. polyphylla var. yunnanenesis was relatively higher and HPLC fingerprints of rhizome of multiple stems P.polyphylla var. yunnanenesis and P. polyphylla var. yunnanensis from different places of origin also have high similarity.
引文
[1]中国药典[S].一部.2015.
[2]Qin X J,Ni W,Chen C X,et al.Seeing the light:shifting from wild Rhizomes to extraction of active ingredients from above-ground parts of Paris polyphylla var.yunnanensis[J].J Ethnopharm,2018,224:134-139.
[3]Guo Y,Liu z,Li K,et al.Paris polyphylla-derived saponins inhibit growth of bladder cancer cells by inducing mutant P53 degradation while up-regulating CDKN1A expression[J].Curr Urol,2018,11(3):131-138.
[4]杨远贵,张霁,张金渝,等.重楼属植物化学成分及药理活性研究进展[J].中草药,2016,47(18):3301-3322.
[5]李海涛,罗先文,管燕红,等.云南省不同地区滇重楼皂苷含量的对比及影响因子分析[J].中国中药杂志,2014,39(5):803-806.
[6]李恒,苏豹,张兆云,等.中国重楼资源现状评价及其种植业的发展对策[J].西部林业科学,2015,44(3):1-7.
[7]中国科学院昆明植物研究所.云南植物志(第8卷)[M].北京:科学出版社,1997.
[8]中国科学院中国植物志编辑委员会.中国植物志.第十五卷[M].北京:科学出版社,1978.
[9]王彩步,李晨,杨敏,等.多茎滇重楼不同部分甾体皂苷活性成分累积的研究[J].辽宁中医药杂志,2018,45(3):582-585.
[10]张绍山,刘璇,王景富,等.UPLC法测定云南省不同地区云南重楼及多芽品系中7种甾体皂苷量及其指纹图谱建立[J].中草药,2016,47(23):4257-4263.
[11]邹亮,周浓,张海珠,等.HPLC测定不同产地滇重楼中的4种重楼皂苷[J].华西药学志,2009,24(5):521-523.
[12]昝珂,高宇明,郑健,等.UPLC法同时测定云南重楼栽培品中11种皂苷的含量[J].药物分析杂志,2017,37(9):1572-1577.
[13]李懿,何佳,赵庭周,等.HPLC同时测定不同产地滇重楼中的6种重楼皂苷[J].中成药,2012,34(1):113-116.
[14]黄圆圆,刘大会,彭华胜,等.15种重楼属植物中8种甾体皂苷的含量测定[J].中国中药杂志,2017,14(17):3443-3451.
[15]秦昆明,杨冰,胡静,等.一测多评法在中药多组分质量控制中的应用现状与思考[J].中草药,2018,49(3):725-731.
[16]田刚,李超,吴菲,等.基于一测多评法对小儿豉翘清热颗粒中9个成分的质量控制[J].药物分析,2018,38(11):1922-1930.
[2]Qin X J,Ni W,Chen C X,et al.Seeing the light:shifting from wild Rhizomes to extraction of active ingredients from above-ground parts of Paris polyphylla var.yunnanensis[J].J Ethnopharm,2018,224:134-139.
[3]Guo Y,Liu z,Li K,et al.Paris polyphylla-derived saponins inhibit growth of bladder cancer cells by inducing mutant P53 degradation while up-regulating CDKN1A expression[J].Curr Urol,2018,11(3):131-138.
[4]杨远贵,张霁,张金渝,等.重楼属植物化学成分及药理活性研究进展[J].中草药,2016,47(18):3301-3322.
[5]李海涛,罗先文,管燕红,等.云南省不同地区滇重楼皂苷含量的对比及影响因子分析[J].中国中药杂志,2014,39(5):803-806.
[6]李恒,苏豹,张兆云,等.中国重楼资源现状评价及其种植业的发展对策[J].西部林业科学,2015,44(3):1-7.
[7]中国科学院昆明植物研究所.云南植物志(第8卷)[M].北京:科学出版社,1997.
[8]中国科学院中国植物志编辑委员会.中国植物志.第十五卷[M].北京:科学出版社,1978.
[9]王彩步,李晨,杨敏,等.多茎滇重楼不同部分甾体皂苷活性成分累积的研究[J].辽宁中医药杂志,2018,45(3):582-585.
[10]张绍山,刘璇,王景富,等.UPLC法测定云南省不同地区云南重楼及多芽品系中7种甾体皂苷量及其指纹图谱建立[J].中草药,2016,47(23):4257-4263.
[11]邹亮,周浓,张海珠,等.HPLC测定不同产地滇重楼中的4种重楼皂苷[J].华西药学志,2009,24(5):521-523.
[12]昝珂,高宇明,郑健,等.UPLC法同时测定云南重楼栽培品中11种皂苷的含量[J].药物分析杂志,2017,37(9):1572-1577.
[13]李懿,何佳,赵庭周,等.HPLC同时测定不同产地滇重楼中的6种重楼皂苷[J].中成药,2012,34(1):113-116.
[14]黄圆圆,刘大会,彭华胜,等.15种重楼属植物中8种甾体皂苷的含量测定[J].中国中药杂志,2017,14(17):3443-3451.
[15]秦昆明,杨冰,胡静,等.一测多评法在中药多组分质量控制中的应用现状与思考[J].中草药,2018,49(3):725-731.
[16]田刚,李超,吴菲,等.基于一测多评法对小儿豉翘清热颗粒中9个成分的质量控制[J].药物分析,2018,38(11):1922-1930.