目的: 采用一测多评(QAMS)法测定维血宁合剂中8个有效成分的含量,建立“中间体-成品”的HPLC指纹图谱,为其质量控制提供参考和依据。方法: 采用高效液相色谱法,使用Acclaim120 C18(250 mm×4.6 mm,5 μm)色谱柱,以甲醇(A)-0.2%磷酸水溶液(B)-0.1%磷酸(C)为流动相,进行梯度洗脱,流速为1.0 mL·min-1,检测波长为 210 nm,柱温为 30 ℃。以虎杖苷为内参物,分别考察儿茶素、表儿茶素、白藜芦醇、槲皮素、木犀草素、染料木素和芒柄花素7个成分的相对校正因子。采用相对校正因子计算3批维血宁合剂提取液、清膏和成品中7 个成分的含量和指纹图谱,与外标法(ESM)进行比较,验证QAMS法的准确性和可行性。结果: 儿茶素、表儿茶素、虎杖苷、白藜芦醇、槲皮素、木犀草素、染料木素和芒柄花素的线性范围分别为1.066~53.300、1.062~53.100、4.384~219.200、1.118~55.900、1.036~51.800、1.110~55.400、1.040~52.000、0.530~53.000 μg·mL-1。以虎杖苷为参照物,其他7个成分的相对校正因子分别为0.375 4、0.348 6、0.585 3、0.597 2、0.517 6、0.588 7、0.659 9,在不同的实验条件下重复性较好。9个批次提取液、清膏、成品中8个成分的含量范围:虎杖苷为45.26~90.45 μg·mL-1,儿茶素为21.29~38.18 μg·mL-1,表儿茶素为15.06~21.46 μg·mL-1,白藜芦醇为6.26~8.57 μg·mL-1,槲皮素为1.22~2.13 μg·mL-1,木犀草素为3.35~4.44 μg·mL-1,染料木素为1.97~3.17 μg·mL-1,芒柄花素为0.65~1.13 μg·mL-1。3批维血宁合剂的指纹图谱有27个共有峰,并且指认了以上8个成分峰,绝大部分相似度在0.9以上。结论: 以虎杖苷为内参物建立的QAMS法操作简单,省时高效,可为维血宁合剂的质量控制提供参考。
Objective: To establish a quantitative analysis of multi-components by single marker(QAMS) method for the determination of 8 isoflavones in Weixuening mixture, and establish the HPLC fingerprint of “intermediate finished product”, so as to provide reference and basis for its quality control. Methods: The Acclaim 120-C18 column (250 mm×4.6 mm,5 μm) was used with methanol (A)-0.2% phosphoric acid aqueous solution(B)-0.1% phosphoric acid(C) as mobile phase. Gradient elution was performed at a flow rate of 1.0 mL·min-1, with the detection wavelength of 210 nm and the column temperature of 30 ℃. Using polydatin as the reference substance, relative correction factors of catechin, epicatechin, resveratrol, quercetin, luteolin, genistein and formononetin were calculated, respectively. The relative correction factors were used to calculate the contents and fingerprints of 7 substances to be tested in 3 batches of Weixuening mixture, and the differences between the two methods were compared to verify the accuracy and feasibility. Results: The linear ranges of catechin, epicatechin, polydatin, resveratrol, quercetin, luteolin, genistein and formononetin were 1.066-53.300, 1.062-53.100, 4.384-219.200, 1.118-55.900, 1.036-51.800, 1.110-55.400, 1.040-52.000, 0.530-53.000 μg·mL-1, respectively. The relative correction factors of catechin, epicatechin, resveratrol, quercetin, luteolin, genistein and formononetin were 0.375 4, 0.348 6, 0.585 3, 0.597 2, 0.517 6,0.588 7, 0.659 9, respectively. The repeatability was good under different experimental conditions. At the same time, the content range of 8 components in the extracts, clear extracts and mixture was polydatin (45.26-90.45 μg·mL-1), catechin (21.29-38.18 μg·mL-1), epicatechin (15.06-21.46 μg·mL-1), resveratrol (6.26-8.57 μg·mL-1), quercetin (1.22-2.13 μg·mL-1), luteolin (3.35-4.44 μg·mL-1), genistein (1.97-3.17 μg·mL-1), and anthocyanin (0.65-1.13 μg·mL-1). There were 27 common peaks in the fingerprint of three batches of Weixuening mixture, and 8 peaks of polydatin, quercetin, catechin, epicatechin, luteolin, anthocyanin and resveratrol were identified, most of which were more than 0.9 similar. Conclusion: The established system is simple, time-saving and efficient, which is conducive to provide reference for the quality control of Weixuening mixture.
[1] 中华人民共和国药典2020年版. 一部[S].2020: 1703
ChP 2020. VolⅠ[S].2020: 1703
[2] 黄伟, 罗雅琴, 王晓.维血宁颗粒治疗免疫性血小板减少症的网络药理作用机制研究[J].中国药物警戒, 2021, 18(11):1048
HUANG W, LUO YQ, WANG X. Network pharmacological mechanisms of Weixuening granules in the treatment of immune thrombocytopenia[J].Chin J Pharmacov, 2021, 18 (11): 1048
[3] 曹林林, 田正良, 宋宗良, 等. 维血宁合剂治疗抗甲状腺药物导致白细胞减少症临床观察[J].时珍国医国药, 2011, 22(8): 1971
CAO LL, TIAN ZL, SONG ZL, et al. Clinical observation of Weixuening mixture in the treatment of leukopenia caused by antithyroid drugs[J].Lishizhen Med Mater Med Res, 2011, 22 (8): 1971
[4] 俞曙星. 维血宁对小儿贫血58例的治疗效果[J].中国卫生标准管理, 2015, 6(28): 155
YU SX. Therapeutic effect of Weixuening on 58 cases of infantile anemia[J].Chin Health Stand Manag, 2015, 6(28): 155
[5] 潘迎英, 杨怡敏, 陶敏贤. 维血宁颗粒治疗化疗后血小板减少38例临床观察[J].长春中医药大学学报, 2010, 26(4): 541
PAN YY, YANG YM, TAO MX. Clinical observation on 38 cases of thrombocytopenia after chemotherapy treated with Weixuening granule[J].J Changchun Univ Tradit Chin Med, 2010, 26(4): 541
[6] 朱鸿敏, 金艺, 李晶, 等. RP-HPLC同时测定维血宁颗粒中芍药苷、虎杖苷、麦角甾苷和白藜芦醇的含量[J].药物分析杂志, 2009, 29(6): 905
ZHU HM, JIN Y, LI J, et al. RP-HPLC simultaneous determination of paeoniflorin, verbascoside, polydatin and resveratrol in Weixuening granules[J].Chin J Pharm Anal, 2009, 29(6):90
[7] 马双成, 王莹, 魏锋. 中药质量控制未来发展方向的思考[J].中国药学杂志, 2021, 56(16):1273
MA SC, WANG Y, WEI F. Ideas on future development direction of quality control of traditional Chinese medicine[J].Chin Pharm J, 2021, 56(16): 1273
[8] 阳长明, 陈霞, 赵巍, 等. 基于源头控制的中药制剂质量研究[J].中草药, 2021, 52(2): 321
YANG CM, CHEN X, ZHAO W, et al. Research on quality of traditional Chinese medicine preparations based on control of source[J].Chin Tradit Herb Drugs, 2021, 52(2): 321
[9] 伍振峰, 郑琴, 杨明, 等. 中药制剂质量控制的方法模式分析与研究[J].中国中药杂志, 2012, 37(9): 1332
WU ZF, ZHENG Q, YANG M, et al. Analysis and study on quality control methods and modes of traditional Chinese medicine preparations[J].Chin Pharm J, 2012, 37(9): 1332
[10] 王智民, 钱忠直, 张启伟, 等. 一测多评法建立的技术指南[J].中国中药杂志, 2011, 36(6): 657
WANG ZM, QIAN ZZ, ZHANG QW, et al. A technical guide established by the one-test-multiple-assessment method[J].Chin Pharm J, 2011, 36(6): 657
[11] 董媛, 李海亮, 王楠, 等.一测多评法测定三七总皂苷中5个皂苷的含量[J].药物分析杂志, 2022, 42(3): 518
DONG Y, LI HL, WANG N, et al. Determination of five saponins in extract of notoginseng total saponinsby quantitative analysis of multi-components by single marker[J].Chin J Pharm Anal, 2022, 42(3): 518
[12] 胡瑞雪, 梁元昊, 徐文丽, 等. 一测多评法在中药中的应用及研究进展[J].药物分析杂志, 2019, 39(11): 1968
HU RX, LIANG YH, XU WL, et al. Application and research progress of QAMS in traditional Chinese medicine[J].Chin J Pharm Anal, 2019, 39(11): 1968
[13] 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
QIN KM, YANG B, HU J, et al. Application status and thinking of quantitative analysis of multi-components by single marker in quality control of multi-components traditional Chinese medicine[J].Chin Tradit Herb Drugs, 2018, 49(3): 725
[14] 龙凯花, 刘峰, 张红, 等. 一测多评法同时测定冠心舒通胶囊中10个成分的含量[J].药学学报, 2022, 57(6): 1880
LONG KH, LIU F, ZHANG H, et al. Simultaneous determination of 10 components in Guanxinshutong capsules by quantitative analysis of multi-components by single marker[J].Acta Pharm Sin, 2022, 57(6): 1880
[15] 赵冬青. 中药复方制剂的质量控制方法探讨[J].中医药管理杂志, 2022, 30(3): 200
ZHAO DQ. Discussion on quality control methods of traditional Chinese medicine compound preparations[J].J Tradit Chin Med Manag, 2022, 30(3): 200
[16] 吕玥, 汤湧, 李立言, 等. 中药复方制剂质量控制体系的研究进展[J].中国中西医结合外科杂志, 2020, 26(1): 170
LÜ Y, TANG Y, LI LY, et al. Research progress on quality control system of traditional Chinese medicine compound preparations[J].Chin J Integr Tradit Chin West Med, 2020, 26(1): 170
[17] 董泰玮, 邹俊波, 张小飞, 等. 基于膜分离法与醇沉法维血宁合剂工艺研究对比[J].中国新药杂志, 2021, 30(20): 1890
DONG TW, ZOU JB, ZHANG XF, et al. Comparison of membrane separation method and alcohol precipitation method as purification process of Weixuening mixture[J].Chin New Drugs J, 2021, 30(20): 1890
