目的:采用超高效液相色谱(UPLC)建立八珍益智方指纹图谱和多成分定量分析方法。方法:采用ACQUITY UPLC HSS T3色谱柱(100 mm×2.1 mm, 1.8 μm),流动相为乙腈(A)-0.2%甲酸水溶液(B),梯度洗脱,流速0.4 mL·min-1,检测波长254 nm(甘草酸)及300 nm(原儿茶酸、甘草苷、橙皮苷、异甘草苷、川陈皮素、桔皮素)。通过对照品比对与Q-TOF/MS技术对共有峰进行指认,采用UPLC-PAD进行含量测定。结果:测得10批次八珍益智方样品的指纹图谱与对照指纹图谱的相似度均大于0.936;共确定八珍益智方指纹图谱19个共有峰,指认14个化学成分,对其中原儿茶酸、甘草苷、橙皮苷、异甘草苷、川陈皮素、桔皮素、甘草酸7个成分进行含量测定。上述7个成分在一定浓度范围内呈良好线性关系,相关系数均大于0.999 9;方法学考察结果显示各项指标均符合要求,回收率范围为92.8%~106.7%,RSD小于4.2%。10批样品中原儿茶酸、甘草苷、橙皮苷、异甘草苷、川陈皮素、桔皮素、甘草酸含量范围分别为0.006~0.017、0.035~0.220、0.622~2.113、0.015~0.045、0.007~0.022、0.003~0.010、0.237~0.642 mg·g-1。结论:所建立的指纹图谱及多指标成分含量测定的分析方法稳定可行,专属性强,可用于八珍益智方的质量控制,为新药研发和临床安全用药提供依据。
Objective: To establish the fingerprint and multi-component determination method of Bazhen Yizhi prescription by using ultra-high performance liquid chromatography (UPLC). Methods: ACQUITY UPLC HSS T3(100 mm×2.1 mm, 1.8 μm) column was used, the mobile phase was acetonitrile (A)-0.2% formic acid aqueous solution (B) with gradient elution at a flow rate of 0.4 mL·min -1, the detection wavelengths were 254 nm (glycyrrhizic acid) and 300 nm (3,4-dihydroxybenzoic acid, liquiritin, hesperidin, isoliquiritin, nobiletin, tangeretin). The common peaks were identified by comparison with reference substances and by Q-TOF/MS technology, and UPLC-PAD was used for content determination. Results: The similarity of fingerprints of 10 batches of Bazhen Yizhi prescription samples to reference fingerprint were all above 0.936. A total of 19 common peaks in the fingerprint of Bazhen Yizhi prescription were assigned, and 14 components were identified. The contents of 3,4-dihydroxybenzoic acid, liquiritin, hesperidin, isoliquiritin, nobiletin, tangeretin and glycyrrhizic acid were determined. The 7 substances showed good linear relationship within certain concentration ranges. The correlation coefficients were all above 0.999 9. The methodological investigation results showed that all indicators meet the requirements, the recovery rates were between 92.8%-106.7%, and the RSDs were less than 4.23%. The content ranges of 3,4-dihydroxybenzoic acid, liquiritin, hesperidin, isoliquiritin, nobiletin, tangeretin and glycyrrhizic acid in 10 batches of samples were 0.006-0.017,0.035-0.220,0.622-2.113,0.015-0.045,0.007-0.022,0.003-0.010 and 0.237-0.642 mg·g-1, respectively. Conclusion: The fingerprint and the determination method of multi-index components are stable, feasible and specific, and can be used for the quality control of Bazhen Yizhi prescription, providing a basis for new drug development and clinical safe drug use.
[1] 牛阳, 张霞, 王荣, 等. 八珍益智颗粒提取工艺研究[J].西北药学杂志, 2012, 27(4): 294
NIU Y, ZHANG X, WANG R, et al. Study on the extraction method for Bazhenyizhi prescription[J].Northwest Pharm J, 2012, 27(4): 294
[2] 牛阳, 张霞, 王荣, 等. 八珍益智颗粒制备工艺优选[J].中国实验方剂学杂志, 2012, 18(7): 34
NIU Y, ZHANG X , WANG R, et al. Optimization of preparation process of Bazhen Yizhi prescription[J].Chin J Exp Tradit Med Form, 2012, 18(7): 34
[3] 牛阳, 周波, 王荣, 等. 八珍益智复方和制备方法及应用: CN110694025A[P]. 2020-01-17
NIU Y, ZHOU X , WANG R, et al. The Preparation Method and Application of Bazhen Yizhi Prescription: CN110694025A[P]. 2020-01-17
[4] BARRECA D, GATTUSO G, LAGANA G, et al. C-and O-glycosyl flavonoids in Sanguinello and Tarocco blood orange(Citrus sinensis (L.) Osbeck) juice: identification and influence on antioxidant properties and acetylcholinesterase activity[J].Food Chem, 2016, 196(APR. 1): 619
[5] WANG SF, CHEN P, JIANG W, et al. Identification of the effective constituents for anti-inflammatory activity of Ju-Zhi-Jiang-Tang, an ancient traditional Chinese medicine formula[J].J Chromatogr A, 2014, 1348: 105
[6] ZHENG YY, ZENG X, PENG W, et al. Characterisation and classification of Citri Reticulatae Pericarpium varieties based on UHPLC-Q-TOF-MS/MS combined with multivariate statistical analyses[J].Phytochem Anal, 2019, 30(3): 278
[7] 张珂, 许霞, 李婷, 等. 利用UHPLC-IT-TOF-MS分析陈皮的化学成分组[J].中国中药杂志, 2020, 45(4): 899
ZHANG K, XU X, LI T, et al. Chemome profiling of Citri Reticulatae Pericarpium using UHPLC-IT-TOF-MS[J].China J Chin Mater Med, 2020, 45(4): 899
[8] ROOWI S, CROZIER A. Flavonoids in tropical citrus species[J].J Agric Food Chem, 2011, 59(22): 12217
[9] DJOUKENG JD, ARBONA V, ARGSAMAILLA R, et al. Flavonoid profiling in leaves of citrus genotypes under different environmental situations[J].J Agric Food Chem, 2008, 56(23): 11087
[10] XING TT, ZHAO X J, ZHANG Y, et al. Fast separation and sensitive quantitation of polymethoxylated flavonoids in the peels of citrus using UPLC-Q-TOF-MS[J].J Agric Food Chem, 2017, 65(12): 2615
[11] 侯蕾, 王亚玲, 王文锦, 等. 益智仁化学成分研究[J].中草药, 2020, 51(2): 315
HOU L, WANG YL, WANG WJ, et al. Chemical constituents from Alpinia oxyphylla[J].Chin Herb Med, 2020, 51(2): 315
[12] 盛康美, 王宏洁. 龙眼肉的化学成分与药理作用研究进展[J].中国实验方剂学杂志, 2010, 16(5): 236
SHENG KM, WANG HJ. Advances in research of chemical constituents and pharmacological activites of Arillus Longan[J].Chin J Exp Tradit Med Form, 2010, 16(5): 236
[13] 董庆海, 吴福林, 王涵, 等. 山药的化学成分和药理作用及临床应用研究进展[J].特产研究, 2018, 40(4): 98
DONG QH, WU FL, WANG H, et al. Research progress on chemical constituents, pharmacological activites and clinical application of Chinese yam[J].Spec Wild Economic Animal Plant Res, 2018, 40(4): 98
[14] 赵小亮, 章军, 马小军, 等. 莲不同药用部位化学成分研究综述[J].中国中医药信息杂志, 2012, 19(1): 106
ZHAO XL, ZHANG J, MA XJ, et al. A review of research on chemical constituents of different medicinal parts of Nelumbinis Semen[J].Chin J Inf Tradit Chin Med, 2012, 19(1): 106
[15] 崔鹤蓉, 王睿林, 郭文博, 等.茯苓的化学成分、药理作用及临床应用研究进展[J].西北药学杂志, 2019, 34(5): 694
CUI HR, WANG RL, GUO WB, et al. Research advances in chemical components, pharmacological activites and clinical application of Poria cocos[J].Northwest Pharm J, 2019, 34(5): 694
[16] 顾思浩, 孔维崧, 张彤, 等. 白术的化学成分与药理作用及复方临床应用进展[J].中华中医药学刊, 2020, 38(1): 69
GU SH, KONG WS, ZHANG T, et al. advances on chemical compositions, pharmacological effects and compound clinical applications of Atractylodes macrocephala Koidz[J].Chin Arch Tradit Chin Med, 2020, 38(1): 69
[17] 李恩灿, 贺玖明, 靳洪涛, 等. 5-羟甲基糠醛的药理和毒理研究进展[J].中国药物警戒, 2018, 15(4): 210
LI EC, HE JM, JIN HT, et al. Research progress of pharmacological activity and toxicity of 5-hydroxymethylfurfural[J].Chin J Pharmacovigilance, 2018, 15(4): 210
[18] 赵玲, 周臣清, 朱婉清, 等. 5-羟甲基糠醛的生物安全性和生物活性研究进展[J].食品工业科技, 2016, 37(11): 372
ZHAO L, ZHOU CQ, ZHU WQ, et al. Progress in the biological safety and activity of 5-hydroxymethylfurfural[J].Sci Technol Food Ind, 2016, 37(11): 372
