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目的建立HPLC梯度洗脱法同时测定天麻首乌片(TSP)中12种活性成分芍药苷、2,3,5,4′-四羟基二苯乙烯-2-O-β-D-葡萄糖苷(二苯乙烯苷)、天麻素、阿魏酸、毛蕊花糖苷、甘草苷、丹参酮IIA、特女贞苷、欧前胡素、芦丁、蟛蜞菊内酯和大黄素的量。方法采用HPLC法,Hypersil GOLD C18(250 mm×4.6 mm,5μm)色谱柱;乙腈-0.1%磷酸水溶液为流动相,体积流量1.0 m L/min,梯度洗脱;进样量为10μL。结果 12种活性成分芍药苷、二苯乙烯苷、天麻素、阿魏酸、毛蕊花糖苷、甘草苷、丹参酮IIA、特女贞苷、欧前胡素、芦丁、蟛蜞菊内酯和大黄素分别在3.12~31.20μg/m L(r=0.999 5)、4.98~49.80μg/m L(r=0.999 4)、1.05~10.50μg/m L(r=0.999 5)、0.99~9.90μg/m L(r=0.999 2)、1.11~11.10μg/m L(r=0.999 2)、3.24~32.40μg/m L(r=0.999 2)、3.63~36.30μg/m L(r=0.999 2)、1.13~11.30μg/m L(r=0.999 3)、1.9~19.0μg/m L(r=0.999 5)、1.55~15.50μg/m L(r=0.999 5)、1.48~14.80μg/m L(r=0.999 5)、102.8~1028.0μg/m L(r=0.999 9)质量浓度与峰面积具有较好的线性关系;精密度、重复性良好,RSD均小于2.0%;平均加样回收率和相应的RSD分别为99.88%、1.55%,101.25%、0.98%,99.67%、1.29%,102.04%、1.17%,101.17%、1.67%,98.27%、1.51%,100.28%、1.20%,99.11%、0.95%,98.49%、1.67%,101.57%、0.94%,102.37%、0.58%,97.89%、0.69%。12批次供试品中12种指标成分质量分数分别为1.241~1.261、2.116~2.133、0.540~0.558、0.077~0.099、0.089~0.110、1.111~1.134、0.158~0.183、1.375~1.399、0.342~0.372、0.542~0.571、0.648~0.672、45.05~45.93 mg/g。结论建立的HPLC梯度洗脱法可同时测定TSP中12种活性成分,该方法操作简便、快速、准确,可作为TSP全面可靠的质量控制方法。
Objective To establish a HPLC gradient elution method for the simultaneous determination of 12 active constituents paeoniflorin, 2,3,5,4’-tetrahydroxystilbene-2-O-β-D-glucoside in Tianma Shouwu tablets (TSP) Stilbene glycoside), gastrodin, ferulic acid, verbascoside, glycyrrhizin, tanshinone IIA, estrogenin, imperatorin, rutin, weenolactone and emodin. Methods The HPLC method was used on a Hypersil GOLD C18 column (250 mm × 4.6 mm, 5 μm). The mobile phase was acetonitrile-0.1% phosphoric acid solution and the volume flow rate was 1.0 mL / min. The injection volume was 10 μL. Results The active components of paeoniflorin, stilbene glycoside, gastrodin, ferulic acid, verbascoside, glycyrrhizin, tanshinone IIA, estrogen, imperatorin, rutin, (R = 0.999 5), 0.99-9.90 μg / m L (r = 0.999 5), 4.98-49.80 μg / m L (r = 0.999 4), 1.05-10.50 μg / r = 0.999 2), 1.11-11.11 μg / m L (r = 0.999 2), 3.24-3240 μg / m L (r = 0.999 2), 3.63- 36.30 μg / m L (R = 0.999 5), 1.48 ~ 14.80 μg / m L (r = 0.999 5) 0.999 5). The mass concentration of 102.8-1028.0 μg / mL (r = 0.999 9) had a good linear relationship with the peak area. The precision and reproducibility were good with RSDs less than 2.0%. The average recoveries of samples and the corresponding RSD were 99.88%, 1.55%, 101.25%, 0.98%, 99.67%, 1.29%, 102.04%, 1.17%, 101.17%, 1.67%, 98.27%, 1.51%, 100.28%, 1.20%, 99.11%, 0.95% , 98.49%, 1.67%, 101.57%, 0.94%, 102.37%, 0.58%, 97.89%, 0.69%. 12 batches of the 12 indicators for the quality indicators were 1.241 ~ 1.261,2.116 ~ 2.133,0.540 ~ 0.558,0.077 ~ 0.099,0.089 ~ 0.110,1.111 ~ 1.134,0.158 ~ 0.183,1.375 ~ 1.399,0.342 ~ 0.372 , 0.542 ~ 0.571, 0.648 ~ 0.672, 45.05 ~ 45.93 mg / g. Conclusion The established HPLC gradient elution method can simultaneously determine 12 kinds of active ingredients in TSP. The method is simple, rapid and accurate and can be used as TSP comprehensive and reliable quality control method.