目的:采用高效液相色谱(HPLC)技术,建立同时测定复方双花片中新绿原酸、绿原酸、隐绿原酸、连翘酯苷A、3,4-二-O-咖啡酰奎宁酸、3,5-二-O-咖啡酰奎宁酸、4,5-二-O-咖啡酰奎宁酸、连翘苷、穿心莲内酯和脱水穿心莲内酯10个成分含量的方法。方法:复方双花片样品用50%甲醇水超声提取,采用Agilent Zorbax SB-C18(250 mm×4.6 mm,5μm)色谱柱分析,流动相为乙腈-0.15%磷酸水溶液,梯度洗脱,流速1.0 mL·min-1,柱温30 ℃,进样量10 μL,检测波长327 nm(检测新绿原酸、绿原酸、隐绿原酸、连翘酯苷A、3,4-二-O-咖啡酰奎宁酸、3,5-二-O-咖啡酰奎宁酸、4,5-二-O-咖啡酰奎宁酸)和226 nm(检测连翘苷、穿心莲内酯、脱水穿心莲内酯)。结果:新绿原酸、绿原酸、隐绿原酸、连翘酯苷A、3,4-二-O-咖啡酰奎宁酸、3,5-二-O-咖啡酰奎宁酸、4,5-二-O-咖啡酰奎宁酸、连翘苷、穿心莲内酯和脱水穿心莲内酯质量浓度分别在4.071~40.71 μg·mL-1(r=0.999 9)、20.16~201.6 μg·mL-1(r=0.999 9)、4.730~47.30 μg·mL-1(r=0.999 9)、4.536~45.36 μg·mL-1(r=0.999 8)、1.817~18.17 μg·mL-1(r=0.999 9)、2.266~22.66 μg·mL-1(r=0.999 7)、3.321~33.21 μg·mL-1(r=0.999 9)、3.462~34.62 μg·mL-1(r=0.999 6)、2.111~21.11 μg·mL-1(r=0.999 9)和2.290~22.90 μg·mL-1(r=0.999 7)与峰面积呈良好的线性关系;平均回收率(RSD)(n=6)分别为101.3%(2.1%)、103.0%(1.5%)、100.9%(2.0%)、101.1%(2.0%)、98.4%(1.6%)、102.2%(2.4%)、98.2%(1.3%)、97.8%(2.0%)、99.0%(2.0%)和96.4%(1.1%);4批样品中上述10个成分的含量分别为1.685~2.649 mg·g-1、12.202~13.780 mg·g-1、2.415~2.594 mg·g-1、1.340~1.919 mg·g-1、0.501~0.791 mg·g-1、0.891~1.342 mg·g-1、1.299~2.105 mg·g-1、1.147~1.504 mg·g-1、0.654~0.694 mg·g-1、0.846~1.151 mg·g-1。结论:所建立的方法简单准确,可用于复方双花片的质量控制与评价。
Objective: To establish an HPLC method for simultaneous determination of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, forsythiaside A, 3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, forsythin, andrographalide and dehydroandrographalide in compound Shuanghua tablets. Methods: The samples were extracted with 50% methanol solution under ultrasonic condition, and were performed on Agilent Zoabax SB-C18 column (250 mm×4.6 mm, 5 μm) by gradient elution of acetonitrile -0.15% phosphoric acid solution at a flow rate of 1.0 mL·min-1. The column temperature was 30 ℃, the injection volume was 10 μL, and the detection wavelength were set at 327 nm (detecting neochlorogenic acid,chlorogenic acid, cryptochlorogenic acid, forsythiaside A,3,4-O-dicaffeoylquinic acid,3,5-O-dicaffeoylquinic acid and 4,5-O-dicaffeoylquinic acid) and 226 nm (detecting forsythin, andrographalide and dehydroandrographalide). Results: The linear ranges of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid,forsythiaside A,3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, forsythin, andrographalide and dehydroandrographalide were 4.071-40.71 μg·mL-1(r=0.999 9), 20.16-201.6 μg·mL-1(r=0.999 9), 4.730-47.30 μg·mL-1(r=0.999 9), 4.536-45.36 μg·mL-1(r=0.999 8), 1.817-18.17 μg·mL-1(r=0.999 9), 2.266-22.66 μg·mL-1(r=0.999 7), 3.321-33.21 μg·mL-1(r=0.999 9), 3.462-34.62 μg·mL-1(r=0.999 6), 2.111-21.11 μg·mL-1(r=0.999 9) and 2.290-22.90 μg·mL-1(r=0.999 7), respectively. The average recoveries (n=6) were 101.3%(2.1%),103.0%(1.5%),100.9%(2.0%),101.1%(2.0%),98.4%(1.6%),102.2%(2.4%),98.2%(1.3%),97.8%(2.0%),99.0%(2.0%)and 96.4%(1.1%), respectively. The contents of the 10 components in 4 bacthes of Fufang Shuanghua tablets were in the range of 1.685-2.649 mg·g-1, 12.202-13.780 mg·g-1, 2.415-2.594 mg·g-1, 1.340-1.919 mg·g-1, 0.501-0.791 mg·g-1, 0.891-1.342 mg·g-1, 1.299-2.105 mg·g-1, 1.147-1.504 mg·g-1, 0.654-0.694 mg·g-1, 0.846-1.151 mg·g-1, respectively. Conclusion: This simple accurate reproducible method can be used for the quality control and evaluate of compound Shuanghua tablets.
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