OBJECTIVE:To establish quantitative analysis of HPLC-quantitative analysis of multi-components by single marker (QAMS) for simultaneous determination of 6 active constituents in Compound huangqin tablets. METHODS:Using baicalin as internal content,HPLC method was used to calculate relative correction factor (RCF) of baicalin to polydatin,wogonoside, baicalein,andrographolide and dehydroandrographolide. The contents of above-mentioned 5 components in Compound huangqin tablets were calculated by using RCF(calculation value). The external standard method(ESM)was used to determine the contents of 6 constituents (measured value). The differences between calculation value and measured value were compared. The determination was performed on ZORBAX Eclipse XDB C18 column with mobile phase consisted of acetonitrile-0.05％ phosphoric acid (gradient elution) at the flow rate of 1.0 mL/min. The detection wavelength was set at 306 nm for polydatin,225 nm for andrographolide,275 nm for baicalin,wogonoside and baicalein,254 nm for dehydroandrographolide. The column temperature was set at 25 ℃ and the injection volume was 10 μL. RESULTS:The linear range was 4.555-81.99 μg/mL for polydatin, 47.03-846.5 μg/mL for baicalin,12.73-229.1 μg/mL for wogonoside,12.88-231.8 μg/mL for baicalein,2.632-47.38 μg/mL for andrographolide,3.508-63.14 μg/mL for dehydroandrographolide (r=0.9992-0.9997),respectively. Limits of detection were 2.25,6.92,1.76,4.69,1.37,1.51 ng;limits of quantitation were 6.89,19.98,5.28,13.81,4.15,4.71 ng. Average recoveries were 98.2％,98.8％,97.9％,98.9％,98.8％,98.7％(RSD=0.91％-1.32％,n=9). RSDs of precision,reproducibility and stability tests (48 h) were lower than 2.0％ (n=6 or n=7). RCFs of baicalin to polydatin, wogonoside, baicalein, andrographolide and dehydroandrographolide were 0.529,0.506,1.004,0.831 and 1.087,respectively. There was no statistical significance between calculated values and measured ones(P>0.05). CONCLUSIONS:The method is simple,effective,accurate and cost saving. It can be used for simultaneous determination of 6 active constituents in Compound huangqin tablets.%目的:建立同时测定复方黄芩片中6种活性成分含量的高效液相色谱(HPLC)-一测多评法(QAMS).方法:采用HPLC法,以黄芩苷为内参物,分别计算其与虎杖苷、汉黄芩苷、黄芩素、穿心莲内酯、脱水穿心莲内酯的相对校正因子(RCF),通过RCF计算复方黄芩片中上述5种成分的含量(计算值),同时采用外标法测定这6种成分的含量(实测值),比较计算值与实测值的差异.色谱柱为ZORBAX Eclipse XDB C18,流动相为乙腈-0.05％磷酸溶液(梯度洗脱),流速为1.0 mL/min,检测波长为306 nm(虎杖苷)、225 nm(穿心莲内酯)、275 nm(黄芩苷、汉黄芩苷、黄芩素)、254 nm(脱水穿心莲内酯),柱温为25℃,进样量为10μL.结果:虎杖苷、黄芩苷、汉黄芩苷、黄芩素、穿心莲内酯、脱水穿心莲内酯的检测质量浓度的线性范围分别为4.555～81.99、47.03～846.5、12.73～229.1、12.88～231.8、2.632～47.38、3.508～63.14μg/mL(r为0.9992～0.9997);检测限分别为2.25、6.92、1.76、4.69、1.37、1.51 ng,定量限分别为6.89、19.98、5.28、13.81、4.15、4.71 ng;平均加样回收率分别为98.2％、98.8％、97.9％、98.9％、98.8％、98.7％(RSD为0.91％～1.32％,n=9);精密度、重复性、稳定性(48 h)试验的RSD<2.0％(n=6或n=7).虎杖苷、汉黄芩苷、黄芩素、穿心莲内酯、脱水穿心莲内酯的RCF分别为0.529、0.506、1.004、0.831、1.087,其计算值和实测值比较差异无统计学意义(P>0.05).结论:该方法简单、有效、结果准确、节约成本,可用于复方黄芩片中上述6种活性成分含量的同时测定.
展开▼
