RP-HPLC法测定杜仲增免液中松脂醇二葡萄糖苷的含量

摘要

The assay was aimed to determine the content of pinoresinol diglucoside (PDG) in Eucommie ulmoides immunoenhancement liquids.The chromatographic conditions were as follows: Inersustain(R)C18 column (250 mm×4.6 mm,5 μm) was under column temperature of 35 ℃, the mobile phase consisted of acetonitrile-0.2% phosphoric acid(15∶85)with a flow rate of 1.0 mL/min,and the UV detection wavelength was 277 nm.Ethyl acetate was used as extraction solvent.In order to determine PDG of the test solution under the chromatographic conditions, the number of theoretical plates and resolution were used as system suitability indicators.Linear regression on reference substance (PDG),linearity range and the precision, stability, and reproducibility of the analysis method, the recovery test of adding samples were all determined.The results showed that under the content determination method, the number of theoretical plates of PDG in the test solution was 8 588, and the resolution was 3.046.PDG performed good linear relation at the linear range between 6 and 192 μg/mL, and the related coefficient was 0.9996.The precision experiments showed that the relative standard deviation (RSD) of reference substance solution was 0.74%.The RSD of reproducibility and stability of PDG in the test solution was 4.50% and 2.69%, respectively.The average recovery was 98.74% with RSD 0.65% (n=9).Under the chromatographic conditions established above, the contents of PDG in the test sample were between 0.124 and 0.127 mg/mL.The conclusion was that the RP-HPLC method performed well system suitability, precision, reoroducibility, stability, and high recovery rate.Meanwhile this method was quick, simple and reliable.It could be used to determine the content of PDG in Eucommie ulmoides immunoenhancement liquids.%试验旨在建立测定杜仲增免液中松脂醇二葡萄糖苷含量的RP-HPLC法.以Inersustain(R)18(250 mm×4.6 mm,5 μm)为色谱柱,乙腈-0.2%磷酸(15∶85)为流动相,流速1.0 mL/min,柱温35 ℃,紫外检测波长277 nm.以乙酸乙酯作为提取溶剂,在所建立的色谱条件下测定供试品溶液,以理论塔板数和分离度作为系统适用性指标.对对照品松脂醇二葡萄糖苷溶液进行线性回归,确定线性范围,考察分析方法的精密度、稳定性、重现性,并进行加样回收试验.结果显示,用所建立的方法测定供试品溶液中松酯醇二葡萄糖苷理论塔板数为8 588,分离度为3.046.松脂醇二葡萄糖苷对照品溶液在6~192 μg/mL浓度范围内线性关系良好,相关系数为0.9996;精密度试验结果显示松脂醇二葡萄糖苷对照品溶液峰面积相对标准偏差(relative standard deviation,RSD)为0.74%;重现性和稳定性试验结果显示供试品溶液中松酯醇二葡萄糖苷峰面积RSD分别为4.50%和2.69%;平均加样回收率为98.74%,RSD为0.65%(n=9);在所建立的色谱条件下,供试品中松脂醇二葡萄糖苷的平均含量为0.124~0.127 mg/mL.结果表明,应用该方法测定杜仲增免液中松脂醇二葡萄糖苷的含量,系统适用性好,精密度高,重现性和稳定性好,回收率高,快速简便、准确可靠,可作为杜仲增免液中松脂醇二葡萄糖苷含量测定方法.