HPLC法测定山野豌豆与狭山野豌豆茎叶中5个黄酮苷的含量

目的：建立HPLC法测定山野豌豆与狭山野豌豆中金丝桃苷、异槲皮苷、山柰苷、槲皮苷及山柰酚-3-O-α-L-阿拉伯糖苷的含量，为山野豌豆与狭山野豌豆的质量控制提供依据。方法：采用Diamonsil C₁₈色谱柱（200 mm×4.6 mm，5 μm）；以乙腈-0.1%磷酸水溶液梯度洗脱，流速1.0 mL·min^-1；紫外检测波长350 nm；柱温25℃；进样量10 μL。结果：金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷的检测下限质量浓度分别为0.20、0.21、0.10、0.22、0.18 μg·mL^-1，线性范围分别为15.12~756.00（r=0.999 9）、14.92~746.00（r=0.999 9）、10.46~523.00（r=0.999 9）、28.84~1442.00（r=0.999 9）、3.20~160.00（r=0.999 9）μg·mL^-1，精密度试验RSD分别为0.21%、0.49%、0.31%、0.12%和0.11%。山野豌豆样品中金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷平均回收率分别为102.3%、99.2%、98.9%、98.3%和99.5%，重复性试验的RSD分别为1.4%、1.9%、1.3%、1.7%和1.2%，稳定性试验的RSD分别为0.85%、0.62%、0.91%、0.25%和0.26%；山野豌豆样品中金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷平均含量分别为0.166、0.080、0.762、0.600和0.192 mg·g^-1。狭山野豌豆样品中金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷的平均回收率分别为101.9%、98.6%、100.1%、98.1%和99.8%，重复性试验的RSD分别为1.6%、2.1%、1.6%、2.1%和1.7%，稳定性试验的RSD分别为0.71%、0.87%、0.82%、0.15%和0.23%；狭山野豌豆样品中金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷平均含量分别为0.735、0.919、0.526、1.312和0.173 mg·g^-1。结论：该方法可用于检测狭山野豌豆与山野豌豆中金丝桃苷、异槲皮苷、山柰苷、槲皮苷、山柰酚-3-O-α-L-阿拉伯糖苷的质量浓度。

Objective: To establish an HPLC method for determining the contents of hyperoside,isoquercitrin, kaempferitrin, quercitrin, kaempferol-3-O-α-L-arabinoside in Vicia amoena Fisch.and Vicia amoena Fisch.var.angusta Freyn., and so as to provide the evidence for quality guarantee for V.amoena var.angusta and V.amoena.Methods: Diamonsil C₁₈ column(200 mm×4.6 mm, 5 μm)was used.Acetonitrile and 0.1% phosphoric acid water solution was used as mobile phase, and the flow rate was at 1.0 mL·min^-1; the detection was at 350 nm;the column temperature was at 25℃, and the injection volume was 10 μL.Results: The lowest detection limits for hyperoside, isoquercitrin, kaempferitrin, quercitrin and kaempferol-3-O-α-L-arabinoside were 0.20 μg·mL^-1, 0.21 μg·mL^-1, 0.10 μg·mL^-1, 0.22 μg·mL^-1, 0.18 μg·mL^-1, respectively, the linear ranges were 15.12-756.00 μg·mL^-1, 14.92-746.00 μg·mL^-1, 10.46-523.00 μg·mL^-1, 28.84-1 442.00 μg·mL^-1 and 3.20-160.00 μg·mL^-1, respectively;the precision RSD values were 0.21%, 0.49%, 0.31%, 0.12% and 0.11%, respectively.The average recoveries of hyperoside, isoquercitrin, kaempferitrin, quercitrin and kaempferol-3-O-α-L-arabinoside in V.amoena were 102.3%, 99.2%,98.9%,98.3% and 99.5%,respectively;the repeatability RSD values were 1.4%,1.9%,1.3%,1.7% and 1.2%,respectively;the stability RSD values were 0.85%,0.62%,0.91%,0.25% 和 0.26%,respectively;the average contents of hyperoside,isoquercitrin,kaempferitrin,quercitrin and kaempferol-3-O-α-L-arabinoside inV.amoena.were 0.166 mg·g^-1,0.080 mg·g^-1,0.762 mg·g^-1,0.600 mg·g^-1 and 0.196 mg·g^-1,respectively.The average recoveries of hyperoside,isoquercitrin,kaempferitrin,quercitrin and kaempferol-3-O-α-L-arabinoside in V.amoena.var.angusta were 101.9%,98.6%,100.1%,98.1% and 99.8%,respectively;the repeatability RSD values were 1.55%, 2.13%, 1.63%, 2.13% and 1.69%, respectively.The stability RSD values were 0.71%, 0.87%, 0.82%,0.15% 和 0.23%,respectively;the average contents of hyperoside,isoquercitrin,kaempferitrin,quercitrin and kaempferol-3-O-α-L-arabinoside in V.amoena var.angusta.were 0.735 mg·g^-1, 0.919 mg·g^-1, 0.526 mg·g^-1,1.312 mg·g^-1 and 0.173 mg·g^-1,respectively.Conclusion: The method is available for determining the contents of hyperoside;isoquercitrin;kaempferitrin;quercitrin and kaempferol-3-O-α-L-arabinoside.

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