荞麦类黄酮成分的含量测定与分析研究

Content determination and analysis of flavonoids in buckwheat

目的建立高效液相色谱法同时测定荞麦中芦丁、金丝桃苷、山奈酚-3-O-芸香糖苷、槲皮素、芹菜素和山奈酚6种黄酮成分的含量,并比较分析不同产地荞麦中黄酮类成分差异情况。方法荞麦样品加入20 mL体积分数为80%甲醇水溶液超声辅助提取。高速离心后,经Shiseido CAPCELL PAK C18 MGⅡS5色谱柱(4.6mm×250mm,5.0μm),以乙腈-0.1%磷酸水溶液为流动相,梯度洗脱;流速为1.2mL/min;柱温为30.0±5℃,检测波长为361 nm。采用统计学方法分析不同产地荞麦的黄酮类成分差异。结果各黄酮类化合物在浓度为0.5～50.0mg/L的范围内线性关系良好(r>0.999),平均加标回收率为73.20%～114.90%,相对标准偏差(relative standard deviations,RSD)为2.02%～6.81%;精密度、稳定性、重复性实验的RSD值均小于5.35%,6种黄酮化合物的检出限为0.07～0.14 mg/L,定量限为0.23～0.45 mg/L。荞麦实际样品分析结果表明不同产地的荞麦中6种黄酮成分含量和总黄酮含量差异显著(P<0.05)。结论该方法分析快速简便、稳定和重复性好,可用于荞麦中芦丁、金丝桃苷、山奈酚-3-O-芸香糖苷、槲皮素、芹菜素和山奈酚6种黄酮成分含量的测定。为荞麦中黄酮的开发利用和含量的测定分析提供参考依据。

Objective To develop a method for simultaneous determination of rutin, hyperoside, kaempferol-3-O-rutinosid, quercetin, apigenin and kaempferol in buckwheat by high performance liquid chromatography, and compare the difference of flavone in buckwheat from different origins. Methods The buckwheat sample was added with 20 mL volume fraction of 80% methanol solution for ultrasonic-assisted extraction. After high speed centrifugation, the samples were separated with a Shiseido CAPCELL PAK C_（18） MGⅡS5 column（4.6 mm×250 mm, 5.0 μm） at the flow rate of 1.2 mL/min by gradient elution using acetonitrile and 0.1% phosphoric solution. The temperature of the column oven was maintained at 30.0±5 ℃ and the detection wavelength was set at 361 nm. The difference of flavonoids in buckwheat from different producing areas was analyzed by statistical method. Results Each compound of flavonoid showed a good linear relationship in the range of 0.5-50.0 mg/L（r〉0.999）, the average recovery rates was 73.20%-114.90% with the relative standard deviations（RSDs） of 2.02%-6.81%. The RSD values of accuracy, stability and repeatability were all less than 5.35%. The limits of detection of 6 flavone compounds were 0.07-0.14 mg/L, and the limits of quantitation were 0.23-0.45 mg/L. The results of sample analysis of buckwheat showed that there were significant differences in the content of 6 kinds of flavonoids and total flavonoids in buckwheat from different producing areas（P〈0.05）. Conclusion The method is rapid, simple, stable and repeatable, and can be used to determine the content of 6 flavonoids in buckwheat, including rutin, hypericin, sanitaphen-3-O-rutin, quercetin, celery and kaempferol, and can provide references for the development and utilization of flavonoids in buckwheat and the determination of flavonoids content.