HPLC法同时测定不同采集地衢枳壳中12种黄酮类成分的含量

Simultaneous Determination of the Contents of 12 Flavonoids in Quzhiqiao from Different Collection Places by HPLC

目的:建立同时测定衢枳壳中12种黄酮类成分含量的方法。方法:采用高效液相色谱法。色谱柱为Agilent Extend C18,流动相为0.1%甲酸水溶液-乙腈溶液(梯度洗脱),流速为1.0 mL/min,柱温为35℃,检测波长为330 nm,进样量为10μL。测定不同采集地的10批衢枳壳样品中12种黄酮类成分(圣草次苷、芸香柚皮苷、柚皮苷、柚皮素、橙皮苷、新橙皮苷、水合橙皮内酯、木犀草素、橙皮内酯、川陈皮素、桔皮素、橙皮油内酯)的含量。结果:圣草次苷、芸香柚皮苷、柚皮苷、柚皮素、橙皮苷、新橙皮苷、水合橙皮内酯、木犀草素、橙皮内酯、川陈皮素、桔皮素、橙皮油内酯检测的质量浓度线性范围分别为1.65~16.51、4.50~45.02、35.41~354.12、4.11~41.12、2.29~22.86、34.96~349.56、1.42~14.15、1.50~15.04、1.83~18.28、1.51~15.08、1.61~16.12、1.28~12.84μg/mL(r均大于0.999 7);检测限分别为0.165 1、0.450 2、3.541 2、0.411 2、0.228 6、3.495 6、0.141 5、0.150 4、0.182 8、0.150 8、0.161 2、0.128 4μg/mL;定量限分别为0.547 8、1.487 4、11.663 3、1.360 3、0.758 3、11.594 9、0.466 3、0.497 1、0.601 2、0.499 9、0.532 3、0.424 6μg/mL;精密度(n=6)、重复性(n=6)、稳定性(24 h,n=7)试验的RSD均小于3%;平均加样回收率分别为99.50%、99.61%、98.18%、98.85%、98.48%、98.50%、98.25%、99.91%、103.13%、98.82%、98.44%、100.29%(RSD=1.49%~2.38%,n=6)。10批不同采集地衢枳壳样品中,上述12个成分的含量分别为1.995 5~2.648 8、4.317 7~5.005 1、33.215 5~34.054 6、3.140 4~3.471 5、3.221 2~3.748 8、42.746 6~44.026 6、0.202 7~0.239 4、0.191 2~0.208 8、0.080 3~0.097 9、0.291 9~0.307 1、0.119 9~0.149 1、0.082 7~0.089 8 mg/g。结论:建立的含量测定方法准确、可靠、简便、高效,可用于同时测定衢枳壳中12个黄酮类成分的含量,并为衢枳壳质量控制标准的建立提供了参考依据。

OBJECTIVE:To establish a method for the simultaneous determination of the contents of 12 flavonoids in Quzhiqiao. METHODS:HPLC method was adopted. The determination was performed on Agilent Extend C18 column with mobile phase consisted of 0.1% formic acid-acetonitrile(gradient elution)at the flow rate of 1.0 mL/min. The column temperature was set at 35 ℃. The detection wavelength was set at 330 nm,and sample size was 10 μL. The contents of 12 components(such as eriocitrin,narirutin,naringin,naringenin,hesperidin,neohesperidin,hesperide hydrate,luteolin,hesperide,nobiletin,hesperetin and hesperidolactone)in 10 batches of Quzhiqiao from different collection places were determined. RESULTS:The linear range of eriocitrin,narirutin,naringin,naringenin,hesperidin,neohesperidin,hesperide hydrate,luteolin,hesperide,nobiletin,hesperetin and hesperidolactone were 1.65-16.51,4.50-45.02,35.41-354.12,4.11-41.12,2.29-22.86,34.96-349.56,1.42-14.15,1.50-15.04,1.83-18.28,1.51-15.08,1.61-16.12,1.28-12.84 μg/mL,respectively(all r>0.999 7). The detection limits were 0.165 1,0.450 2,3.541 2,0.411 2,0.228 6,3.495 6,0.141 5,0.150 4,0.182 8,0.150 8,0.161 2,0.128 4 μg/mL,respectively. The limits of quantitation were 0.547 8,1.487 4,11.663 3,1.360 3,0.758 3,11.594 9,0.466 3,0.497 1,0.601 2,0.499 9,0.532 3,0.424 6μg/mL,respectively. RSDs of precision(n=6),reproducibility(n=6)and stability(24 h,n=7)tests were all lower than 3%.The average recoveries were 99.50%,99.61%,98.18%,98.85%,98.48%,98.50%,98.25%,99.91%,103.13%,98.82%,98.44%, 100.29%(RSD=1.49%-2.38%, n=6). The contents of the above 12 components in 10 batches of samples from different collection places were 1.995 5-2.648 8,4.317 7-5.005 1,33.215 5-34.054 6,3.140 4-3.471 5,3.221 2-3.748 8,42.746 6-44.026 6,0.202 7-0.239 4,0.191 2-0.208 8,0.080 3-0.097 9,0.291 9-0.307 1,0.119 9-0.149 1,0.082 7-0.089 8 mg/g. CONCLUSIONS:The method is accurate,reliable,simple and efficient,which can be used to simultaneous determination of the contents of 12 flavonoids in Quzhiqiao,and to provide reference for the establishment of quality control standards of Quzhiqiao.