HPLC-DAD法同时测定荆防藿朴解毒合剂(新冠3号)中14个成分的含量

Simultaneous determination of 14 components in Jingfang Huopu Jiedu mixture(Xinguan No.3)by HPLC-DAD

目的:建立HPLC法同时测定荆防藿朴解毒合剂中14个成分(新绿原酸、咖啡酸、绿原酸、苦杏仁苷、隐绿原酸、紫丁香苷、1,3-二咖啡酰奎尼酸、阿魏酸、升麻素苷、5-O-甲基维斯阿米醇苷、橙皮苷、欧前胡素、和厚朴酚和厚朴酚)的含量。方法:采用ZXRBAX SB-C18(4.6 mm×250 mm,5μm)色谱柱,以乙腈-甲醇-0.1%磷酸水溶液为流动相,进行梯度洗脱,流速为1.0 mL·min^-1,柱温40℃,检测波长分别为327 nm(新绿原酸、咖啡酸、绿原酸、隐绿原酸、1,3-二咖啡酰奎尼酸、阿魏酸)、207 nm(苦杏仁苷和橙皮苷)和254 nm(升麻素苷、5-O-甲基维斯阿米醇苷、紫丁香苷、欧前胡素、和厚朴酚、厚朴酚)。结果:荆防藿朴解毒合剂中上述14个化学成分均可实现较好的分离,质量浓度在一定的范围内与峰面积的线性关系良好,r>0.9990,方法学考察结果表明该仪器精密度、方法重复性、各样品溶液的稳定性均良好(RSD<2.5%,n=6),平均加样回收率为98.2%~101.5%(RSD<2.5%,n=6),3批样品中上述14个成分的含量范围依次为0.0172~0.0179、0.0278~0.0287、0.0884~0.0908、0.3829~0.3922、0.0243~0.0252、0.0172~0.0180、0.0613~0.0641、0.0283~0.0291、0.0281~0.0289、0.0132~0.0136、0.0455~0.0472、0.0014~0.0015、0.0082~0.0085和0.0127~0.0132 mg·mL^-1。结论:本实验建立的含量测定方法稳定可靠,可用于该复方中14个成分的同步含量测定,可为疫情期间该组方的临床使用安全及有效及质量可控提供方法学参考,也可为后续临床药理研究提供实验基础。

Objective:To establish an HPLC method for simultaneous determination of fourteen constituents(neochlorogenic acid,caffeic acid,chlorogenic acid,amygdalin,cryptochlorogenic acid,syringin,3-dicaffeoylquinic acid,ferulic acid,prim-O-glucosylcimifugin,5-O-methylvisammioside,hesperidin,imperatorin,honokiol and magnolol)in Jingfang Huopu Jiedu mixture.Methods:The analysis was performed on a ZXRBAX SB-C18(4.6 mm×250 mm,5μm)column with gradient elution of acetonitrile-methanol-0.1%phosphoric acid at a flow rate was 1.0 mL·min^-1.The column temperature was 40℃.The detection wavelengths were set at 327 nm(neochlorogenic acid,caffeic acid,chlorogenic acid,cryptochlorogenic acid,1,3-dicaffeoylquinic acid and ferulic acid),207 nm(amygdalin and hesperidin)and 254 nm(prim-O-glucosylcimifugin,5-O-methylvisammioside,syringin,imperatorin,honokiol and magnolol).Results:The excellent linearity between the concentration and peak area was obtained for the above 14 components in Jingfang Huopu Jiedu mixture in the determined ranges(r>0.9990)with the average recovery of 98.2%-101.5%(RSD<2.5%,n=6),and the results of methodological investigation showed that the precision of the instrument,the repeatability of the method and stability of each sample solution were all good(RSD<2.5%,n=6).The contents of the above 14 components in 3 batches of sample were 0.0172-0.0179,0.0278-0.0287,0.0884-0.0908,0.3829-0.3922,0.0243-0.0252,0.0172-0.0180,0.0613-0.0641,0.0283-0.0291,0.0281-0.0289,0.0132-0.0136,0.0455-0.0472,0.0014-0.0015,0.0082-0.0085 and 0.0127-0.0132 mg·mL^-1.Conclusion:The contents determination method established in this experiment is stable and reliable,which can be used to determine the content of 14 components in the compound simultaneously and provide methodological reference ensure for the compound’s safe,effective and quality-controlled clinical use during COVID-19 outbreaks,it can also provide experimental basis for subsequent clinical pharmacological studies.