基于多波长切换HPLC同时测定五加皮中10个成分研究

Study on simultaneous determination of ten components in Acanthopanacis Cortex based on multi-wavelength switching HPLC

目的:建立多波长切换HPLC法同时测定五加皮多指标质量评价方法。方法:采用Waters AtlantisTMT3色谱柱(250 mm×4.6 mm,5μm),流动相为乙腈(A)-0.1%磷酸水溶液(B),梯度洗脱(0~15 min,10%A→20%A;15~25 min,20%A→40%A;25~30 min,40%A→85%A;30~45 min,85%A),流速1.0 mL·min^(-1),柱温35℃,检测波长为277 nm(0~30 min,检测新绿原酸、松柏苷、紫丁香苷、绿原酸、隐绿原酸、咖啡酸、1,3-二咖啡酰奎宁酸、异绿原酸A、异绿原酸C)、202 nm(30~45 min,检测异贝壳杉烯酸),进样量为20μL。结果:在优化的色谱条件下,新绿原酸、松柏苷、紫丁香苷、绿原酸、隐绿原酸、咖啡酸、1,3-二咖啡酰奎宁酸、异绿原酸A、异绿原酸C、异贝壳杉烯酸的质量浓度分别在1~100μg·mL^(-1)(r=1.000)、0.5~50μg·mL^(-1)(r=1.000)、0.25~25μg·mL^(-1)(r=1.000)、3.75~375μg·mL^(-1)(r=0.9999)、0.25~25μg·mL^(-1)(r=0.9999)、0.25~25μg·mL^(-1)(r=0.9999)、0.25~25μg·mL^(-1)(r=1.000)、2~200μg·mL^(-1)(r=0.9998)、0.5~50μg·mL^(-1)(r=0.9999)、14.45~1445μg·mL^(-1)(r=0.9998)范围内均呈现良好的线性关系,精密度、重复性和稳定性的RSD均<3%,加样回收率(n=6)为90.4%~104.0%,RSD为0.4%~2.4%。11批样品中上述成分含量范围分别为0.03~0.19,0.06~0.87、0.02~0.48、0.16~4.51、0.04~0.20、0.04~0.13、0.02~0.45、0.04~3.03、0.22~1.03、5.26~21.18 mg·g-1。结论:所建立的五加皮多指标质量评价方法快速、准确,有较好的重现性和稳定性,可用于五加皮的质量评价研究。

Objective:To establish a multi-index quality evaluation method for Acanthopanacis Cortex based on multi-wavelength switching HPLC.Methods:A Waters Atlantis~(TM)T_3(250 mm×4.6 mm,5μm)column was used in the HPLC assay.The mobile phase was acetonitrile(A)-0.1%phosphoric acid aqueous solution(B)by gradient elution(0-5 min,10%A→20%A,15-25 min,20%A→40%A,25-30 min,40%A→85%A,30-45 min,85%A).The flow rate was 1.0 mL·min^(-1),the column temperature was maintained at 35℃and the detection wavelengths were set at 270 nm(0-30 min)for neochlorogenic acid,abinoside,syringin,chlorogenic acid,cryptochlorogenic acid,caffeic acid,1,3-dicaffeioylquinic acid,isochlorogenic acid A,isochlorogenic acid C and 202 nm(30-45 min)for kaurenic acid.The injection volume was 20μL.Results:Under the above conditions,neochlorogenic acid,abinoside,syringin,chlorogenic acid,cryptochlorogenic acid,caffeic acid,1,3-dicaffeioylquinic acid,isochlorogenic acid A,isochlorogenic acid C and kaurenic acid showed good linearity in the ranges of 1-100μg·mL^(-1)(r=1.000),0.5-50μg·mL^(-1)(r=1.000),0.25-25μg·mL^(-1)(r=1.000),3.75-375μg·mL^(-1)(r=0.9999),0.25-25μg·mL^(-1)(r=0.9999),0.25-25μg·mL^(-1)(r=0.9999),0.25-25μg·mL^(-1)(r=1.000),2-200μg·mL^(-1)(r=0.9998),0.5-50μg·mL^(-1)(r=0.9999),14.45-1445μg·mL^(-1)(r=0.9998).The RSDs of precision,repeatability and stability were all less than 3.0%.The average recoveries(n=6)were 90.4%-104.0%with the RSD value of 0.4%-2.4%.The content ranges of the above-mentioned components in 11 samples were 0.03-0.19,0.06-0.87,0.02-0.48,0.16-4.51,0.04-0.20,0.04-0.13,0.02-0.45,0.04-3.03,0.22-1.03 and 5.26-21.18 mg·g~(-1),respectively.Conclusion:The multi-index quality evaluation method of Acanthopanacis Cortex is simple,accurate,reproducible and reliable,which can be used to evaluate the quality of it.