HPLC法测定红豆杉细胞培养物中的α-萘乙酸

Determination of α-Naphthylacetic Acid In Cell Culture of Taxus by HPLC Method

建立高效液相色谱法检测红豆杉细胞培养物中萘乙酸残留的方法。细胞培养液直接用二氯甲烷萃取,鲜细胞用99%乙醇浸提后,再用二氯甲烷萃取,分别得到回收率均达到87%以上的萘乙酸试样,色谱分离采用Nova-pak C18柱,柱温30℃,紫外检测波长为281 nm,进样体积5μL,外标法定量。结果显示萘乙酸的质量在0.001～0.125 ug(R=0.9998)范围内与色谱峰面积呈良好的线性关系,萘乙酸在细胞培养液中的检出限为0.02 mg/L,在鲜细胞中的检出限为0.2 mg/kg。本方法简便快速、灵敏度高,可监测红豆杉细胞培养生产紫杉醇过程中萘乙酸的残留。

This paper was to establish method of determination of α-Naphthylacetic acid in cell culture of taxus by HPLC.Cell culture medium was extracted with dichloromethane,while fresh cell was extracted with ethanol（99%） first and then partitioned with dichloromethane.The NAA sample were obtained at a return rate of 87% or more.Chromatographic separation using Nova-pak C18 column,column temperature was 30 ℃,UV detection wavelength was 281 nm.Results：The results showed that the calibration curve was linear over the concentration of α-Naphthylacetic acid in the range of 0.001 ug - 0.125 ug（R=0.9998）.The limits of detection of α-Naphthylacetic acid in cell culture medium was 0.02 mg/L,and the limits of detection of α-Naphthylacetic acid in fresh cells was 0.2 mg/kg.The method is simple,rapid,sensitive,and can determine the residues of α-Naphthylacetic acid in cell culture of taxus.