Determination of four major steroid glycosides in Anemarrhena asphodeloides by HPLC-ELSD
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摘要: 目的 建立同时测定知母药材中4种主要皂苷成分的HPLC-ELSD方法。 方法 采用Agilent Zorbax SB-C18柱(4.6 mm×250 mm, 5 μm), 以水(冰醋酸调节pH 3.3)(A)-乙腈(B)为流动相进行梯度洗脱,0~8 min,12%~23% B;8~25 min,23% B;25~35 min,23%~45% B;35~40 min,45%~95% B。蒸发光散射检测器漂移管温度55℃,以氮气为雾化气,压力为4.0 Bar。 结果 知母皂苷B、知母皂苷E1、知母皂苷BⅡ、知母皂苷A Ⅲ浓度分别在42.10~252.6 μg/ml(r=0.999 3)、48.80~292.8 μg/mlr=0.999 1)、192.2~1 153 μg/ml(r=0.999 7)、8.512~85.12 μg/ml(r=0.998 5)的范围内呈良好的线性关系。4种成分精密度试验RSD<1%,48 h内稳定性RSD<1%,加样回收率为96.04%~102.8%。 结论 该含量测定方法简便,分离效果好,能同时测定知母皂苷B、知母皂苷E1、知母皂苷BⅡ和知母皂苷AⅢ 4种有效成分的含量,结果准确可靠。Abstract: Objective To develop a new high performance liquid chromatography (HPLC) coupled with Evaporative Light Scattering Detector (ELSD) method for simultaneous determination of four major steroid glycosides (Timosaponin B, Timosaponin E1, Timosaponin BⅡ and Timosaponin AⅢ). Methods HPLC analysis was performed on an Agilent Zorbax SB-C18 column (4.6 mm×250 mm, 5 μm) with a mobile phase of water (adjust to pH 3.3 by acetic acid) (A) and acetonitrile (B). The gradient elution program was as follow:0~8 min, 12%~23% B;8~25 min, 23% B;25~35 min, 23%~45% B;35~40 min, 45%~95% B. The temperature of drift tube was 55℃ and the nebulizer nitrogen flow rate was 4.0 Bar. Results The linearity was obtained over 42.10~252.6 μg/ml (r=0.999 3) for Timosaponin B, 48.80~292.8 μg/ml (r=0.999 1) for Timosaponin E1, 192.2~1153 μg/ml (r=0.999 7) for Timosaponin BⅡ and 8.512~85.12 μg/ml (r=0.998 5) for Timosaponin AⅢ. The RSDs of precision and stability of the samples were both less than 1% in 48 hours. The average recovery was between 96.04%~102.8%. Conclusions The present method, with satisfactory efficacy, was simple which could simultaneously determine multiple steroid glycosides in Anemarrhena asphodeloides Bge. from different areas.
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