高效液相色谱-蒸发光散射检测法测定藜芦中的介藜芦碱和藜芦胺

建立了高效液相色谱-蒸发光散射检测(HPLC-ELSD)测定藜芦中介藜芦碱、藜芦胺含量的方法,并对4种藜芦属药材样品进行了测定。采用的色谱柱为Kromasil C18柱(250 mm×4.6 mm, 5 μm),以乙腈和0.1%三氟乙酸水溶液为流动相进行梯度洗脱,洗脱程序为:0～5 min, 20%乙腈; 5～30 min, 20%乙腈～40%乙腈, 30～40 min, 40%乙腈～20%乙腈; 40～45 min, 20%乙腈；流速为0.8 mL/min;柱温为35 ℃;采用ELSD检测,漂移管温度为98 ℃,载气流速2.2 L/min 。介藜芦碱和藜芦胺的线性范围分别为42.05～980 mg/L和43.77～1020 mg/L;平均回收率分别为99.2%和101.4%,相对标准偏差分别为1.7%和2.1% (n=6);信噪比为3时,测得介藜芦碱和藜芦胺最低检测限分别为18.37 mg/kg和21.50 mg/kg。该方法快速简便、灵敏度和分离度好,适用于藜芦药材中活性生物碱的测定。

Determination of jervine and veratramine in veratrum plants using high performance liquid chromatography coupled with evaporative light scattering detection

A method of reversed-phase high performance liquid chromatography (HPLC) coupled with evaporative light scattering detection (ELSD) was developed for the determination of jervine and veratramine in veratrum plants. The extraction method of total active alkaloids from the raw material was also established. The separation and quantification were achieved using a Kromasil C8column (250 mm x 4.6 mm, 5 microm), and a mobile phase of acetonitrile and 0.1% trifluoroacetic acid with the following gradient elution: 20% acetonitrile at the first 5 mm, 20%-40% acetonitrile at the 5-30 mm, 40%-20% acetonitrile at the 30-40 mm, 20% acetonitrile at the 40-45 mm with a flow rate of 0.8 mL/min; column temperature of 35 t and monitored by an ELSD detector with the drift tube of 98 t and the nitrogen flow rate of 2.2 L/min. The calibration curves for jervine and veratramine were linear over the ranges of 42.05-980 mg/L and 43.52-1020 mg/L, respectively. The recoveries were 99.2% and 101.4% with relative standard deviations of 1.7% and 2.1% (n=6), respectively. The limits of detection for jervine and veratramine in raw material were 18.37 mg/kg and 21.50 mg/kg, respectively, with 3 times of the signal to noise ratio. This HPLC-ELSD method is rapid, simple, accurate and convenient. It can be used as one of the direct and reliable means for quantitative determination of the active alkaloids in veratrum plants.