高效液相色谱法同时测定明党参原植物和离体培养材料中3种呋喃香豆素含量

Simultaneous Determination of Furanocoumarins in Cultivated Changium smyrnioides Wollf and Its in Vitro Cultures by HPLC

目的建立测定明党参原植物及离体培养材料中3个代表性呋喃香豆素成分花椒毒素、佛手酚和佛手柑内酯的高效液相色谱方法。方法采用HPLC-PDA,Agilent-C18色谱柱(4.6 mm×250 mm,5μm),流动相为甲醇-水梯度洗脱,流速1.0 m L·min^(-1),检测波长314 nm,进样量10μL。结果所建方法可同时测定明党参样品中3种香豆素含量,花椒毒素、佛手酚和佛手柑内酯的线性方程分别为Y=17 057ρ-87.689(r=1.000 0)、Y=23 828ρ-380.44(r=0.999 9)、Y=37 123ρ-441.16(r=0.999 9)。原植物和再生植株中花椒毒素与佛手柑内酯含量明显高于愈伤组织和悬浮细胞,但佛手酚在前两者中未检出而在后两者中有少量积累,源于叶和柄细胞的样品3种香豆素含量普遍较高。结论所建方法用于测定明党参中3种香豆素含量,准确、灵敏、重复性良好。该法为明党参资源有效开发与利用,尤其为利用生物技术进一步探索香豆素类有效成分的调控奠定基础。

OBJECTIVE To determine simultaneously the contents of xanthotoxin,bergaptol, and bergapten in cultivated Changium smyrnioides Wollf and its in vitro cultures by HPLC.METHODS Agilent-C18 column （4.6 mm×250 mm,5 μm） was used for chromatographic separation and PAD was applied as detector. The flow rate was 1.0 mL·min-1with a mobile phase of methanol-water in gradient elution mode. The detection wavelength was set at 314 nm while the injection volume was 10 μL.RESULTS The linear regression equations of xanthotoxin, bergaptol, and bergapten were Y=17 057ρ-87.689 （r=1.000 0）, Y=23 828ρ-380.44 （r=0.999 9） and Y=37 123ρ-441.16（r=0.999 9）, respectively. The contents of xanthotoxin and bergapten in cultivated and regenerated specimens were obviously higher than those in calli and suspension cells, while bergaptol was only detected in the latter two samples. A larger amount of furanocoumarins accumulated in the cells from leaves and petioles. CONCLUSION The established method is simple and effective with high sensitivity and good repeatability. It can be adopted in studies on the utilization of Changium smyrnioides, especially the regulation and induction of furanocoumarins.