Determination of three active component in Anoectochilus roxburghii(Wall) Lindl with ultrasound-microwave combined on extraction by RP-HPLC

Objective To establish a method for extraction and determination of flavonodis in Anoectochilus roxburghii(Wall)Lindl.Methods The effect of ultrasound-microwave combined on extraction of Anoectochilus roxburghii(Wall)Lindl has been studied.The optimized condition of extraction of flavonoids was studied with orthogonal design.The reversed phase HPLC system consisting of an Agilent ODS column(4.6 mm×250 mm,5 μm)and a mixture of CH3OH-H2O-H3PO4(52 ∶ 48 ∶ 0.4)as the mobile phase was used.The detection wavelength was at 368 nm and the flow rate was at 1.0 ml/min.Results The quercetin,kaempferorl and isorhamnetin were well separated by this method.Linearities of quercetin,kaempferorl and isorhamnetin were good(r=0.999 5,0.999 6,0.999 7)in ranges of 5.0～80.0,5.0～80.0,10.0～160.0 μg,respectively.The average recoveries were 98.2%,97.9%,98.4%.The contents of quercetin,kaempferol and isorhamnetin was from 0.109～0.197 mg/g,0.089～0.126 mg/g and 0.281～0.356 mg/g,groups of Anoectochilus roxburghii(Wall)Lindl from different location.Conclusion The method for extraction and determination is simple,accurate and can be used for the quality study of Anoectochilus roxburghii(Wall)Lindl.