Study On The Isolation Methods Of Lignans From Phyllanthus Niruri Linn

Lignans from Phyllanthus niruri Linn attract people's attention for their anti-HBV activity.Two isolation methods of lignans were developed in,this paper.The crude extract of lignans was obtained from Phyllanthus niruri Linn using ethanol first and then petroleum ether.With flash column chromatography,preparative thin layer chromatography,selective precip itation and recrystallization,three compounds were isolated from the crude extract of lignans,their structures were elucidated by spectroscopic methods,including IR,NMR,as well as MS.As a result,the compounds were identified as stereoisomer of niranthin,hypophyllanthin,andphyllanthin.However,the stereo configuration of stereoisomer of niranthin needed a further confirmation.High-speed countercurrent chromatography(HSCCC)was applied to the isolation of lignans from the crude extract.Firstly,the effects of polyethylene glycol(PEG)10000 on the isolation of lignans were respectively tested in solvent system composed of n-hexane-ethyl acetate-methanol-water(3:2:3:2,V/V(1)and n-hexane-ethyl acetate-ethanol-water(6:4:5.5:5,V/V)(2).The results demonstrated that the effects were weak.Secondly,without PEG,temperature and revolution speed were respectively optimized in solvent system(1)and(2),gave the optimized parameters,25? and 800 r/min,in both solvent systems.Resolution of hypophyllanthin and phyllanthin were attained under optimized conditions in both solvent systems,in solvent system (1)it was 0.952,and in solvent system(2)it was 0.929.Besides,in both solvent systems,resolution of stereoisomer of niranthin and hypophyllanthin were larger than 1.5,which indicating a complete separation.Thus,conclusion can be drawn that the optimized conditions in HSCCC were solvent system:n-hexane-ethyl acetate-methanol-water(3:2:3:2,V/V);mobile phase:the lower phase;flow rate:2 mL/min;temperature:25?;revolution speed:800 r/min.F inally,under optimized conditions a large amount of sample,257.3 mg,was injected to evaluate the preparative capacity of HSCCC at large sample load,and it worked well,gave 84.8 mg stereoisomer of niranthin,31.9 mg hypophyllanthin and 68.2 mg phyllanthin with purities of 69.97%,90.24%,and 86.78%,respectively.A comparison between flash column chromatography and HSCCC was discussed.