Rapid Characterization And Separation,Identification Of Lipase Inhibiting Activity Components In Leaves Of Ilex Cornuta

Obesity is easy to cause a series of diseases such as fatty liver,coronary heart disease,hypertension and diabetes.Obesity seriously affects people’s health and has become one of the public health issues of wide concern in the world.At present,most of the weight-losing drugs have no obvious curative effect and have serious side effects for treating obesity.Therefore,it is necessary to develop drug having a good effect on obesity treatment and less side effects.At present,there are a large number of reports that the leaves of Ilex cornuta have the effect of reducing weight.Firstly,activity screening was carried out by detecting whether the 75% ethanol extract from the leaves of Ilex cornuta could inhibit the expression of PPARγ in 3T3-L1-Lenti-PPARγ-Luc cell line.Secondly,TLC-bioautography-lipase inhibitors screening method was used.The main active ingredients were found to be concentrated in the 60% ethanol elution site.At the same time,this combined method of EFISI-MS with TLC bioautography were used to characterize the structure of the components with lipase inhibitory activity in the 60% ethanol elution site.In order to verify the structure of the active ingredient TLC-EFISI-MS to characterize,a systematic chemical composition study was carried out on the main active part(60% ethanol elution site)and twenty-five compounds were isolated,purified and identified,of which five new compounds.1 Rapid recognition of lipase inhibiting activity constituents in the leaves of Ilex cornutaFractions from leaves of Ilex cornuta after separation by macroporous resin was screened the inhibiting activity of lipase by TLC-bioautography.There were more activity spots in the 60 % elution than the others.fourteen lipase-inhibitory compounds were characterized by TLC-EFISI-MS.And ten ipase-inhibitory compounds were verified by reference substances.These compounds are: asiatic acid(active spots 1),ziyuglycoside II(active spots 2),pomolic acid 28-O-β-D-glucopyra-noside(active spots 3),ziyuglycoside I(active spots 4),ilexside I(active spots 5),ilecornutoside F(active spots 5),ilexcornutoside D(active spots 6-B),(3β,12β)-3-O-[β-D-glucopyranosyl-(1→2)-a-L-arabinopyranosy]-12-hydroxy-19-oxo-18,19-secours-13(18)-en-28,21-lactone(active spots 6-A),ilexpublesnin Q(7),ilexside II(active spots 8),3β-O-[α-L-arabinopyraosyl-(1→2)-β-D-glucuronopyrano-syl]-19α-hydroxyurs-12-en-28-oic acid 28-O-β-D-glucopyranosyl ester(active spots 10-B).Among them,the active spot 6-A has four different configurations(ilexcornutoside A-C and ilexcornutoside F).And the active spot 5 contains not only ilexside I,but also a new compound ilexcornutoside E,which is consistent with its pyrolysis mode.Ilexcornutoside A-E are new compounds.Ilexpublesnin Q was first identified from the sacral leaves.2 Separation and identification of lipase inhibiting activity components in leaves of Ilex cornutaTo identify the structure of the active constituents in the leaves of Ilex cornuta and verified the results of mass spectrometry,60% ethanol elution were separated and purified by traditional column chromatography and preparative liquid chromatography.Twenty-five compounds were isolated,purified and identified,including sixteen triterpenes ilexcornutoside A(1),ilexcornutoside B(2),ilexcornutoside C(3),ilexcornutoside F(6),ilexcornutoside D(4),ilexcornutoside E(5),ursolic acid(7),lupeol(8),uvaol(9),pomolic acid 28-O-β-D-glucopyranoside(10),ziyuglycosides I(12)and II(11),ilexside I(13),3β-O-[α-L-arabinopyraosyl-(1→2)-β-D-glucuronopy-ranosyl]-19α-hydroxyurs-12-en-28-oic acid 28-O-β-D-glucopyranosyl ester(14),ilexoside II(15),and ilexpublesnin Q(16).three flavonoids quercitrin(17),isorhamnetin-3-O-β-D-glucoside(18),kaempferol(19).three lignin(+)-syringaresinol(20),medioresinol(21),(+)-pinoresinol(22).two reduced sesquiterpenes(+)-dehydrovomifoliol(23),(6R,9R)-3-oxo-a-ionol-O-β-D-glucopyranoside(24)and a fatty acid component palmic acid(25),in which compounds 1-5 are new compounds.The absolute configuration of compound 6 was first determined.Compound 1(2)and compound 6(3)are diastereoisomer to each other.Compound 16 and 20 were isolated from this plant for the first time,and compounds 21-24 were isolated from this genus for the first time.The isolated monomeric compounds were screened by lipase inhibitor screening method,and twenty-one compounds of them showed better lipase inhibitory activity.At the same time,we detected whether compound 1-6 inhibited the expression of PPARγ in 3T3-L1-Lenti-PPARγ-Luc cells.Componds(1-3,6)showed inhibitory PPARγ expression activity and compounds 1 and 3 showed a comparable activity in decrease of the PPARγ expression to the positive control(T0070907)at 5 μM.