Study On Distribution Of Biflavanones In Seed Coat,Kernels And Slices Of Garcinia Kola Seeds And Anti-inflammatory Activity Of Components Compatibility

Garcinia kola(G.kola)seeds were the dried seeds of the Garcinia kola Heckel.They play an important role in African traditional medicine,which are widely used to treat stomachache,gastritis,malaria,venereal diseases and laryngitis.Biflavanones are the main components according to the qualitative analysis of G.kola seeds in the previous study which have various pharmacological activities of hypoglycemic,hypolipidemic and hepatocyte protection.But there have been few studies on investigation of biflavanones distribution among G.kola seeds in coat,kernels and slices.Also,attention was hardly paid to resin exuded from fresh slice.Aiming at above problems,this study focused on the quantitative study of four biflavanones in G.kola seeds and carried out the characteristics of the content of biflavanones in G.kola seeds from different habitats and specification(weight),the characteristics of biflavanones distribution in coat and kernels,optimization of the slicing process and identification of compounds from gum resin exuded from fresh slice.On this basis,the antiinflammatory activities of crude extracts of G.kola seeds,different compatible components(biflavanone and garcinoic acid components)and garcinoic acid were carried out.The active ingredients and its component compatibility characteristics of G.kola seeds were systematically revealed.1.Focusing on the main and biologically active components,a rapid high performance liquid chromatography(HPLC)method was established for quantitative study of the four biflavanones,which was validated methodologically.All calibration curves showed good linearity.The RSD values of the precision were not more than 0.8%.The RSD values of the stability were not more than 0.7%.The repetitive RSD values were not more than 2.4%.The recovery rates of the samples were between 98.31% and 99.89% with RSD values not more than 2.4%.2.The content of the biflavanones in different habitats and specification(weight),different parts(seed coat and kernels)and thickness of different slices were systematically developed.The main constituents of gum resin exuded from fresh slices during the slicing process were identified by mass spectrometry.There was no significant difference among the contents of the tested biflavanones in G.kola seeds from different habitats.Moreover,no obvious influence of seeds weight was observed on the biflavanones content in G.kola seeds from the same habitat.By the aid of “spider-web” mode,it was unveiled that the accumulation of biflavanones mainly occurred in seed kernels.Furthermore,slice thickness(1-2 mm)was optimized by comprehensive evaluation of biflavanones content,moisture,cutting-time and drying-time to efficiently dry the slices.Interestingly,during slicing,the brownish yellow or brownish red spots gradually appeared on the surface of fresh slices,which was derived from gum resin exuded.By employing of UHPLC/ESI Q-Orbitrap MS,garcinoic acid and its derivatives were surprisingly identified in gum resin,which was significantly different from the biflavanones contained in the slices.3.In order to study the anti-inflammatory active part of G.kola seeds and its component compatibility characteristics,pre-HPLC was used to separate crude extract of G.kola seeds into biflavanone component and garcinoic acid component.Quantitative characterization of the main compounds,GB-2,GB-1,kolaflavanone,GB-1a and garcinoic acid,in the crude extract and components was performed by HPLC.Rapid separation and preparation of biflavanone and garcinoic acid components were successfully achieved by pre-HPLC.GB-2,GB-1,kolaflavanone and GB-1a compounds accounted for 27.71% of the crude extract and gracinoic acid accounted for 4.998%.GB-2,GB-1,kolaflavanone and GB-1a accounted for 29.80% of the biflavanone component,and garcinoic acid made up 55.93% of the garcinoic acid component.4.An LPS-induced RAW264.7 inflammation model was established for anti-inflammatory activity study on the crude extract of G.kola seeds,the component of biflavanone,the component of garcinoic acid,simulated crude extract,different proportion components and garcinoic acid,which took NO,TNF-?,IL-1? and IL-6 proinflammatory factors in the cell supernatant as indicators.We found that the above groups can exert anti-inflammatory effects at different degrees.The biflavanone component and the garcinoic acid component showed a good synergistic anti-inflammatory effect.The garcinoic acid component was the main active component that exerted anti-inflammatory effects and the garcinoic acid was the main compound of the garcinoic acid component which was a potential active compound.Through the study of this subject,the distribution characteristics of biflavanones in G.kola seeds from different habitats,specification(weight),different parts and different thicknesses were clarified.The chemical constituents of brownish yellow or brownish red gum resin were identified,the anti-inflammatory activities of different components and compounds of G.kola seeds were clarified.It is of great significance for the comprehensive development and utilization of G.kola seeds.