DOI: 10.3724/SP.J.1123.2017.01026

Chinese Journal of Chromatography (色谱) 2017/35:6 PP.578-586

Investigation on silymarin impact on lipopolysaccharide induced inflammation model based on arachidonic acid metabolism pathway

The objective of this research is to investigate the suppressive effect of silymarin on vitro cell culture model of inflammatory macrophage RAW264.7 induced by Kdo2-Lipid A, and explore its mechanism based on cell metabonomics. Ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) method was used in the cell metabonomic assay to quantitative analysis of metabolites related to eicosanoids pathway. Then chemometric approaches such as principal component analysis were used to process the metabolic data. Within the established method, a total of 59 eicosanoids standards (containing 15 deuterated internal standards) were simultaneously separated in a single 5 min run, and the analytical method is proved to be rapid, sensitive and accurate. Whereafter, the metabolites with VIP> 1 and P value< 0.05 were considered as biomarkers. 12-OxoLeukotriene B4 (12-OxoLTB4) was eventually identified as metabolic biomarkers of silymarin treatment group in this research, and according to the related inflammatory pathways, we speculated silymarin has anti-inflammatory activities by inhibiting the 5-lipoxygenase (5-LOX) activity and blocking lipid peroxidation in 5-LOX metabolic pathways to reduce the formation of peroxides and oxygen free radicals. This study provide a novel approach to the mechanism research on the silymarin treatment on RAW264.7 cells based on cell metabonomics.

Key words:ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS),eicosanoids,deuterated internal standards,silymarin,metabonomics,inflammatory

ReleaseDate:2017-06-16 16:24:25

[1] Dong Y, Kong C Y. Chinese Traditional Patent Medicine, 2010, 32(7): 1225 董岩, 孔春燕. 中成药, 2010, 32(7): 1225

[2] Liu H L. Chinese Journal of Ethnomedicine and Ethnopharmacy, 2008, 17(7): 23 刘洪玲. 中国民族民间医药, 2008, 17(7): 23

[3] Fiebrich F, Koch H. Experientia, 1979, 35(12): 1548

[4] Gupta O P, Sing S, Bani S, et al. Phytomedicine, 2000, 7(1): 21

[5] Yang C, Mai D T, Pan Z M, et al. Chinese Journal of Chromatography, 2016, 34(5): 449 杨婵, 麦旦提, 潘喆敏, 等. 色谱, 2016, 34(5): 449

[6] Masoodi M, Nicolaou A. Rapid Commun Mass Spectrom, 2006, 20(20): 3023

[7] Zehethofer N, Pinto D M. Anal Chim Acta, 2008, 627(1): 62

[8] Han X, Gross R W. J Lipid Res, 2003, 44(6): 1071

[9] US Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Veterinary Medicine (CVM). Federal Register, 2001, 66(4): 206

[10] Hang T J. Pharmaceutical Analysis. 8th ed. Beijing: People's Medical Publishing House, 2011 杭太俊. 药物分析. 8版. 北京: 人民卫生出版社, 2011

[11] Dumlao D S, Buczynski M W, Norris P C, et al. Biochimica et Biophysica Acta, 2011, 1811(11): 724

[12] Buczynski M W, Stephens D L, Bowers-Gentry R C, et al. J Biol Chem, 2007, 282(31): 22834

[13] Yu Y, Cao Y, Chen Y M, et al. Chinese Journal of Chromatography, 2010, 28(7): 644 俞颖, 曹毅, 陈益民, 等. 色谱, 2010, 28(7): 644

[14] Lee J W, Mok H J, Lee D Y, et al. Int J Mol Sci, 2016, 17(4): 508

[15] Nie B, Yu C H, Wang F F, et al. Lishizhen Medicine and Materia Medica Research, 2008, 19(1): 65 聂犇, 余陈欢, 王芳芳, 等. 时珍国医国药, 2008, 19(1): 65

[16] Funk C D, Chen X S. Am J Respir Crit Care Med, 2000, 161: S120

[17] Wang F. [PhD Dissertation]. Hangzhou: Zhejiang University, 2006 汪峰. [博士学位论文]. 杭州: 浙江大学, 2006