title: An in vitro approach for lipolysis measurement using high-resolution mass spectrometry and partial least squares based analysis.
abstract: The elevation of free fatty acids (FFAs) has been regarded as a universal metabolic signature of excessive adipocyte lipolysis. Nowadays, in vitro lipolysis assay is generally essential for drug screening prior to the animal study. Here, we present a novel in vitro approach for lipolysis measurement combining UHPLC-Orbitrap and partial least squares (PLS) based analysis. Firstly, the calibration matrix was constructed by serial proportions of mixed samples (blended with control and model samples). Then, lipidome profiling was performed by UHPLC-Orbitrap, and 403 variables were extracted and aligned as dataset. Owing to the high resolution of Orbitrap analyzer and open source lipid identification software, 28 FFAs were further screened and identified. Based on the relative intensity of the screened FFAs, PLS regression model was constructed for lipolysis measurement. After leave-one-out cross-validation, ten principal components have been designated to build the final PLS model with excellent performances (RMSECV, 0.0268; RMSEC, 0.0173; R2, 0.9977). In addition, the high predictive accuracy (R2= 0.9907 and RMSEP = 0.0345) of the trained PLS model was also demonstrated using test samples. Finally, taking curcumin as a model compound, its antilipolytic effect on palmitic acid-induced lipolysis was successfully predicted as 31.78% by the proposed approach. Besides, supplementary evidences of curcumin induced modification in FFAs compositions as well as lipidome were given by PLS extended methods. Different from general biological assays, high resolution MS-based method provide more sophisticated information included in biological events. Thus, the novel biological evaluation model proposed here showed promising perspectives for drug evaluation or disease diagnosis.
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China.
authors.forename: Wen-Qi
authors.initials: WQ
authors.lastname: Chang
authors.affiliation: Zhejiang Institute for Food and Drug Control, Hangzhou 310052, China.
authors.forename: Jian-Liang
authors.initials: JL
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Li
authors.affiliation: Key Laboratory of New Drug Delivery Systems of Chinese Materia Medica, Jiangsu Provincial Academy of Chinese Medicine, Jiangsu, Nanjing 210028, China.
authors.forename: Zi-Qi
authors.initials: ZQ
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China.
authors.forename: Li
authors.initials: L
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Li
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China. Electronic address: liulifang69@126.com.
authors.forename: Li-Fang
authors.initials: LF
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, No. 24 Tongjia Lane, Nanjing, China. Electronic address: xinguizhong1984@126.com.
authors.forename: Gui-Zhong
authors.initials: GZ
authors.lastname: Xin
mesh_terms: D017667:Adipocytes / Q000187:drug effects*; D000818:Animals; D002138:Calibration; D002478:Cells, Cultured; D003474:Curcumin / Q000737:chemistry; D004353:Drug Evaluation, Preclinical; D016018:Least-Squares Analysis; D008066:Lipolysis; D008297:Male; D013058:Mass Spectrometry; D008813:Mice, Inbred ICR; D019308:Palmitic Acid / Q000737:chemistry
chemical_list: D019308:Palmitic Acid; D003474:Curcumin
keywords: Free fatty acids; Lipolysis measurement; Mass spectrometry; Orbitrap; Partial least squares