title: Highly sensitive and selective detection of human carboxylesterase 1 activity by liquid chromatography with fluorescence detection.
abstract: Human carboxylesterases 1 (hCE1), one of the most important human drug metabolizing enzymes, catalyzes the hydrolysis of a large number of structurally diverse of endogenous and exogenous substrates. However, a practical, reliable and sensitive method for the precise measurement of hCE1 activities in complex biological samples has been rarely reported. In this study, a liquid chromatography-fluorescence detection (LC-FD) based method was developed for highly selective and sensitive measurement of hCE1 activities in human tissue and cell preparations. This method was based on the fluorimetric detection of HMBT, the hydrolyzed product of BMBT which was a newly developed specific probe substrate for hCE1. The developed LC-FD method was fully validated in terms of specificity, sensitivity, linearity, precision, recovery and stability. With the help of LC separation, most polar endogenous compounds in biological samples could be eluted in the column dead time, which is very beneficial for accurate determination of hCE1 activities in complex biological samples. The lower limit of quantification for HMBT (product of hCE1) of this LC-FD based method was as low as 20nM, which was quite lower than other reported methods. The method also exhibited good precision, both intra- and inter- assay variances were both lower than 2.5%. Furthermore, the newly developed method was successfully applied to measure hCE1 activity in human liver preparations from individual donors (n=12), as well as in homogenates from eleven different human cell lines. All these findings combined with this practical method are very helpful for the deep understanding of the expression and function of hCE1 in human biological samples.
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China; University of the Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Dan-Dan
authors.initials: DD
authors.lastname: Wang
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China; University of the Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Jin
authors.affiliation: Dalian Medical University, Dalian 116044, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Hou
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Feng
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China; University of the Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Na
authors.initials: N
authors.lastname: Li
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Shi-Yang
authors.initials: SY
authors.lastname: Li
authors.affiliation: Dalian Medical University, Dalian 116044, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Zhou
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Li-Wei
authors.initials: LW
authors.lastname: Zou
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China. Electronic address: geguangbo@dicp.ac.cn.
authors.forename: Guang-Bo
authors.initials: GB
authors.lastname: Ge
authors.affiliation: The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China. Electronic address: dewangjg@hotmail.com.
authors.forename: Jin-Guang
authors.initials: JG
authors.lastname: Wang
authors.affiliation: Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Ling
authors.initials: L
authors.lastname: Yang
mesh_terms: D002265:Carboxylic Ester Hydrolases / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D008862:Microsomes, Liver / Q000201:enzymology; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D002265:Carboxylic Ester Hydrolases; C474609:CES1 protein, human
keywords: BMBT hydrolysis; Biological samples; Human carboxylesterase 1 (hCE1); Liquid chromatography-fluorescence detection (LC-FD)