Format

Send to

Choose Destination
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Jan 1;1008:212-218. doi: 10.1016/j.jchromb.2015.11.046. Epub 2015 Nov 27.

Highly sensitive and selective detection of human carboxylesterase 1 activity by liquid chromatography with fluorescence detection.

Author information

1
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.
2
Dalian Medical University, Dalian 116044, China.
3
Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
4
Laboratory of Pharmaceutical Resource Discovery, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China. Electronic address: geguangbo@dicp.ac.cn.
5
The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China. Electronic address: dewangjg@hotmail.com.

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.

KEYWORDS:

BMBT hydrolysis; Biological samples; Human carboxylesterase 1 (hCE1); Liquid chromatography-fluorescence detection (LC-FD)

PMID:
26673230
DOI:
10.1016/j.jchromb.2015.11.046
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Elsevier Science
Loading ...
Support Center