Metallothionein quantification in clams by reversed-phase high-performance liquid chromatography coupled to fluorescence detection after monobromobimane derivatization

被引：41

作者：

Alhama, J ^{[1
]}

Romero-Ruiz, A ^{[1
]}

López-Barea, J ^{[1
]}

机构：

[1] Univ Cordoba, Dept Biochem & Mol Biol, E-14071 Cordoba, Spain

来源：

JOURNAL OF CHROMATOGRAPHY A | 2006年 / 1107卷 / 1-2期

关键词：

biomarkers of environmental pollution; metals; bivalve mollusks; Chamaellea gallina; SDS-PAGE;

D O I：

10.1016/j.chroma.2005.11.057

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

In this paper, we describe a highly specific, sensitive and reliable method for total metallothionein (MT) quantification by RP-HPLC coupled to fluorescence detection following reaction with monobromobimane of thiols from metal-depleted MT after heat-denaturation of extracts in the presence of sodium dodecyl sulphate (SDS). SDS-polyacrylamide gel electrophoresis (SDS-PAGE) confirmed the identity of the peak resolved (t(R) = 16.44) with MT: a highly fluorescent protein of approximate to 58.3 kDa, in agreement with the high thiol content and low MT size. Other heat-resistant and Cys-containing proteins of 35 kDa were efficiently separated. The new method was successfully used to quantify NIT content in digestive gland of clams from southern Spanish coastal sites with different metal levels, and is proposed as a tool for using MTs as biomarker in monitoring programmes. (c) 2005 Elsevier B.V. All rights reserved.

引用

页码：52 / 58

页数：7

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