Simultaneous Determination of Human Serum Albumin and Low-Molecular-Weight Thiols after Derivatization with Monobromobimane

Molecules

卷:26

Rafał Głowacki¹  Grażyna Chwatko¹  Katarzyna Kurpet² 

[1] Department of Environmental Chemistry, Faculty of Chemistry, University of Lodz, Pomorska 163, 90-236 Lodz, Poland;

[2] Doctoral School of Exact and Natural Sciences, University of Lodz, Banacha 12/16, 90-237 Lodz, Poland;

关键词: low-molecular-weight thiols;    human serum albumin;    α-lipoic acid;    blood plasma;    derivatization;    monobromobimane;   

DOI  :  10.3390/molecules26113321

来源: DOAJ

【 摘 要 】

Biothiols are extremely powerful antioxidants that protect cells against the effects of oxidative stress. They are also considered relevant disease biomarkers, specifically risk factors for cardiovascular disease. In this paper, a new procedure for the simultaneous determination of human serum albumin and low-molecular-weight thiols in plasma is described. The method is based on the pre-column derivatization of analytes with a thiol-specific fluorescence labeling reagent, monobromobimane, followed by separation and quantification through reversed-phase high-performance liquid chromatography with fluorescence detection (excitation, 378 nm; emission, 492 nm). Prior to the derivatization step, the oxidized thiols are converted to their reduced forms by reductive cleavage with sodium borohydride. Linearity in the detector response for total thiols was observed in the following ranges: 1.76–30.0 mg mL⁻¹ for human serum albumin, 0.29–5.0 nmol mL⁻¹ for α-lipoic acid, 1.16–35 nmol mL⁻¹ for glutathione, 9.83–450.0 nmol mL⁻¹ for cysteine, 0.55–40.0 nmol mL⁻¹ for homocysteine, 0.34–50.0 nmol mL⁻¹ for N-acetyl-L-cysteine, and 1.45–45.0 nmol mL⁻¹ for cysteinylglycine. Recovery values of 85.16–119.48% were recorded for all the analytes. The developed method is sensitive, repeatable, and linear within the expected ranges of total thiols. The devised procedure can be applied to plasma samples to monitor biochemical processes in various pathophysiological states.

【 授权许可】

Unknown