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

Abstract

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.

Engineered bacteria titrate hydrogen sulfide and induce concentration-dependent effects on host in a gut microphysiological system

“…Solvent B was 90% methanol and 0.25% glacial acetic acid, adjusted to pH 3.9. The protocol was adapted and modified from previously described protocols 74,75 .…”

Metal ion‐mediated carbon dot nanoprobe for fluorescent turn‐on sensing of N‐acetyl‐l‐cysteine

“…[6][7][8][9] On account of its superiority in the fields of biology and medicine, NAC identification and quantification have been studied for many years and several methods have been well designed and performed for NAC detection. Instrument methods including chromatography, [10][11][12] electrophoresis analysis, [13] mass spectrometry, [14] chemiluminescence analysis, [15,16] electrochemistry method, [17] spectrophotometry, [18] and fluorescence analysis [19][20][21][22] have been successfully established for NAC determination in human tissue and pharmaceutical samples. Among these methods, fluorescence analysis holds the merits of simple operation, rapid response, sensitive, and convenient detection, making it a promising method for pharmaceutical analysis.…”