Application of the dibromohydroxyphenylfluorone–molybdenum(VI) complex to the sensitive spectrophotometric determination of protein

• Published in: Analytica chimica acta 2000-01, Vol.403 (1), p.225-233
• Main Authors: Guo, Zhong-Xian, Hao, Yong-Mei, Cong, Xin, Shen, Han-Xi
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; Binding reaction; Biological and medical sciences; Body fluids; Dibromohydroxyphenylfluorone; Fundamental and applied biological sciences. Psychology; General aspects, investigation methods; Molybdenum(VI); Protein; Proteins; Spectrophotometry
• ISSN: 0003-2670; 1873-4324
• DOI: 10.1016/S0003-2670(99)00564-4

A new method is described for the spectrophotometric determination of protein, based on the binding interaction of protein, molybdenum(VI) and dibromohydroxyphenylfluorone (DBHPF). At pH 2.0–3.4 and in the presence of Triton X-100, the binding reaction is complete within 8 min at room temperature, and causes a change of the absorption spectrum and an absorbance decrease at 533 nm of the DBHPF–Mo(VI) complex. The calibration graphs for bovine and human serum albumins are both linear up to 8.0 mg 1 −1 with 3 σ detection limits of 70 and 80 ng ml −1, respectively. Compared with some accepted and reported assays, besides being highly sensitive, the method is reproducible and simple, and there is a lack of interferences. Results for a culture, cell homogenate and human body fluids are in agreement with those obtained by the Bradford method, with relative standard deviations of 1.5–2.5% ( n = 5). The binding numbers and association constants of serum albumins with the complex are estimated by a modified Scatchard graphical method, and the binding mechanism is discussed.