title: Facile colorimetric assay of alkaline phosphatase activity using Fe(II)-phenanthroline reporter.
abstract: We report a versatile approach for the colorimetric assay of alkaline phosphatase (ALP) activity based on the distinctive metal-to-ligand charge-transfer (MLCT) absorption properties of Fe(II)-phenanthroline reporter. In the presence of ALP, the applied substrate ascorbic acid 2-phosphate is enzymatically hydrolyzed to produce ascorbic acid, which then reduces Fe3+to Fe2+. The complexation of Fe2+with the bathophenanthroline disulfonate (BPS) ligand generates a blood-red Fe(BPS)34-reporter, which is characterized by an intense MLCT absorption band at 535 nm in the visible range. Under optimal conditions, the spectral output exhibits a good quantitative relationship with ALP activity over the range of 0-220 mU mL-1with a detection limit of 0.94 mU mL-1. Moreover, the activity of ALP can also be conveniently judged through naked-eye observations. Results indicate that it is highly selective and can be applied to the screening of ALP inhibitors. In addition, it has been successfully employed to detect the endogenous ALP level of undiluted human serum samples, with a detection limit of 1.05 mU mL-1being achieved. This approach avoids any elaborately designed substrates and holds considerable simplicity and flexibility for reporter design. This study broadens the horizon of the applications of phenanthroline-based transition metal complexes. Furthermore, an efficient and practical method like this has the potential to be widely used in clinical applications and in the point-of-care testing.
authors.affiliation: School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094, PR China.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Hu
authors.affiliation: School of Chemical Engineering, Nanjing University of Science and Technology, Nanjing, 210094, PR China.
authors.forename: Baojing
authors.initials: B
authors.lastname: Zhou
authors.affiliation: School of Chemical Engineering, Nanjing University of Science and Technology, Nanjing, 210094, PR China.
authors.forename: Pengyun
authors.initials: P
authors.lastname: Dang
authors.affiliation: School of Chemistry and Chemical Engineering, Liaocheng University, Liaocheng, 252000, PR China.
authors.forename: Lianzhi
authors.initials: L
authors.lastname: Li
authors.affiliation: School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094, PR China. Electronic address: j.kong@njust.edu.cn.
authors.forename: Jinming
authors.initials: J
authors.lastname: Kong
authors.affiliation: School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing, 210094, PR China; Chemistry Department, College of Arts and Sciences, University of South Florida, East Fowler Ave, Tampa, FL, 33620-4202, United States. Electronic address: xueji@usf.edu.
authors.forename: Xueji
authors.initials: X
authors.lastname: Zhang
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis*; D003124:Colorimetry; D005296:Ferrous Compounds / Q000737:chemistry; D006801:Humans; D057230:Limit of Detection; D010618:Phenanthrolines / Q000737:chemistry*; D044967:Serum / Q000737:chemistry
chemical_list: D005296:Ferrous Compounds; D010618:Phenanthrolines; D000469:Alkaline Phosphatase
keywords: Alkaline phosphatase; Bathophenanthroline disulfonate; Colorimetric; Density functional theory; Electrochemical; Metal-to-ligand charge-transfer