Naked-eye sensitive detection of alkaline phosphatase (ALP) and pyrophosphate (PPi) based on a horseradish peroxidase catalytic colorimetric system with Cu(ii)

Abstract

In this paper, a novel colorimetric method for the detection of alkaline phosphatase (ALP) and pyrophosphate (PPi) was designed based on a Cu(2+)-horseradish peroxidase (HRP)-3,3',5,5'-tetra-methylbenzidine (TMB)-H2O2 system. In the presence of ALP, l-ascorbic acid-2-phosphate (AAP) could be hydrolyzed to ascorbic acid which could reduce Cu(2+) to Cu(+) to inhibit the enzymatic activity of HRP in the colorimetric system. The change in absorbance was found to be proportional to the ALP concentration with a linear detection range and a limit of detection of 5.4 mU mL(-1). In the presence of PPi, because Cu(2+) was chelated by PPi, the conversion of Cu(ii) by AA was effectively inhibited. The color of the HRP-TMB-H2O2 system with Cu(2+) showed blue. The HRP-TMB-H2O2 system with the Cu(2+) colorimetric system could also detect PPi with a satisfying result. In summary, this method possesses sensitivity, reproducibility, and cost-effectiveness without labelling and separation and the use of a colorimetric method is more in line with the requirements of on-site detection and green chemistry.

Gold Triangular Nanoplates Based Single-Particle Dark-Field Microscopy Assay of Pyrophosphate

“…In addition, the levels of PPi in cells have become a potential indicator for early diagnosis of cancer, , and it is also used as an additive in toothpaste, detergents, baking powder, and food. , Therefore, the detection of PPi with high sensitivity and high selectivity is still urgently needed. Until now, different approaches including fluorescence and electrochemistry methods have been the commonly used methods. − However, these methods are laborious, operationally cumbersome, and insufficient in real-time measurement.…”

Pyrophosphate Prompted Aggregation‐Induced Emission: Chemosensor Studies, Cell Imaging, Cytotoxicity, and Hydrolysis of the Phosphoester Bond with Alkaline Phosphatase

“…Alkaline phosphatase (ALP) is the enzyme that containing two zinc atom for its catalytic center and active at basic pH . ALP is regularly used enzyme for diagnosis of numerous diseases especially related to liver and bone . It is also employed as a biomarker for liver disease in various pathological laboratory …”

Development of chromogenic detection for biomolecular analysis

“…There are various colorless substrates of peroxidase, including 3,3,5,5‐tetramethylbenzidine (TMB), dopamine (DA), 2,2‐azo (3‐ethylbenzothiazolin‐6‐sulfonic acid) diammonium salts (ABTS), and 3,3‐diaminobenzidine (DAB), which can be oxidized to different colored products. [ 38–44 ] At present, numerous HRP‐based chromogenic detections have been constructed for biomolecular analysis. For example, Cao's group reported a thrombin assay based on HRP‐catalyzed TMB oxidation and proposed a simple electrophoresis titration‐redox boundary model as shown in Figure 1A.…”