Ultrasensitive optical detection of hydrogen peroxide by triggered activation of horseradish peroxidase

Abstract

Hydrogen peroxide is a very reactive byproduct of many metabolic pathways. We describe an ultra-sensitive colorimetric method to detect hydrogen peroxide based on the reconstitution of apo-horseradish peroxidase with the hemin derivative, hemin di(N,N'-acetyl-hydrazide). Oxidation of the latter by hydrogen peroxide yields hemin, which is able to reconstitute apo-horseradish peroxidase, forming an active peroxidase. We have also applied this analyte-triggered reconstitution of horseradish peroxidase to detect the activity of the enzyme glucose oxidase.

An updated view on horseradish peroxidases: recombinant production and biotechnological applications

“…The majority of studies on HRP in biosensor systems focuses on the detection of H 2 O 2 (e.g. Kafi et al 2008 ; Virel et al 2010 ; Zhong et al 2011 ; Ahammad et al 2011 ). In recent years however, a considerable number of studies also dealt with the detection of other molecules, such as glucose (Alonso-Lomillo et al 2005 ), ethanol (Azevedo et al 2005 ), DNA and RNA (Fan et al 2013 ; Tran et al 2014 ; Saikrishnan et al 2014 ), l -phenylalanine (Kubota et al 2013 ), citrinin (Zachetti et al 2013 ), pyrogallol and hydroquinone (Raghu et al 2013 ), phenols (Kafi and Chen 2009 ; Liu et al 2011 ), the milk allergen β-lactoglobulin (Ruiz-Valdepeñas Montiel et al 2015 ), rotavirus titers (Li et al 2014 ), and tumor markers (Chen et al 2009 ; Kim et al 2014b ; Patris et al 2014 ) via H 2 O 2 .…”

Well-redispersed ceria nanoparticles: Promising peroxidase mimetics for H2O2 and glucose detection

“…Based on this phenomenon, we developed a new facile colorimetric method for glucose detection with the CeO 2 NPs as peroxidase mimetics. Compared with various glucose sensors using horseradish peroxidase (HRP), [29][30][31] this method with CeO 2 NPs as peroxidase mimetics has numerous advantages including high stability, low-cost, and ease of preparation. Furthermore, this method is used for glucose detection in human blood serum samples; the results indicate that the highly sensitive and selective glucose detection method has a wide range of potential applications in bioanalysis.…”

Mechanism of Reconstitution/Activation of the Soluble PQQ-Dependent Glucose Dehydrogenase from Acinetobacter calcoaceticus: A Comprehensive Study

“…In numerous cases, this enzyme activation process, also termed enzyme reconstitution, can be achieved in vitro through a precise control of the reaction conditions (temperature, buffer composition, pH …), paving thus the way for fundamental investigations of enzyme structure/reactivity relationships (notably by taking advantage of engineered binding partners such as apoenzyme mutants and/or cofactor analogs) 1,2,3,4,5,6,7,8 or even for discovering new enzyme functionalities. 9,10,11 Besides to the capacity of switching on the activity of an enzyme by simply adding its cofactor, enzyme reconstitution has also been advantageously exploited in different biotechnological applications, 5,10,12,13,14,15,16,17,18,19 ranging from the design of artificial signal transduction systems for analytical purposes to the development of new applications in biotechnology, bioelectronics or synthetic biology. 10,20,21,22,23,24,25 Among the enzymes whose catalytic activity can be easily and efficiently switched on via reconstitution, the soluble quinoprotein glucose dehydrogenase (sGDH, code UniprotKB F0KFV3) is certainly the most prevailing and attractive.…”