3,3′,5,5′-tetramethylbenzidine as multi-colorimetric indicator of chlorine in water in line with health guideline values

Abstract

Sanitizing solutions against bacterial and viral pathogens are of utmost importance in general and, in particular, in these times of pandemic due to Sars-Cov2. They frequently consist of chlorine-based solutions, or in the direct input of a certain amount of chlorine in water supply systems and swimming pools. Colorimetry is one of the techniques used to measure the crucial persistence of chlorine in water, including household chlorine test kits commonly based on colorimetric indicators. Here, we show a simple and cheap colorimetric method based on 3,3′,5,5′-tetramethylbenzidine (TMB), commonly used as chromogenic reagent for enzyme-linked immunosorbent assays. TMB is converted by chlorine to a colored molecule through a pH-dependent multi-step oxidation process where the chromaticity of TMB is directly proportional to chlorine content. This molecule offers several advantages over other commonly used reagents in terms of safety, sensitivity, and, peculiarly, hue modulation, giving rise to the detection of chlorine in water with a multi-color change of the indicator solution (transparent/blue/green/yellow). Moreover, through the appropriate setting of reaction conditions, such coloration is finely tunable to cover the range of chlorine concentration recommended by international health agencies for treatment of drinking water and swimming pools and to test homemade solutions prepared by dilution of household bleach during health emergency events such as during the current pandemic.

Serotonin-Derived Fluorophore: A Novel Fluorescent Biomaterial for Copper Detection in Urine

“…Based on these results, and considering that, to the best of our knowledge, there are no studies on copper ion detection exploiting the fluorescence quenching of a polymer derived by an endogenous molecule, such as SEDF, we decided to apply this method to real urine samples, where the presence of large amounts of copper ions would be indicative of a pathological condition (see below), whereas the concomitant presence of the other quenching ions, in particular Au 3+ and Cr 2 O 7 2− , is insignificant in urine [ 38 , 39 ], and the urinary level of iron in healthy people (0.2 mg L −1 , 0.003 mM) is approximately two orders of magnitude below the concentration of Fe 3+ here tested (0.5 mM), and one order of magnitude below the minimum amount of Cu 2+ here spiked in urine (0.05 mM, vide infra). However, there is also the possible interference of larger urinary levels, as a consequence of other iron-overload diseases, which could be simply minimized by using the proper sequestering buffer, as already reported [ 40 , 41 ]. Moreover, we also evaluated the interference of the main organic compounds in urine on SEDF fluorescence quenching.…”

Determination of the Dissociation Constant for Polyvalent Receptors Using ELISA: A Case of M13 Phages Displaying Troponin T-Specific Peptides

“…When the reaction is stopped by the addition of sulfuric acid, the enzyme is denatured. As a result, the blue complex dissociates into a yellow product (adsorption maximum at 450 nm) and transparent TMB. , …”

Determination of the Dissociation Constant for Polyvalent Receptors using ELISA: a Case of M13 Phages Displaying Troponin T-specific Peptides

“…The surface density of HRP enzyme (i.e., the amount of HRP immobilised on the liquid|solid interface) is a crucial factor affecting the rate of substrate (3,3',5,5'-tetramethylbenzidine, TMB) oxidation. The 14 reaction rate may be expressed by the rate of the change of absorbance at 370 nm (which is the maximum of absorption of yellow by-product 43 ) under the assumption that the kinetics of the enzymatic reaction is zero order (Michaelis-Menten model and enzyme saturation). In this case, the measured signal is the enzyme activity defined as the slope of the linear time function of absorbance (Eq.13).…”

“…When the reaction is stopped by addition of sulfuric acid, the enzyme is denatured. As a result, the blue complex dissociates into a yellow product (adsorption maximum at 450 nm) and transparent TMB 43,44 .…”