Horseradish peroxidase (HRP)-based assays feature particular interests because of the simple colorimetric readout. In these assays, 3,3′,5,5′-tetramethylbenzidine (TMB) is the most widely used chromogenic substrates for HRP. The later research in nanozyme and DNAzyme also used TMB (the chosen substrate) because they are both HRP-mimics. It should be noted that the substrate of HRP is not just limited to TMB but, in fact, a broad range of benzidine derivatives. However, except decreased carcinogenicity due to tetrasubstitution of benzidine, the rationale for the chosen substrate TMB is not clear yet. Here, we addressed such a fundamental issue from the chemistry point of view. Nine benzidine derivatives featuring varied properties (different substitution groups and varied number of substitutions) were selected and investigated with four typical TMB-involved chromogenic systems. Among the existing benzidine substrates that are used for peroxidase-based assays, TMB exhibited the highest sensitivity, better color purity of colored products, and reasonable stability of oxidation products. Moreover, two tetrasubstituted benzidine derivatives other than TMB (4OCH 3 and 2OCH 3 2CH 3 ) were synthesized for comparison. It turned out that the performances (sensitivity, color purity, and stability of the colored products) of TMB are still superior, thus chemically confirming its status of “the chosen substrate” in colorimetric assays.

D2O plays important roles in a variety of fields (such as the nuclear industry and bioorganic analysis), and thus its isotopic purity (H2O contents) is highly concerned. Due to its highly similar physical properties to H2O and large excess amounts of H2O over D2O, it is challenging to distinguish D2O from H2O. On the basis of the characteristic NIR-II phosphorescence of singlet oxygen (1O2), and the fact that H2O is a more efficient quencher for 1O2 than D2O, here, we proposed to simply use the 1275 nm emission of 1O2 for the analysis of the isotopic purity of D2O. In normal cases (a xenon lamp for excitation), such steady-state emission is extremely weak for valid analytical applications, we thus employed laser excitation for intensification. To this goal, a series of photosensitizers were screened, and eventually polythiophene PT10 was selected with high singlet oxygen quantum yield (ΦΔ = 0.51), high H2O/D2O contrast, and excellent photostability. Upon excitation with a 445 nm laser, a limit of detection (LOD, 3σ) of 0.1% for H2O in D2O was achieved. The accuracy of the proposed method was verified by the analysis of the isotopic purity of several D2O samples (with randomly added H2O). More interestingly, the hygroscopicity of D2O was sensitively monitored with the proposed probe in a real-time manner; the results of which are important for strengthening the care of D2O storage and the importance of humidity control during related investigations. Besides D2O isotopic purity evaluation, this work also indicated the potential usefulness of the NIR-II emission of singlet oxygen in future analytical detection.

Ultraviolet radiation (UVR) is both useful to human beings and can cause irreversible harm of varying degrees (UVA, UVB, and UVC). Especially, in areas with excessive sunlight, the appearance of UVB results in an increased risk of skin cancer. On the other hand, UV lamps (254 nm, UVC) are widely used in disinfection (air, water, and factory food) and hospital sterilization; the leakage of UVC is thus sometimes inevitable, which may cause fatal injuries to the related staff. Therefore, low-cost UV dosimetry-based personal protective equipment (PPE) and industrial monitoring devices are of great importance. Here, for the first time, we found that 3,3′,5,5′-tetramethylbenzidine (TMB) could be rapidly oxidized upon UVB and UVC irradiation in a dose-dependent manner, in which TMB acts as a self-photosensitizer. Since TMB is a typical and widely used chromogenic substrate in enzyme-linked immunosorbent assay (ELISA), it is well-commercialized with low cost and vast availability worldwide, which permitted the development of low-cost naked-eye UVB and UVC dosimetry. A wearable bracelet mounted with TMB-loaded paper was developed for successful indication of whether the UVB exposure in the sunlight exceeded the minimum erythema dose (MED). In addition, we also developed a clock dial equipped with a TMB solution for unattended detection of UVC leakage from UVC disinfection lamps. The UVB-and UVC-selective coloration and low cost of TMB offered remarkable potential in facile detection of UVR in our daily life.

No abstract

As key molecules in most biological pathways, proteins physically contact one or more biomolecules in a highly specific manner. Several driving forces (i.e., electrostatic and hydrophobic) facilitate such interactions and a variety of methods have been developed to monitor these processes both in vivo and in vitro. In this work, a new method is reported for the detection of protein interactions by visualizing a color change of a cyanine compound, a supramolecule complex of 3,3-di-(3-sulfopropyl)-4,5,4',5'-dibenzo-9-methyl-thiacarbocyanine triethylammonium salt (MTC). Nuclear magnetic resonance (NMR) studies suggest that the hydrophobic nature of the protein surfaces drives MTC into different types of aggregates with distinct colors. When proteins interact with other biomolecules, the hydrophobic surface of the complex differs, resulting in a shift in the form of MTC aggregation, which results in a color change. As a result, this in vitro method has the potential to become a rapid tool for the confirmation of protein-biomolecule interactions, without the requirements for sophisticated instrumentation or approaches.