3-amino-5-mercapto-1,2,4-triazole (AMT), a 5-membered heterocyclic compound containing a thiol group and an amino group, has been recognized as a good corrosion inhibitor for copper and copper alloys because of its strong adsorption ability on metal surfaces. Yu et al.  investigated the corrosion inhibition effect of AMT against copper corrosion in 3.5 wt.% NaCl solution. The inhibition efficiency (IE) increased with the AMT concentration, and the IE% maintained about 94% with an AMT concentration of 5 × 10?3 mol/L. 3-amino-5-mercapto-1,2,4-triazole is a low-molecular-weight chemical used in the synthesis of organic chemicals, in the processing of silver halide photographic materials, as an antioxidant for aluminum and lubricating oils, and as a viscosity index improver and dispersant. The crystal structure, quantum-chemical calculations, spectroscopic properties and thermal decomposition products of 1,2,4 triazole derivatives have been studied by many authors.

Dermal Exposure to 3-Amino-5-mercapto-1,2,4-triazole Induces Sensitization

Occupational exposure to xenobiotic agents represents a significant cause of asthma, with approximately 28% of all adult cases reported to be related to workplace exposure. A cluster of eight occupational asthma (OA) cases, potentially caused by occupational exposure to N-(2,6-difluorophenyl)-5-methyl-[1,2,4]triazolo[1,5-a]pyrimidine-2-sulfonamide (DE498) and 3-amino-5-mercapto-1,2,4-triazole (AMT), was investigated by the National Institute for Occupational Safety and Health. AMT is a low-molecular-weight chemical used (1) in the synthesis of organic chemicals, (2) in the processing of silver halide photographic materials, (3) as an antioxidant for aluminum and lubricating oils, and (4) as a viscosity index improver and dispersant. To meet these usage needs, 250,000 pounds of AMT was imported into the United States in 1993. Despite the wide potential for exposure to this chemical, there is little toxicity data available. A single study reported a decrease in thyroid peroxidase and thyroxine (T4) concentrations, with a concurrent increase in thyroid weight and thyroid stimulating hormone (TSH) levels in Fischer rats following oral exposure to 3-amino-5-mercapto-1,2,4-triazole.[1]

Initial screening using QSAR yielded negative results for irritancy for both AMT and DE498. Earlier studies using the Draize ocular irritancy rabbit test reported slight irritation at 24 h post DE498 challenge with resolution by 48 h. The absence of any indication of irritancy in the TOPKAT evaluation is incongruent, given that the training set for the irritation module for the TOPKAT program is based on Draize test results (Accelyrs, Inc.). No previous evaluation of the irritancy potential of 3-amino-5-mercapto-1,2,4-triazole was found in the literature.  In DE498, the 3-mercapto group of AMT is incorporated into a sulfonamide structure, while the 5-amino group of AMT is incorporated into a pyrimidine base. Lacking these reactive moieties, the DE498 was negative for sensitization potential in the TOPKAT QSAR model and the LLNA.  In DE498, the 3-mercapto group of AMT is incorporated into a sulfonamide structure, while the 5-amino group of 3-amino-5-mercapto-1,2,4-triazole is incorporated into a pyrimidine base. Lacking these reactive moieties, the DE498 was negative for sensitization potential in the TOPKAT QSAR model and the LLNA. Although the mechanism is not yet defined, the development of dermal sensitization and subsequent pulmonary hyperreactivity in BALB/c mice suggests the potential for 3-amino-5-mercapto-1,2,4-triazole to induce occupational asthma.

Triplet excited state decay of 3-amino-5-mercapto-1,2,4-triazole

In particular, 3-amino-5-mercapto-1,2,4-triazole (AMT) can be considered as one pull–push electronic structure due to both electron donor (–NH2) and acceptor (–SC or –SH) groups bridged by one aromatic 1,2,4-triazole ring. Due to the donor–π–acceptor (D–π–A) structure, AMT demonstrates distinguished physical and chemical behaviour besides the above-mentioned characteristics. Generally, popular hydrogen bonds (H-bonds) involve a combination of an O–H or an N–H proton donor and an N or an O proton acceptor, which demonstrates the double fluorescent phenomenon and large Stokes shift. However, there are only a few reports on H-bonds that involve a sulfur atom as either the proton donor (–SH) or acceptor (S). Moreover, 3-amino-5-mercapto-1,2,4-triazole contains both donor and acceptor of hydrogen, which provides the prerequisite to the proton transfer reaction. Proton transfer has long been an issue of fundamental importance. In this paper, we reported the ground structure and excited state decay dynamics of AMT in different environments using resonance Raman spectra, steady and nanosecond transient absorption spectroscopies combined with theoretical calculation. The decay channels of the singlet excited S2(ππ*) state were investigated by the DFT and TD-DFT calculations.[2]

The crystalline AMT structure was initially confirmed by FT-IR and FT-Raman spectra, as shown. To further assign the experimental spectra, the possible tautomers were calculated at the B3LYP-D3(BJ)/6-311++G(d, P) level. As shown, eight monomeric isomers were found and structure (a) is the lowest one in energy. In conclusion, resonance Raman spectroscopy was employed to assign the vibrational spectra of 3-amino-5-mercapto-1,2,4-triazole (AMT) in methanol and water by a combination of FT-IR, FT-Raman, UV-vis spectroscopy, and density functional theoretical (DFT) calculations. The FT-Raman spectra confirmed that AMT formed a monomer in its neat solid form. In methanol and water, we speculated that the hydrogen bond between the solvent and the AMT molecule might be at –N4H11O and H9N7H8O, and the Franck–Condon region structural dynamics along various reaction coordinates are solvent dependent. After 3-amino-5-mercapto-1,2,4-triazole is excited, we propose three decay channels on the S2 state.

Determination of 3-amino-5-mercapto-1,2,4-triazole in serum

The chemical, 3-amino-5-mercapto-1,2,4-triazole (AMT) is used in a number of commercial applications including pesticide synthesis, processing of silver halide photographic materials, as a viscosity index improver, dispersant, and as an antioxidant for lubricating oils and aluminum. A sensitive and selective analytical method is required in order to carry out toxicokinetic or biomonitoring studies of AMT. This paper describes the development of a method suitable for the detection of AMT in serum. High performance liquid chromatography (HPLC) with fluorescent detection of a highly fluorescent species (AMT-MBB) obtained from the conjugation of AMT to the thiol specific probe, monobromobimane (MBB) was used. Initial work was carried out using aqueous 3-amino-5-mercapto-1,2,4-triazole samples. AMT has a strong UV absorbance from 200 to 280 nm, with a maximum absorbance at 250 nm and local minima around 220 nm. Monitoring of both wavelengths proved useful with respect to establishing peak purity in a complex matrix. Despite good sensitivity (LOD 0.05 μg/ml) and linear standard plots obtained, UV detection of AMT was not suitable for the analysis of AMT because of its poor retention on C18. [3]

In the development of the analytical procedure different approaches to facilitate protein removal and possible 3-amino-5-mercapto-1,2,4-triazole concentration were investigated. In preliminary work a solid phase extraction (SPE) procedure using C18 cartridges was used to concentrate and separate AMT-MBB from serum protein. Preliminary work on AMT spiked urine samples with the omission of the protein removal step indicate that this method could readily be adapted to determine AMT levels in urine samples. Linear standard curves (r2>0.999) within the concentration range 0.78–50 μg/ml (67–4305 pmole on column) were obtained. Problems associated with the apparent instability of MBB hydrolysis products that gave rise to secondary break down products, one of which could not be separated from the AMT-MBB conjugate precluded use of SPE with AMT-MBB. An HPLC method using fluorescence detection was developed for the analysis of AMT in serum. The method involves the reduction of any oxidized 3-amino-5-mercapto-1,2,4-triazole using TBP, precolumn derivatization of free thiol with MBB, separation and quantitation of the AMT adduct using reversed-phase chromatography with fluorescence detection. The assay was found to be simple, specific and sensitive. It should provide a suitable tool for the determination of 3-amino-5-mercapto-1,2,4-triazole in serum and may also have potential industrial hygiene application.

References

[1]Klink, K J, and B J Meade. “Dermal exposure to 3-amino-5-mercapto-1,2,4-triazole (AMT) induces sensitization and airway hyperreactivity in BALB/c mice.” Toxicological sciences : an official journal of the Society of Toxicology vol. 75,1 (2003): 89-98. doi:10.1093/toxsci/kfg171

[2]Duan A, An S, Xue J, Zheng X, Zhao Y. Absorption, fluorescence, Raman spectroscopic and density functional theoretical studies on the singlet and triplet excited state decay of 3-amino-5-mercapto-1,2,4-triazole. RSC Adv. 2020 Apr 5;10(23):13442-13450. doi: 10.1039/d0ra01628d. PMID: 35492984; PMCID: PMC9051463.

[3]Depree, G J, and P D Siegel. “Determination of 3-amino-5-mercapto-1,2,4-triazole in serum.” Journal of chromatography. B, Analytical technologies in the biomedical and life sciences vol. 801,2 (2004): 359-62. doi:10.1016/j.jchromb.2003.11.031