Spectroscopic and Microestructural Evidence for T-2 Toxin Adsorption Mechanism by Natural Bentonite Modified with Organic Cations.

Aluminosilicates are adsorbents able to bind mycotoxins, and their chemical modification increases their affinity to adsorb low-polarity mycotoxins. To further investigate if the inclusion of salts in bentonite modifies its adsorptive capacity, we studied T-2 toxin adsorption in natural bentonite (NB) and when modified with quaternary ammonium salts differing in polarity and chain length: myristyl trimethyl ammonium bromide (B14), cetyl trimethyl ammonium bromide (B16) and benzyl dimethyl stearyl ammonium chloride (B18). The results showed that quaternary salts made bentonite: displace monovalent (Na+1, K+1) and divalent (Mg+2, Ca+2) ions; reduce its porosity; change its compaction and structure, becoming more crystalline and ordered; and modify the charge balance of sheets. T-2 adsorption was higher in all modified materials compared to NB (p ≤ 0.0001), and B16 (42.96%) better adsorbed T-2 compared to B18 (35.80%; p = 0.0066). B14 (38.40%) showed no differences compared to B16 and B18 (p > 0.05). We described the T-2 adsorption mechanism in B16, in which hydrogen bond interactions, Van der Waals forces and the replacement of the salt by T-2 were found. Our results showed that interaction types due to the inclusion in B16 might be more important than the hydrocarbon chain length to improve the adsorptive capacity of bentonite.

Molecular Docking and In Vitro Studies of Ochratoxin A (OTA) Biodetoxification Testing Three Endopeptidases.

Ochratoxin A (OTA) is considered one of the main mycotoxins responsible for health problems and considerable economic losses in the feed industry. The aim was to study OTA's detoxifying potential of commercial protease enzymes: (i) Ananas comosus bromelain cysteine-protease, (ii) bovine trypsin serine-protease and (iii) Bacillus subtilis neutral metalloendopeptidase. In silico studies were performed with reference ligands and T-2 toxin as control, and in vitro experiments. In silico study results showed that tested toxins interacted near the catalytic triad, similar to how the reference ligands behave in all tested proteases. Likewise, based on the proximity of the amino acids in the most stable poses, the chemical reaction mechanisms for the transformation of OTA were proposed. In vitro experiments showed that while bromelain reduced OTA's concentration in 7.64% at pH 4.6; trypsin at 10.69% and the neutral metalloendopeptidase in 8.2%, 14.44%, 45.26% at pH 4.6, 5 and 7, respectively (p < 0.05). The less harmful α-ochratoxin was confirmed with trypsin and the metalloendopeptidase. This study is the first attempt to demonstrate that: (i) bromelain and trypsin can hydrolyse OTA in acidic pH conditions with low efficiency and (ii) the metalloendopeptidase was an effective OTA bio-detoxifier. This study confirmed α-ochratoxin as a final product of the enzymatic reactions in real-time practical information on OTA degradation rate, since in vitro experiments simulated the time that food spends in poultry intestines, as well as their natural pH and temperature conditions.