Pub Date : 2021-08-27DOI: 10.21203/rs.3.rs-846230/v1
Natalia Klanovicz, F. Stefanski, A. F. Camargo, W. Michelon, H. Treichel, A. C. Teixeira
The occurrence of micropollutants in aqueous matrices has become a global concern and a challenge for wastewater treatment plants. Monitoring their toxicity has shown that these compounds, even at low concentrations, pose risks to human and environmental health. Therefore, our study aimed to prospect immobilization strategies for non-commercial oxidoreductase enzymes and insert them in the context of pollutant remediation. The enzymatic extract was obtained by submerged fermentation of the fungus Trichoderma koningiopsis in an alternative substrate, consisting of fresh microalgal biomass from the phycoremediation process. The immobilization efficiency of peroxidase (POD) was evaluated by monitoring the residual activity (RA) and the discoloration potential (DP) of a synthetic dye solution. Concomitantly, the catalytic properties of free POD were explored, and the most promising storage strategy to maintain the enzymatic activity was studied. Guaiacol peroxidase from T. koningiopsis expressed specific activity of up to 7801.1 U mg− 1 in the free form, showing stability when subjected to up 80°C in a pH range between 4.0–8.0. Furthermore, the bioproduct immobilized on magnetic nanoparticles expressed up to 688% RA and 100% DP. An increase in the RA of the enzyme, both in free and immobilized form, was also observe after storage for up to 8 months. The synthesized magnetic nanozymes showed good reusability, maintaining 13546.4 U mg− 1 after ten cycles and removing 93% of color in a second batch. Toxicological evaluation with Allium cepa indicated that the enzymatic process of color removal with immobilized POD, despite maintaining unwanted cytotoxic effects, was essential to eliminate genotoxic effects. In this sense, the immobilization processes of T. koningiopsis peroxidase presented in our work are promising for the enzyme market and for the wastewater treatment sector.
水基中微量污染物的存在已成为全球关注的问题，也是污水处理厂面临的挑战。对其毒性的监测表明，这些化合物即使浓度很低，也会对人类和环境健康构成风险。因此，本研究旨在探索非商业氧化还原酶的固定化策略，并将其应用于污染物修复。该酶提取物是通过在由藻修复过程中产生的新鲜微藻生物量组成的替代底物中对真菌克宁质木霉进行深层发酵获得的。通过监测合成染料溶液的残留活性(RA)和变色势(DP)来评价过氧化物酶(POD)的固定化效率。同时，对游离POD的催化性能进行了探讨，并对最有希望保持酶活性的储存策略进行了研究。在游离状态下，愈创木酚过氧化物酶的比活性高达7801.1 U mg−1，在高达80°C、pH值在4.0-8.0之间的条件下表现出稳定性。此外，磁性纳米颗粒固定化的生物产物表达率高达688% RA和100% DP。在储存8个月后，游离和固定形式的酶的RA也有所增加。所合成的磁性纳米酶具有良好的可重复使用性，10次循环后仍保持13546.4 U mg−1，第二批脱色率达93%。对葱的毒理学评价表明，固定化POD除色的酶促过程，尽管保持了不必要的细胞毒性作用，但对于消除遗传毒性作用是必不可少的。从这个意义上说，我们的工作中提出的koningiopsis过氧化物酶的固定化工艺对酶市场和废水处理领域有很大的前景。
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