Cytotoxicity Assessment of Aflatoxin B1 (AFB1) after High Voltage Atmospheric Cold Plasma (HVACP) Generated Reactive Gas Species (RGS) Treatment Using Artemia salina and HepG2 Mammalian Cell MTT Bioassays

Abstract

Mycotoxins are toxic, heat-resistant metabolites of mold and fungi prevalent in food and animal feeds. Recent
studies have shown that air passed through high voltage atmospheric cold plasma (HVACP) becomes
temporarily ionized, forming reactive gas species (RGS) capable of destroying mycotoxins before reverting to
normal air. This study evaluated aflatoxin B1 (AFB1) cytotoxicity with and without RGS treatment using two
commonly applied toxicity evaluation model systems: Artemia salina [Brine Shrimp Test (BST)] and HepG23-(4-
5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide salt (HepG2-MTT) in vitro bioassays. The BST dose
response assay tested 0.25-1.13 μg AFB1/mL. Results obtained showed 11-73% mortality after 72h and 50%
mortality (LD50) at 0.6 μg AFB1/mL. Untreated AFB1 (1.7 μg/mL) caused 92% mortality of Artemia while RGStreated
AFB1 caused significantly less, 12% (p<.0001). The HepG2-MTT dose response assay tested 0.2-2.8
AFB1 μg/mL, causing 10-32% cell death and 13-57% for 24h and 48h respectively. Untreated AFB1 (0.8 μg/mL)
caused 22% and 29% cell death after 24h and 48h while RGS-treated AFB1 caused significantly less, 3% and 2%
(p<.05). Overall, this study confirms HVACP generated RGS significantly reduces AFB1 and the AFB1
byproducts formed are less toxic to cells and living organisms.