Chitosan-Based Nanoencapsulation of Ocimum americanum Essential Oil as Safe Green Preservative Against Fungi Infesting Stored Millets, Aflatoxin B1 Contamination, and Lipid Peroxidation

Present study deals with the first-time report on encapsulation of Ocimum americanum essential oil (OAEO) into chitosan matrix with enhanced antifungal, aflatoxin B-1 (AFB(1)) inhibition, antioxidant activity, and in situ efficacy in the millet food system. GC-MS analysis suggested citral (66.72%) as the major component of OAEO. Physicochemical characterizations through SEM, FTIR, and XRD analyses confirmed the successful loading of OAEO into chitosan nanoemulsion (OAEO-CsNe). In vitro release profile of nanoencapsulated OAEO exhibited biphasic burst and controlled volatilisation, a prerequisite for long-term antifungal effect in the stored food system. OAEO-CsNe completely inhibited the growth and AFB(1) production of Aspergillus flavus at 0.2 and 0.175 mu L/mL, respectively. Inhibition of ergosterol biosynthesis followed by the release of vital cellular ions, and 260, 280 nm absorbing materials from AFLHPSi-1 cells suggested plasma membrane as a potential site of antifungal action of OAEO-CsNe. Significant reduction of cellular methylglyoxal (an AFB(1) inducer) level in AFLHPSi-1 cells after fumigation with OAEO-CsNe confirmed the novel biochemical mechanism of anti-aflatoxigenic activity. Additionally, in silico modelling of citral (major component of OAEO) with Ver-1 and Omt-A proteins suggested the hydrogen bond-dependent molecular interaction for inhibition of AFB(1) biosynthesis. OAEO-CsNe showed significant in situ antifungal, anti-aflatoxigenic, and lipid peroxidation-suppressing potentialities without altering the organoleptic and germination properties of Setaria italica seeds. Moreover, the appreciative safety profile (LD50 = 11,162.06 mu L/kg) of OAEO-CsNe in a mammalian model system (Mus musculus) strengthens its recommendation as an effective green preservative against fungal infestation, AFB(1) contamination, and reactive oxygen species-mediated lipid peroxidation in stored food commodities.