In Silico and In Vitro Studies of Mycotoxins and Their Cocktails; Their Toxicity and Its Mitigation by Silibinin Pre-Treatment

Abstract

Mycotoxins found in randomly selected commercial milk thistle dietary supplement were evaluated for their toxicity in silico and in vitro. Using in silico methods, the basic physicochemical, pharmacological, and toxicological properties of the mycotoxins were predicted using ACD/Percepta. The in vitro cytotoxicity of individual mycotoxins was determined in mouse macrophage (RAW 264.7), human hepatoblastoma (HepG2), and human embryonic kidney (HEK 293T) cells. In addition, we studied the bioavailability potential of mycotoxins and silibinin utilizing an in vitro transwell system with differentiated human colon adenocarcinoma cells (Caco-2) simulating mycotoxin transfer through the intestinal epithelial barrier. The IC50 values for individual mycotoxins in studied cells were in the biologically relevant ranges as follows: 3.57–13.37 nM (T-2 toxin), 5.07–47.44 nM (HT-2 toxin), 3.66–17.74 nM (diacetoxyscirpenol). Furthermore, no acute toxicity was obtained for deoxynivalenol, beauvericin, zearalenone, enniatinENN-A, enniatin-A1, enniatin-B, enniatin-B1, alternariol, alternariol-9-methyl ether, tentoxin, and mycophenolic acid up to the 50 nM concentration. The acute toxicity of these mycotoxins in binary combinations exhibited antagonistic effects in the combinations of T-2 with DON, ENN-A1, or ENN-B, while the rest showed synergistic or additive effects. Silibinin had a significant protective effect against both the cytotoxicity of three mycotoxins (T-2 toxin, HT-2 toxin, DAS) and genotoxicity of AME, AOH, DON, and ENNs on HEK 293T. The bioavailability results confirmed that AME, DAS, ENN-B, TEN, T-2, and silibinin are transported through the epithelial cell layer and further metabolized. The bioavailability of silibinin is very similar to mycotoxins poor penetration.

Mycotoxins: Biotransformation and Bioavailability Assessment Using Caco-2 Cell Monolayer

“…Microorganisms from guts have been reported to exhibit the capacity for degrading mycotoxins [ 131 , 132 , 133 , 134 ]. Additionally, P-glycoprotein (P-gp) and multidrug resistance protein (MRP), members of the ATP–binding cassette (ABC) superfamily of transport proteins, are able to pump mycotoxins out of the intestinal cells, leading to limit bioavailability of the substrates [ 71 , 135 ]. Both CYP450 and P-gp in the gut play a crucial role in defense mechanisms against mycotoxins that reach the intestinal mucosa [ 92 ].…”

“…Berger et al [ 215 ] showed that OTA was absorbed by the human intestinal mucosa by passive diffusion of the undissociated form of OTA and it was not appreciably metabolized by Caco-2 cells [ 215 ]. DON and NIV were not significantly metabolized or accumulated in Caco-2 cells as well [ 71 , 202 , 203 , 207 , 216 , 217 ]. Therefore, upon ingestion, these mycotoxins can be absorbed from the gut via intestine cells, then entered into the systemic circulation and thus transported to the whole body.…”

“…( A ): Demethylation; ( B , C , H ): Hydroxylation; ( D , I ): Methylation; ( E , F ): Sulfation, glycosylation, and glucuronidation; ( G ): Epoxide reduction. CYP: Cytochrome P; and UGTs: Uridine 5′-diphospho-glucuronosyltransferase [ 71 , 117 , 121 , 172 , 173 , 174 , 177 , 178 , 179 , 180 , 181 ].…”

“…Bioaccessibility can be considered as an indicator for the maximal absorption of the toxin, which can be used for realistic worst-case risk assessment of the toxin in a consumer product [ 70 ]. In fact, foodborne mycotoxins can be degraded or modified by metabolic processes of the human body, and only a fraction of the initial content can pass the intestinal membrane to enter the bloodstream [ 71 ]. In this sense, bioavailability is defined as the portion of ingested contaminant in food that reaches the systemic circulation [ 72 ].…”

Prevalent Human Gut Bacteria Hydrolyse and Metabolise Important Food-Derived Mycotoxins and Masked Mycotoxins

“…Similarly, de-acetylation of trichothecenes is considered a detoxification pathway for DAS [ 54 , 55 ] and DAS is de-acetylated by mixed human faecal microbiota [ 7 ]. In vitro studies also demonstrated that HT-2 is less cytotoxic than the T-2 [ 56 , 57 ]. The current study identified P. copri DSM 18205 as efficiently de-acetylating type A trichothecenes T-2 and DAS.…”