Interaction Of Aflatoxin B1 Research Articles

Current Knowledge of Individual and Combined Toxicities of Aflatoxin B1 and Fumonisin B1 In Vitro.

Mycotoxins are considered the most threating natural contaminants in food. Among these mycotoxins, aflatoxin B1 (AFB1) and fumonisin B1 (FB1) are the most prominent fungal metabolites that represent high food safety risks, due to their widespread co-occurrence in several food commodities, and their profound toxic effects on humans. Considering the ethical and more humane animal research, the 3Rs (replacement, reduction, and refinement) principle has been promoted in the last few years. Therefore, this review aims to summarize the research studies conducted up to date on the toxicological effects that AFB1 and FB1 can induce on human health, through the examination of a selected number of in vitro studies. Although the impact of both toxins, as well as their combination, were investigated in different cell lines, the majority of the work was carried out in hepatic cell lines, especially HepG2, owing to the contaminants' liver toxicity. In all the reviewed studies, AFB1 and FB1 could invoke, after short-term exposure, cell apoptosis, by inducing several pathways (oxidative stress, the mitochondrial pathway, ER stress, the Fas/FasL signaling pathway, and the TNF-α signal pathway). Among these pathways, mitochondria are the primary target of both toxins. The interaction of AFB1 and FB1, whether additive, synergistic, or antagonistic, depends to great extent on FB1/AFB1 ratio. However, it is generally manifested synergistically, via the induction of oxidative stress and mitochondria dysfunction, through the expression of the Bcl-2 family and p53 proteins. Therefore, AFB1 and FB1 mixture may enhance more in vitro toxic effects, and carry a higher significant risk factor, than the individual presence of each toxin.

Structural dynamics studies on the binding of aflatoxin B1 to chicken egg albumin using spectroscopic techniques and molecular docking

Aflatoxin B1, a mycotoxin produced by large number of Aspergillus species including Aspergillus flavus and Aspergillus parasiticus, has been described as the most potent carcinogenic mycotoxin. In this study, we have used a multiple spectroscopic and molecular docking approach to investigate the interaction of aflatoxin B1 (AFB1) with chicken egg albumin (CEA). Fluorescence spectroscopy, UV-Vis spectroscopy, and three-dimensional fluorescence spectroscopic techniques were employed to gain insight into the conformational changes in CEA in the presence of AFB1. Fluorescence spectroscopy revealed ligand-induced quenching in the fluorescence emission spectra of CEA upon binding with AFB1. Hyperchromic effect was observed in case of the ground state complex formation between CEA and AFB1 by UV-Vis spectroscopy. To gain further comprehension into the site of binding of AFB1 to CEA, competitive site marker displacement assay was performed using warfarin site marker. The magnitude of ΔG value calculated from fluorescence-based method was negative which confirmed spontaneous process. The results obtained suggest that the binding is enthalpy driven and van der Waals force and hydrogen bonds are stabilizing the AFB1-CEA complex. Three-dimensional fluorescence studies also confirmed the quenching in the fluorescence intensity around tryptophan residues in CEA. Circular dichroism assessment revealed reduction in the alpha helical content of CEA in the presence of AFB1. Molecular docking studies showed hydrophobic interaction, van der Waals forces, and hydrogen bonds as major forces present in interaction between CEA and AFB1. The overall study confirms conformational and structural alteration in the protein due to binding of AFB1.Communicated by Ramaswamy H. Sarma

Gut Microbiota Profiling of Aflatoxin B1-Induced Rats Treated with Lactobacillus casei Shirota.

Aflatoxin B1 (AFB1) is a ubiquitous carcinogenic food contaminant. Gut microbiota is of vital importance for the host’s health, regrettably, limited studies have reported the effects of xenobiotic toxins towards gut microbiota. Thus, the present study aims to investigate the interactions between AFB1 and the gut microbiota. Besides, an AFB1-binding microorganism, Lactobacillus casei Shirota (Lcs) was tested on its ability to ameliorate the changes on gut microbiota induced by AFB1. The fecal contents of three groups of rats included an untreated control group, an AFB1 group, as well as an Lcs + AFB1 group, were analyzed. Using the MiSeq platform, the PCR products of 16S rDNA gene extracted from the feces were subjected to next-generation sequencing. The alpha diversity index (Shannon) showed that the richness of communities increased significantly in the Lcs + AFB1 group compared to the control and AFB1 groups. Meanwhile, beta diversity indices demonstrated that AFB1 group significantly deviated from the control and Lcs + AFB1 groups. AFB1-exposed rats were especially high in Alloprevotella spp. abundance. Such alteration in the bacterial composition might give an insight on the interactions of AFB1 towards gut microbiota and how Lcs plays its role in detoxification of AFB1.

Molecular modeling studies on the interactions of aflatoxin B1 and its metabolites with the peripheral anionic site of human acetylcholinesterase

Aflatoxins are secondary metabolites of the fungi Aspergillus flavus and A. parasiticus. Among them, aflatoxin B1 (AFB1) is the most frequent type in nature and the most carcinogenic for mammals. It can contaminate many kinds of food like seeds, oil, olives, milk, dairy products, corn and meat, causing acute and chronic damages to the organism, especially in the liver, being, for this reason, considered highly hepatotoxic. AFB1 is also a mixed inhibitor of the enzyme acetylcholinesterase (AChE). This fact, together with its high toxicity and carcinogenicity, turns AFB1 into a potential chemical and biological warfare agent, as well as its metabolites. In order to investigate this, we performed inedited molecular modeling studies on the interactions of AFB1 and its metabolites inside the peripheral anionic site of human AChE (HssAChE), to verify their stability, suggest the preferential ways of inhibition, and compare their behavior to each other. Our results suggest that all metabolites can be better inhibitors of HssAChE than AFB1 and that AFBO and AFM1, the most toxic and carcinogenic metabolites of AFB1, are also the most effective HssAChE inhibitors among the AFB1 metabolites.Communicated by Ramaswamy H. Sarma

Molecular Modeling Studies on the Interactions of Aflatoxin B1 and Its Metabolites with Human Acetylcholinesterase. Part II: Interactions with the Catalytic Anionic Site (CAS).

The most common type of aflatoxin (AFT) found in nature is aflatoxin B1 (AFB1). This micotoxin is extremely hepatotoxic and carcinogenic to mammals, with acute and chronic effects. It is believed that this could be related to the capacity of AFB1 and its metabolites in inhibiting the enzyme acetylcholinesterase (AChE). In a previous work, we performed an inedited theoretical investigation on the binding modes of these molecules on the peripheral anionic site (PAS) of human AChE (HssAChE), revealing that the metabolites can also bind in the PAS in the same way as AFB1. Here, we investigated the binding modes of these compounds on the catalytic anionic site (CAS) of HssAChE to compare the affinity of the metabolites for both binding sites as well as verify which is the preferential one. Our results corroborated with experimental studies pointing to AFB1 and its metabolites as mixed-type inhibitors, and pointed to the residues relevant for the stabilization of these compounds on the CAS of HssAChE.

Interactions of aflatoxin B1 and related secondary metabolites with native cyclodextrins and their potential utilization

The attenuation or prevention of aflatoxins effect on agricultural products is still an unresolved problem due to the various influencing factors. Fluorescence fingerprint technique (such as excitation-emission matrix, EEM) has been developed for rapid detection of aflatoxins in liquid and solid samples. Unfortunately, the accuracy is seriously affected by background fluorescence compounds, such as kojic acid derivative (KAD). Therefore, development of more sensitive detection methods based on the enhancement of fluorescence signal is highly important. In this study, the EEM fluorescence technique was used to study the effects of α-, β-, and γ-cyclodextrins (CDs), and organic solvents on the fluorescence intensity of aflatoxin B1 (AFB1) and KAD, and molecular modeling study was also performed for the deeper understanding of these host-guest interactions. Results demonstrated that CDs and methanol could induce a higher fluorescence intensity of AFB1, while KAD presented a certain inhibitory effect on the enhancement of fluorescence signal of AFB1. Among the tested compounds β-CD formed the most stable conformation with AFB1 (the logarithm of binding constant values are approximately 2.22–2.49) and the clathrate compounds caused the highest fluorescence enhancement of AFB1 as well (the ratio of fluorescence intensity values (I/I0) for the presence and absence of CD at 430 nm are about 3.76–7.96). However, no obvious elevation for KAD fluorescence intensity caused by various CDs and organic solvents was observed at 493 nm. A significantly increase of fluorescence intensity was observed at the ratio of I430/I493 by CDs. This suggested that CDs could be used to decrease the interference of KAD, and to improve the accuracy and sensitivity for detection of AFB1, especially for the low concentration level.

Investigation of Non-Covalent Interactions of Aflatoxins (B1, B2, G1, G2, and M1) with Serum Albumin.

Aflatoxins are widely spread mycotoxins produced mainly by Aspergillus species. Consumption of aflatoxin-contaminated foods and drinks causes serious health risks for people worldwide. It is well-known that the reactive epoxide metabolite of aflatoxin B1 (AFB1) forms covalent adducts with serum albumin. However, non-covalent interactions of aflatoxins with human serum albumin (HSA) are poorly characterized. Thus, in this study the complex formation of aflatoxins was examined with HSA applying spectroscopic and molecular modelling studies. Our results demonstrate that aflatoxins form stable complexes with HSA as reflected by binding constants between 2.1 × 104 and 4.5 × 104 dm3/mol. A binding free energy value of −26.90 kJ mol−1 suggests a spontaneous binding process between AFB1 and HSA at room-temperature, while the positive entropy change of 55.1 JK−1 mol−1 indicates a partial decomposition of the solvation shells of the interacting molecules. Modeling studies and investigations with site markers suggest that Sudlow’s Site I of subdomain IIA is the high affinity binding site of aflatoxins on HSA. Interaction of AFB1 with bovine, porcine, and rat serum albumins was also investigated. Similar stabilities of the examined AFB1-albumin complexes were observed suggesting the low species differences of the albumin-binding of aflatoxins.

Gold coated porous silicon nanocomposite as a substrate for photoluminescence-based immunosensor suitable for the determination of Aflatoxin B1

A rapid and low cost photoluminescence (PL) immunosensor for the determination of low concentrations of Aflatoxin B1 (AFB1) has been developed. This immunosensor was based on porous silicon (PSi) covered by thin gold layer (Au) and modified by antibodies against AFB1 (anti-AFB1). PSi layer was formed on silicon substrate, then the surface of PSi was covered by 30nm layer of gold (PSi/Au) using electrochemical and chemical deposition methods and in such ways PSi/Au(El.) and PSi/Au(Chem.) structures were formed, respectively. In order to find PSi/Au the most efficiently suitable for PL-based sensor design, structure several different PSi/Au(El.) and PSi/Au(Chem.) structures were designed while using different conditions for electrochemical or chemical deposition of gold layer. It was shown that during the formation of PSi/Au structure crystalline Au nanoparticles uniformly coated the surface of the PSi pores. PL spectroscopy of PSi/Au nanocomposites was performed at room temperature and it showed a wide emission band centered at 700nm. Protein A was covalently immobilized on the surface of PSi/Au(El.) and PSi/Au(Chem.) forming PSi/Au(El.)/Protein-A and PSi/Au(Chem.)/Protein-A structures, respectively. In the next step PSi/Au(El.)/Protein-A and PSi/Au(Chem.)/Protein-A structures were modified by anti-AFB1 and in such way a structures (PSi/Au(El.)/Protein-A/anti-AFB1 and PSi/Au(Chem.)/Protein-A/anti-AFB1) sensitive towards AFB1 were designed. The PSi/Au(El.)/Protein-A/anti-AFB1- and PSi/Au(Chem.)/Protein-A/anti-AFB1-based immunosensors were tested in a wide range of AFB1 concentrations from 0.001 upon 100ng/ml. Interaction of AFB1 with PSi/Au(El.)/Protein-A/anti-AFB1- and PSi/Au(Chem.)/Protein-A/anti-AFB1-based structures resulted PL quenching. The highest sensitivity towards AFB1 was determined for PSi/Au(El.)/Protein-A/anti-AFB1-based immunosensor and it was in the range of 0.01–10ng/ml. The applicability of PSi/Au-based structures as new substrates suitable for PL-based immunosensors is discussed.

Interaction of aflatoxin B1 and fumonisin B1 in mice causes immunotoxicity and oxidative stress: Possible protective role using lactic acid bacteria

Aflatoxins (AF) are important foodborne mycotoxins implicated in human health and have immunocytotoxic effects. The aims of this study were to evaluate a new aflatoxin B1 (AFB1) and fumonisin B1 (FB1)-binding/degrading micro-organism for biological detoxification, to examine its ability to degrade AFB1 and FB1 in liquid medium, and to evaluate its potential in vivo protective role against any combined effects from AFB1 and FB1 on host splenocyte caspase-3 activity (reflecting DNA damage/cell death) and mRNA levels of select inflammation-regulating cytokines. Balb/c mice were divided into groups (10/group) and treated daily for 2 weeks by oral gavage with AFB1 (80 µg/kg BW), FB1 (100 µg/kg), AFB1 + FB1, or lactic acid bacteria (Lactobacillus paracasei BEJ01, 2 × 109 CFU/L, ∼2 mg/kg) – alone or in combination with the AFB1 and/or FB1. After the exposures, spleens were collected for measures of caspase-3 activity, lipid peroxidation (LP), and glutathione (GSH) content, expression of anti-oxidation protective enzymes (GPx and SOD), and mRNA levels of inflammation-regulating cytokines (e.g. IL-10, IL-4, IFNγ, TNFα). Thymii were also removed for analysis of apoptosis. The results indicated that, in the spleen, exposure to the mycotoxins led to increased caspase-3 activity, LP, and IL-10 and IL-4 mRNA levels, but decreased GSH content and down-regulated expression of GPx and SOD, and of IFNγ and TNFα mRNA. Co-treatment using Lactic Acid Bacteria (LAB) with AFB1 or FB1 suppressed levels of DNA fragmentation, normalized splenic LP and increased GSH levels, up-regulated expression of GPx and SOD, and normalized mRNA levels of the analyzed cytokines. It is concluded that AFB1 and FB1 might have combinational (synergistic moreso than additive) toxic effects in situ. Further, it can be seen that use of LAB induced protective effects against the oxidative stress and (immuno)toxicity of these agents in part through adhesion (and so likely diminished bioavalability).

Interactions of aflatoxin B1 with SRP components can disrupt protein targeting.

Spectrofluorimetric studies have revealed that aflatoxin B1 (AFB1) interacts with signal recognition particle (SRP), which acts as an escort for polyribosomes with signal peptides to be transported and bound to the cytoplasmic face of the endoplasmic reticulum (ER). We further report that the binding of AFB1 to SRP is selective as it only binds to two (SRP9 and 14) out of its three constituent polypeptides studied. Binding of AFB1 to proteins is known to alter their conformations. Interactions of AFB1 with SRP polypeptides may generate structural and functional alterations in this particle and hinder secretory protein synthesis.

Molecular mechanics calculations of the noncovalent interaction of aflatoxin B1 and its ultimate carcinogen with various dna sequences

Molecular mechanics calculations have been used to study the external noncovalent complexes formed between DNA and the strong carcinogen, aflatoxin B1. Three different sequences of hexameric duplexes were used for the DNA. Both the aflatoxin B1 parent molecule and its ultimate carcinogenic form, a carbocation, were modeled. The results are compared with recent experimental data on sequence specificity of the covalent attachment of aflatoxin to sites on the nucleotide bases. The comparison is discussed in light of a recently proposed hypothesis suggesting that the locus of carbocation formation is in acidic domains near the surface of the macroion and this determines the site of covalent adduct formation.

Interactions of aflatoxin B1 and alkylating agents with DNA: structural and functional studies.

Interactions of aflatoxin B1 and alkylating agents with DNA: structural and functional studies.

Interaction of aflatoxin B1 with some biologically important substances.

The interaction of aflatoxin B1 with a number of biologically important substances has been investigated by spectrophotometric and fluorimetric techniques. The ultra-violet absorption of tryptophan, adenosine, RNA and DNA were altered by the addition of aflatoxin B1. Some amino-acids, purine and pyrimidine derivatives, bovine-serum albumin and DNA quenched the fluorescence of aflatoxin B1. Fluorescence polarization data suggested that the interaction of aflatoxin B1 with bovine-serum albumin was stronger than that with DNA.

Interaction of Aflatoxin B<sub>1</sub> with DNA

The interaction of aflatoxin B1 with DNA was investigated. In the presence of native DNA the absorption spectrum of the toxin showed an obvious spectral shift in the region of 300-420 nm with an isosbestic point at 376 nm. DNA-bound aflatoxin is hydrolyzed in alkaline media more easily than the free toxin. The hydrolized form has a labile structure and can decompose further. The bound toxins are easily dissociated by heat as well as by salt, and all toxin molecules are released from DNA which remains double-stranded. Aflatoxin can bind to thermally denatured DNA as well, with an accompanying spectral shift which depends on the particular preparation of denatured DNA, and there was an isosbestic point at 380 nm. The complex of toxin and denatured DNA was stabilized by salt up to 0.1 M. Thus it was concluded that aflatoxin was bound with denatured DNA in a different form from native DNA. The number of binding sites of DNA was estimated by constructing Scatchard plots based on both spectral analysis and equilibrium dialysis. However, these show no definite value but fall in the region of 0.07 to 0.013, that is one toxin molecule per 80-140 nucleotide of native DNA. Several lines of evidence suggest the possibility that aflatoxin exists in aqueous solution as aggregates. The mechanism of the binding was discussed. It is noteworthy that the number of binding sites of DNA doubles by the presence of histones.

The Interactions of Aflatoxin B1 and 4-Hydroxycoumarin with Human Serum Albumin

The Interactions of Aflatoxin B1 and 4-Hydroxycoumarin with Human Serum Albumin

The interaction of aflatoxin B1 with polynucleotides and its effect on ribonucleic acid polymerase.

1. The interaction of aflatoxin B(1) with different polynucleotides was studied spectrophotometrically. Equations were derived that enable the degree of binding to be determined without first determining the extinction coefficient of the bound form. 2. The interaction with calf thymus DNA obeys first-order relationships with an association constant of 0.40mm(-1), but there is some evidence for a secondary binding process from results obtained at 390nm. 3. The spectral shifts decreased in the order polyadenylic acid+polyuridylic acid>DNA>polyadenylic acid>polyadenylic acid+polyinosinic acid. Polycytidylic acid, polyuridylic acid, polyinosinic acid (both single- and triple-stranded), AMP, CMP, GMP and UMP did not interact with aflatoxin. It was concluded that there is a requirement for the amino group of adenine (or possibly guanine) for binding of aflatoxin to polynucleotides to occur. 4. Binding is reversed by increasing ionic strength, and by Mn(2+) and Mg(2+) in the concentration range studied (0-5mm). The effect of the Mn(2+) or Mg(2+) was far greater than would be expected on the basis of their ionic strength. With both the bivalent cations and sodium chloride the reversal is greatest with double-stranded polynucleotides. 5. Inhibition in vitro of the DNA-dependent RNA polymerase of Escherichia coli by aflatoxin B(1) was detected only in the absence of Mg(2+) and at concentrations of Mn(2+) below the optimum for RNA synthesis in vitro. 6. The degree of inhibition (maximally 30%) was dependent on the concentration of Mn(2+) and decreased during incubation.

Interactions of aflatoxin with histones and DNA.

The interactions of aflatoxin B(1) with certain histone fractions and DNA were investigated by means of viscosity measurements and equilibrium dialysis. Two main histone fractions (F2b and F1), both lysine-rich, were examined after treatment with the toxin. Fraction 2b and 1 differ in amino acid composition and behave differently, in regard to gross conformation, in the presence of electrolytes. Aflatoxin increased the viscosity of fraction 2b but affected the viscosity of fraction 1 only slightly. Equilibrium dialysis experiments showed that aflatoxin was bound to both histone fractions. Aflatoxin also increased the viscosity of DNA and equilibrium dialysis showed that 1 molecule of the toxin was bound to approximately every 5 nucleotides of the nucleic acid. Binding constants for the aflatoxin complexes were calculated as 1000 for F2b, 700 for F1, and 5500 for DNA. The biological implications of these data, in regard to the effect aflatoxin has on the information-transcription process, are discussed.