*1.2.2 Chemical structure*

Chemically aflatoxin B occur they are difuro-coumorins –cyclopentenone and difurocoumaro lactone series which are freely soluble in chloroform and methanol [49, 50], Other aflatoxins have different substitutes but share basic coumarine structure. The epoxidation of the 8, 9-double bond and cyclopentenone ring of B series is responsible for the order of acute and chronic toxicity as compared with the six-membered lactone ring of the G series AFB1 > AFG1 > AFB2 > AFG2 (**Figure 1**) [49].

## *1.2.3 Physical structure*

Structurally they are dihydrofuran-coumorins moiety containing double bond which are freely soluble in chloroform and methanol. They are stable at high temperatures but unstable to UV light or polar solvents [49, 51]. Aflatoxins are toxic secondary metabolites upon exposure to fluorescence ultra violet (UV) light, aflatoxin B appear blue in color and G appear green in color (**Table 1**) [49, 52].

#### **Figure 1.**

*Chemical properties of aflatoxin B and G (A–F). Source: Adapted from Agriopoulou et al. [38].*


*Adapted from: Reddy et al. [49]*

#### **Table 1.**

*Physical properties of aflatoxins.*

#### **1.3 Occurrence of aflatoxin in food and feed**

Eggs, milk and meat are sometimes contain residues of aflatoxins because of consumption of aflatoxin contaminated feed ingredients such as peanuts, cottonseed, nuts, almonds, figs, spices, soybean, rice and maize [53].

#### **1.4 Mode of action**

Cytochrome P450 enzymes (phase I metabolisation) convert aflatoxins to a reactive 8,9-epoxide form, which is essential for the toxicity. In mammals CYP1A2 and CYP3A4 are the enzymes responsible for conversion [54] in chicken and turkeys, the corresponding enzymes are CYP2A6 and to a lesser extent CYP1A1 orthologs [55, 56]. DNA and protein binds to guanine residues of nucleic acids to produced epoxide metabolite causing genotoxicity and cytotoxicity [57]. Aflatoxin B1-DNA adducts result in guaninecytosine (GC) to thymine-adenine (TA) transversions [48], which leads to irreversible DNA damage, therefore results to hepatocellular carcinomas [58]. Gluthatione conjugation or hydrolysis detoxified the toxic epoxide metabolite and epoxide hydrolase to phase II metabolisation and AFB1–8,9-dihydrodiol (AFB1-dhd) respectively. AFBI Metabolisation to less toxic compounds such as aflatoxin M1 (AFM1) or Q1 (AFQ1) [54, 56]. AFM1 metabolite possesses carcinogenic properties which are 10 times lower than AFB1. These metabolites obtained from cattle milk. The maximum limits in milk permissible for human consumption have been established (0.05 μg/kg) [12, 59], 20 ppb in grain and 4 ppb in food and agricultural commodities [59].

#### *1.4.1 Carcinogenesis*

The International Agency for Research on Cancer [60] classify aflatoxin as class 1 carcinogen, transversion of G to T occur in guanine codon 249 of tumor suppressor gene p53 of DNA that induce mutagenesis by alkylation of nuclear DNA, leading to carcinogenesis and teratogenesis [61]. 8, 9,-epoxide is a potent carcinogen and induces chromosomal aberrations, mutation and cell toxicity [62].

#### *1.4.2 Immunesuppression*

Immunosuppressive effects on NK cell activity, humoral and cellular immune function are impair by aflatoxin through reducing the primary and secondary

immune responses [63–66]. AFB1 induces; thymic aplasia, reduce T-lymphocyte function, lymphokines, suppress phagocytic and complement activity [67, 68]. Aflatoxin suppresses the levels of IL-1, IL-2, IL-6, IFN, TNF alpha, mRNA and proinflammatory cytokines [69, 70]. Embryonic chicks exposed to AFB1 showed a depressed graft-versus-host response, thymic bursal involution, delayed cutaneous hypersensitivity, macrophages function, reduced antibody titers to vaccines for Newcastle, Mareks and infectious bursal disease [32, 52, 71, 72].
