**3. Types of aflatoxins, structure and properties of AFB1**

To date twenty (20) different types of AFBs have been discovered. Of these 20 currently known AFBs, the major ones include AFB1, AFB2, AFG1, AFG2, aflatoxin M1 (AFM1) and aflatoxin M2 (AFM2). The AFB1, AFB2, AFG1 and AFG2 are made by fungi while AFM1 and AFM2 are made as intermediate products when AFB1 and AFB2 respectively are metabolized. The B and G descriptions of AFBs refer to the type of color generated when AFBs are placed under short-wave UV (ultraviolet) illumination during thin layer chromatography. The B description of AFB1 and AFB2 refers to the blue light generated under UV illumination while the G description refers to the green light generated under UV illumination on thin layer chromatographic plates. The AFM1 and AFM2 were initially detected in raw milk of livestock that had ingested feed contaminated with AFBs thus the description M. The subscript numbers 1 and 2 indicate the major and minor compounds respectively [18–20].

Structurally, AFB1 like all other AFBs consist of bifuran ring that is fused to a coumarins ring. AFB molecules differ from AFG molecules in that AFB molecules have cyclopentenone ring while the AFG molecules have lactone ring. There are double bonds at loci 8 and 9 on the terminal furan ring of the AFB1 structure and these double bonds confer the unique carcinogenic characteristics to AFB1 (**Figure 1**) [21].

Generally, AFBs occur as crystals which appear as uncolored or lemon-yellow at 25 to 28°C [22]. AFBs are partially soluble in water and hydrocarbons but cannot be dissolved in hydrophobic solvents. AFBs are completely dissolvable in polar solvent like alcohol (e.g. methanol), acetone and chloroform. To degrade AFBs, they can be placed in light and air. AFBs can also be degraded by exposing them to UV light,

**Figure 1.** *Chemical structure of AFB1.*

strong acid solution, strong basic solution and oxidizing agents. AFBs can be broken down by exposing them to very high temperature conditions ranging between 237 to 299°C. Complete destruction of AFBs can be achieved when they are autoclaved with ammonia or when they are treated with bleach containing sodium hypochlorite. Normal cooking temperatures cannot degrade AFBs.

### **4. Biotransformation of aflatoxins**

Metabolism of AFB1 largely occurs in the liver by a group of enzymes called cytochrome P450 (CYP 450). When AFB1 is ingested, it is transported to the liver where the CYP 450 enzymes convert AFB1into different compounds which include AFM1, aflatoxicol, aflatoxin P1 (AFP1) and aflatoxin Q1 (AFQ1). Additionally, the CYP450 enzymes especially CYP1A2 and CYP3A4 convert AFB1 into reactive oxygen species (ROS) AB1–8, 9-epoxide which exists in two (2) forms; endo-AFB1–8, 9-epoxide and exo-AB1–8, 9-epoxide. Whereas the CYP3A4 produces exo-AB1–8, 9-epoxide and a small quantity of AFQ1, the CYP1A2 produces both endo and exo-AFB1–8, 9-epoxides as well as AFM1 [23]. Of the two epoxide species, exo-AFB1–8, 9-epoxide is considered to be the toxic species that confers genotoxic characteristics on AFB1 [2, 3, 24]. The AFB1–8, 9 epoxide metabolites formed can form conjugates with glutathione leading to the formation of a stable, harmless, soluble product which is excreted in the bile. The conjugation process is catalyzed by the enzyme glutathione-S-transferase (GST). The conjugation and the subsequent excretion of the soluble product formed is the mechanism by which AFB1 is detoxified as a hepatocarcinogen. The AFB1–8, 9 epoxideglutathione complex formed is also broken down in the liver and kidney is excreted in the urine as mercapturic acid [25]. On the other hand, when individuals are exposed to high levels of AFB1 beyond the capability of GST enzymes to break down the epoxides into harmless forms, or when the activity of GST enzymes is reduced through

**Figure 2.**

*Schematic flow chart on metabolism of AFB1.*

mutations of the GST gene, the AFB1–8, 9 epoxides can bind to liver proteins and cause their failure which can result in acute hepatotoxicity or aflatoxicosis.

Conversely, the 8, 9 epoxides can cause mutations of DNA in the liver cells and as a results pro-mutagenic lesions may be formed. When the pro-mutagenic lesions are formed, proto-oncogenes are activated and this may cause the tumor suppressor genes to become inactivated. The AFB1–8, 9 epoxide has affinity for the N7 atom of guanine and so bind with it which leads to the production of a pro-mutagenic DNA adduct (AFB1-N7 -Gua adduct). The AFB1-N7 -Gua adduct is not stable and so goes through depurination process which results in the adduct being excreted in the urine. Animals such as mice that are more immune to the carcinogenic effects of AFB1 have much greater GST action compared to animals such as rats which more susceptible to the carcinogenic effects of AFB1. In humans, the action of GST enzymes is much lower when compared with rats and mice. This suggests that the ability of humans to detoxify AFB1–8, 9 epoxides is lower [26] and therefore humans stand a greater chance of suffering from the carcinogenic effects of AFB1 when compared with rats and mice. **Figure 2** below shows the schematic flow chart on how metabolism of AFB1 occurs in the liver.

### **5. Effects of aflatoxins on human health**

AFBs are very genotoxic agents that can still cause ailment in human beings when individuals are exposed to small quantities [27]. Even though AFBs can cause disease in many parts of the human system, they are largely known to cause acute and or chronic disease in the liver as well as liver cancer. There are so many ways by which AFBs manifest their toxic side effects when ingested into the body. AFBs can modify the integrity of the intestines [28] and regulate the expression of cytokines. These negative effects of AFBs can lead to impaired growth and weakening of the immune system in children [29]. The quantity or amount of AFBs consumed or ingested coupled with how long the individual has been exposed largely determine the negative impact of AFBs in humans and other animals. Acute exposure of humans to AFBs occur when large amount of AFBs are consumed within shortest possible time. Chronic exposure occurs when humans consume minute quantities of AFBs over a prolonged period of time.

When humans are exposed to large quantities of AFBs over a relatively short time period, it can results in vomiting, stomach aches, mental retardation, improper digestion of food, liver disease, coma and hepatotoxicity or aflatoxicosis. About 25% of individuals who experience acute AFBs exposure die from AFBs-related diseases [30]. There are environmental factors which predispose humans to acute aflatoxicosis. These factors are scarcity of food, high temperatures and humid environment which promotes the growth of the fungi that produce AFBs and inadequate systems to regulate and monitor food stuffs for the presence AFBs. Globally acute aflatoxicosis has become a recurrent public health problem [31, 32].

Research indicates that prolong exposure of individuals to small quantities of AFBs can cause impairment of the immune system, reduction in absorption of nutrients from the small intestines which may ultimately result in stunted growth especially in children and young infants [33, 34]. Reports from research conducted within Togo as well as Benin, countries located in the West African sub-region indicate that there is a correlation between levels of AFB-protein (albumin) adduct and growth impairment [35, 36]. In 2005 Jiang et al. [37] undertook a study in Ghana and reported that individuals who had higher levels of AFB1-albumin adducts had reduced levels of certain leukocytes types. Similarly, Turner et al. [38] undertook a study in Gambia and reported that infants who had high levels of aflatoxin-albumin adducts had lower quantities of IgA antibodies in their saliva.
