# A Review: Sample Preparation and Chromatographic Technologies for Detection of Aflatoxins in Foods
## 1 Introduction  
Aflatoxins are a class of toxic metabolites generated by certain fungi that are commonly found in the environment. _Aspergillus flavus_ and _Aspergillus parasiticus_ are the dominant fungal species that produce aflat under drought, warm and humid conditions [1; 2; 3]. In the 1960s, Turkey \"X\" disease resulted in the death of over 100,000 turkeys and other poultry in England [4; 5; 6; 7]. The outbreak, associated with contaminated feed, led to the identification of aflat and commenced research on aflatoxin toxicity. Since then aflatoxin contamination has been gradually recognized as a threat to global food safety and trade [8; 9; 10; 11]. Continuous and collaborative efforts from government, academia and industry have been dedicated to understanding and mitigating the impact of aflat on public health and economy [12; 13; 14]. Aflatoxins are well known to contaminate crops in the field and during storage [15]. Commonly consumed food commodities such as cereals, peanuts and dairy products are a major concern for aflatoxin contamination because current agricultural practices and food processing techniques cannot completely remove aflat [1; 16; 17]. Consequently, aflat can enter the food chain and give rise to health and trade related issues [18; 19; 20]. Among various types of aflat, the predominant ones--aflatoxin B\({}_{1}\), B\({}_{2}\), G\({}_{1}\) and G\({}_{2}\) and aflatoxin M\({}_{1}\), a hydroxylated metabolite of aflatoxin B\({}_{1}\)--can pose a severe threat to human and animal health via dietary exposure. Toxicity of aflat is well-documented. As reported by the International Agency for Research on Cancer (IARC), aflat are carcinogenic and hepatotoxic to humans. Aflatoxin B\({}_{1}\), B\({}_{2}\), G\({}_{1}\) and G\({}_{2}\) are classified as human carcinogens (group 1B). Aflatoxin B\({}_{1}\) is the most potent carcinogen within the group. Although aflatoxin M\({}_{1}\) is less hepatotoxic and immunotoxic than its parent compound, aflatoxin B\({}_{1}\), it is stable under pasteurization temperature used to process milk. It is classified as a possible human carcinogen (group 2B) by the IARC. [21]. In response, regulatory bodies worldwide have set regulatory levels for aflat, especially for aflatoxin B\({}_{1}\) and aflatoxin M\({}_{1}\), in various commodities [22]. For example, in peanuts, regulatory levels range from 2 ug/kg for aflatoxin B\({}_{1}\) and 4 ug/kg for total aflat (sum of aflatoxin B\({}_{1}\) + B\({}_{2}\) + G\({}_{1}\) + G\({}_{2}\)) by the European Union [23] to 20 ug/kg for total aflat by the United States Food and Drug Administration [24]. Similarly, the Food Safety and Standards Authority of India [25] has set the limit of total aflat at 10 ug/kg for ready-to-eat nuts and 15 ug/kg for processed nuts. In the US, the action level of aflatoxin M\({}_{1}\) in raw milk is 0.5 ug/kg, while in the EU the maximum level of aflatoxin M\({}_{1}\) in milk is 0.05 ug/kg and 0.025 ug/kg in infant formulas and dietary foods for special medical purposes intended specifically for infants. Large variability among these regulatory levels exists because of the requirements (feasibility and economic impact) and approaches to risk analysis are defined in national/regional legislation [26]. To comply with established regulatory levels and ensure food and feed are safe for consumption and trade, development of analytical methods that are suitable for monitoring and surveillance of aflat are critical. A well-established aflatoxin method is based on a thorough understanding of the target analytes (polarity, water solubility, thermostability, partitioning coefficient, pKa, etc. ), food composition (e.g., fat, water and sugar content), extraction conditions, sample clean-up and instrumentation. Figure 1 shows the important physicochemical properties of aflat B\({}_{1}\), B\({}_{2}\), G\({}_{1}\), G\({}_{2}\) and M\({}_{1}\). Figure 1: Important physicochemical properties of aflatoxin B\({}_{1}\), B\({}_{2}\), G\({}_{1}\), G\({}_{2}\) and M\({}_{1}\).