A Review of Ochratoxin A Occurrence, Condition for the Formation and Analytical Methods

Ochratoxin A (OTA) is a mycotoxin produced by several fungal species including Aspergillus ochraceus, A. carbonarius, A. niger and Penicillium verrucosum. Various studies report shown that Ochratoxin A can be leads several health problems for both animal and human health through the consumption of Ochratoxin A contaminated plant and animal origin foods. For instance, Ochratoxin A has been shown to be nephrotoxic, teratogenic, immunotoxic, and carcinogenic in human health. Therefore, the main aim of this review focused on the occurrence, analytical methods and the condition for the formation of Ochratoxin A. Number of studies fi nding report indicate that Ochratoxin A was existed in several processed and unprocessed food stuffs, species and different alcoholic beverage. Primarily, cereals and cereals contained food products have highly vulnerable for Ochratoxin A due the presence of high moisture contents. On the other hand, several environmental conditions are playing an important for the formation of Ochratoxin A in different food stuffs. For example, most important abiotic factors which infl uence the growth and Ochratoxin A production by such Spoilage fungi include water availability, temperature and gas composition. Finally, several analytical methods are used for detection of Ochratoxin A from different plant and animal origin foods such as Thin layer chromatography, Enzyme linked immunosorbent assay and High performance liquid chromatography. However, based on the sensitivity, resolution and effi ciency currently high performance liquid chromatography techniques are more popular and advanced analytical techniques for different mycotoxins groups particularly, Ochratoxin A and Afl atoxins detection. Review Article A Review of Ochratoxin A Occurrence, Condition for the Formation and Analytical Methods


Introduction
Mycotoxins are secondary metabolites produced by a wide variety of fi lamentous fungi, including species from the genera Aspergillus, Fusarium, Penicillium, Alter aria and Clavicles that grow under different climatic conditions on agricultural commodities [1]. The Food and Agricultural Organization (FAO) of the United Nations has estimated that up to one quarter (25%) of the world's food crops are signifi cantly contaminated with secondary metabolites of different toxigenic moulds (FAO 2004). Ochratoxins belong to a family of structurally related, secondary fungal metabolites produced by various Penicillium and Aspergillus strains [2]. Based on their structure and chemical compounds Ochratoxin can be categorized into three. Among them, Ochratoxin A (OTA) is the most commonly occurring metabolite and the most hazardous, since it is a nephrotoxic and carcinogenic agent [2]. Additionally, exposure of the human being for a long period of time with Ochratoxin A contaminated diets it leads several health problems such as, kidney, liver cancer and weakening of the immunity system [3], and has been classified by the International Agency for Research on Cancer (IARC) as a possible carcinogen to humans [4]. Likewise, Ochratoxin A it can be leads serious health problem to animal during the feeds of contamination agricultural products by these mycotoxin types. For example, hepatotoxic, carcinogenic, genotoxic, Immunotoxic, teratogenic, and neurotoxic health impacts on animals [5]. Furthermore, Ochratoxin A is toxic metabolites of Aspergillus fungi that can contaminate various foods and feed products.
One of the recent research fi nding result shows that the presence of Ochratoxin A in wheat bread and maize bread is (12.9 and 70)μg/kg respectively [6]. And the result showed that the concentration level of maize breads has high Ochratoxin A contents and maize and maize contain food stuffs is a worldwide problem of Ochratoxin A. Especially, maize kernels are a good substrate for mould infection and production of mycotoxins harmful to both humans and animals. However, these products (maize) play an important role in developing countries particularly the diets of Ethiopian people. The infection of these crops by Ochratoxin A fungi and hence contamination with Ochratoxin A is generally higher [7]. On the other hand, majority of Ethiopian people depend their own life by agricultural product especially cereal products occupy the primary position. These indicate that the damage caused by Ochratoxin A in Ethiopia is increasing this may be due to lack of well documented information, poor management and lack of awareness on occurrence Ochratoxin A from agricultural product. Consumption of Ochratoxin A in higher amount through feed and foodstuffs may impact animal and human health. The effects of Ochratoxin A not only human and animal health impact but also play a signifi cant role for economic loss and boundary less distribution. Therefore this review aims to address the occurrence, condition for the formation and analytical methods for Ochratoxin A. In addition, they are benefi cial to the society, scientifi c community and research institution.  Figure 1 [9]. Among them Ochratoxin A is the most toxic member of the Ochratoxin which is structurally similar to the amino acid phenylalanine. Chemical name for Ochratoxin A contains 7-carboxy-5-chloro-8-hydroxy-4,4-dihydro-3(R)methyl isocoumarin (Ochratoxin ; OT) that is linked through the 7-carboxy group to l--phenylalanine by a peptide bond.

Material and methods
Briefl y, the chemical abstract number, molecular formula and molecular mass of each Ochratoxins type is described in

Occurrence of Ochratoxin A in food and feeds
Ochratoxin A is one of the secondary metabolite of mycotoxin group and can be existed in several agricultural products and in animal feeds [10]. Number of studies fi nding report indicate that Ochratoxin A was existed in several processed and unprocessed food stuffs, species and different alcoholic beverage. For instance, in cereal, milk, meat and species [12][13][14][15] in alcoholic beverage such as beer and wine [16,17]. Likewise, some dried fruit and coffee was infected by Ochratoxin A in [17,18]. But cereals occupy the first position of the total exposure to Ochratoxin A with 60% because of cereal have high moisture contents some times more than 20%  [19]. Hence, to protect consumer from different health risks different International Organization was set the maximum permissible limits for Ochratoxin A in different food stuffs and alcoholic beverage. Then, the maximum limits of 5.0 ng/g Ochratoxin A in raw cereal grains whereas 3.0 ng/g in cerealprocessed. Similarly, coffee and dried fruit 10 ng/g wines and cereal based baby food 2 μg/L and 0.5 ng/g respectively set by European commission [20]. Likewise, Ochratoxin A has also been determined in foods of animal origin such as pork blood products, pork kidney, pork liver, or pork meat. The reason the pigs are predominantly exposed to Ochratoxin A through their contaminated feed [18]. In short, Ochratoxin A is a secondary metabolite produced either by penicillin in cereal and cereal proceeds food or Aspergilli in wine, grapes, coffee and cocoa [21]. According to this in recent time several scientifi c communities has received increased primary attention towards the effects of Ochratoxin A because of its hazard to human and animal health [22]. In general, animals are directly exposed to mycotoxins through the consumption of mould feedstuff

Effects of Ochratoxin A on animal health
Similar to that of human being, animals are exposed to Ochratoxin A causes when the feeds have been known to exhibit the form of intoxication which can leads to death through consumptions [30]. Once the animals are exposed to Ochratoxin

Condition formation of Ochratoxin A in different commodities
The most important abiotic factors which infl uence the growth and Ochratoxin A production by such Spoilage fungi include water availability, temperature and gas composition [34]. Water activity is perhaps the most critical factor infl uences the germination, growth and establishment of molds on nutrient worthy substrates. The previous studies shown, that Ochratoxin A contamination was produced by P. verrucosum at water activity of 0.95 and a temperature of 25 0 C [35]. Similar to water activity, temperature is also second factor and highly Several literature data indicate that Penicillium verrucosum is found mostly in grains with moisture contents are higher than about 14.5% [41]. In addition, the development P. verrucosum and formation of Ochratoxin A on different grain (wheat) when the moisture content ranging from 10-30% and 18-22% [42].
However, when the moisture content was detected below 17% the probability for the growth of P. verrucosum and production of Ochratoxin A is very low [34]. Similar to temperature and moisture contents, gas compositions are other factors for the growth of fungi and production of Ochratoxin A in different food stuffs, dried fruit, alcoholic beverage and coffee. According to [41]

Analytical methods for Ochratoxin A
Several methods have been developed for the determination of Ochratoxin A in a variety of food commodities including cereals (barley, corn, wheat bran, and flour), coffee, cocoa, wine, beer, and dried fruits. In the present review some of the analytical methods that used for determination, separation and quantifi cation of Ochratoxin A from different food stuffs, feeds and alcoholic beverage were discuss as follow.

Thin layer chromatography
Thin Layer Chromatography is a technique used to isolate non-volatile mixtures. During conducting the experiments, sheet of aluminum foil, plastic, or glass which is coated with a thin layer of adsorbent materials and materials usually including aluminum oxide, cellulose or silica gel [45]. Similar to the other chromatography methods, thin layer chromatography is depends on the separation principles. The separation relies on the relative affi nity of compounds towards both the phases.
The mobile phase move over the surface of the stationary phase [45]. On the other hand, thin layer chromatography the most widely used and established separation and detection technique for afl atoxin since its developments in the 1960s [46]. Not only for afl atoxin analysis but also used for determination of Ochratoxin A from different agricultural crops product. For instance, detection of 2.4 -4 mg/Kg of Ochratoxin A, and 10 mg/Kg of Ochratoxin A from rice and wheat respectively [47].
The use of thin layer chromatography analysis for mycotoxins is still popular for both quantitative and semi-quantitative purposes. The main reason is due to its high throughput of samples, low operating cost and ease of identification of target compounds, using UV-vis spectral analysis especially in developing country [48]. However, when compared with other chromatographic techniques, thin layer chromatography detection limit is high, length of separation is limited and lack of automation [49].

Enzyme-Linked Immunosorbent Assay (ELISA)
Enzyme linked immunosorbent assay is a plate based assay technique which is used for detecting and quantifying substances such as peptides, proteins, antibodies and hormones.
Similar to the previously discussed analytical methods, also enzyme linked immunosorbent assay methods have their own advantages and limitations during the conducting of analysis.
Enzyme linked immunosorbent assay methods have advantages due to their simplicity, and number of samples that can be analyzed at the same time. In addition, it requires low volume samples, quicker sample cleanup, simple and specifi c [45].
However, enzyme linked immunosorbent assay is less accurate and relatively low sensitivity and low effi ciency compared with other chromatography techniques [50]. In addition, false positive or negative results are observed because of cross-reactions among molecules or interferences. Therefore, enzyme linked immunosorbent assay kits should not be used as a quantitative method and should only be used with foods for which they have been extensively tested and demonstrated to work [51]. Moreover, one of the documented reports showed that during the uses of enzyme linked immunosorbent assay methods suffi cient controls must be employed for each test, to ensure the validity of the quantifi cation unless diffi cult to obtain accurate result [52].

High Performance Liquid Chromatography (HPLC)
In fact, more advanced and sensitive analytical methods analysis compared with other chromatographic methods [53]. For example, compared with thin layer chromatography methods HPLC is extremely quick and effi cient [54]. The reason that it uses a pump, rather than gravity, to force a liquid solvent through a solid adsorbent material, with different chemical components separating out as they move at different speeds [49]. Moreover, the process can be completed in roughly 10 to 30 minutes, and it delivers high resolution.