Detection some Aflatoxins in some locally marketed Raw Peanuts

 

Rima Khalil¹, Moufid Yassin², Sundus J. Yaseen³

¹Master Student in Analytical and Food Chemistry Department, Faculty of Pharmacy, Tishreen University, Latakia, Syria and a Member of a Technical Committee at the University of Kalamoon University,

Deir Atieh, Syria.

²Professor in Analytical and Food Chemistry Department, Faculty of Pharmacy, Tishreen University,

Latakia, Syria.

³Doctor in Biochemistry and Microbiology, Faculty of Pharmacy, Kalamoon University, Deir Atieh, Syria.

 

ABSTRACT:

Aflatoxins are toxic secondary metabolites which are produced by some fungi that invade many different crops before and after harvest especially peanuts. In this study twenty samples of raw peanut kernels were randomly collected from different supermarkets in Latakia city, Syria for detection of their contamination with aflatoxins. The levels of aflatoxins in collected samples were determined using high performance liquid chromatography equipped with post-column photochemical reactor for enhanced detection and fluorescence detector. Extraction and cleanup of aflatoxins were achieved by 70% methanol and immunoaffinity column (IAC). A detection limit of (0.02-0.04ng/ml) and a quantification limit of (0.06-0.132 ng/ml) were obtained for the studied types of aflatoxins. The linearity was in the range (0.156-20ng/ml) for AFB₁, AFB₂, AFG₁ and AFG₂. 55% and 25% of the samples were contaminated with Total aflatoxins and AFB₁, respectively. The concentrations in all investigated samples ranged from not detected to 149.96µg/kg with mean concentration ±SD (16.49±34.08 µg/kg) for total aflatoxins and from not detected to 37.10µg/kg with mean ±SD (3.46±8.94µg/kg) for AFB₁. 25% and 50% of tested samples exceeded the maximum acceptable limits set by the European commission.

 

 

 

1. INTRODUCTION:

Peanut (also called ground nut);¹ is taxonomically classified as Arachis hypogaea, Family: Leguminosae,² It involves in different industries,³ including food industries like butter, roasted peanut, snacks, confections⁴ and oil production⁵ Peanut products are widely consumed from all age groups,⁶ it provides the body with important nutrients such as proteins, fat, vitamins and minerals.⁷ On the other hand, peanuts could cause allergy for some people,⁸ while the greatest risk facing peanut production is the high potential for contamination with aflatoxins (AFs).⁹

 

Aflatoxins are produced as secondary metabolites by many fungi, mainly Aspergillus flavus, Aspergillus parasiticus and Aspergillus nomius.¹⁰ Fungi are one of the most important microorganisms that exist in the soil,¹¹ they can produce compounds that are medically useful,^12,13 on the other hand, some fungi produce compounds such as aflatoxins that are very toxic products.¹⁴ AFs production occurs preferably in tropical and subtropical regions where optimum conditions are available.^15,16

 

There are at least eighteen different types of AFs,¹⁷ the most important ones are AFB₁, AFB₂, AFG₁ and AFG₂,¹⁸ in addition to AFM₁ and AFM₂ which are two metabolites of aflatoxin B₁ and B₂, respectively. AFM₁ and AFM₂ were detected for the first time in cow's milk.^19,20 Chemically, aflatoxins are di-furano-coumarin derivatives with a bi-furan group attached to the coumarin nucleus and a pentanone ring (in case of AFs type B) or a lactone ring (in case of AFs type G), Figure (1).¹⁷

 

Figure (1): Chemical structures of the most important aflatoxins; the symbols A, B, C and D indicate Aflatoxin B₁, (B) Aflatoxin B₂, (C) Aflatoxin G₁ and (D) Aflatoxin G₂, respectively.^21]

 

Aflatoxins could be found in many foods.²² In general, foods that are contaminated with aflatoxins pose a real threat to human and animal health.^23,24 Aflatoxicosis is a disease caused by consumption of food contaminated with aflatoxins.²⁵ Aflatoxicosis is classified as acute aflatoxicosis²⁶ which can result in death;²⁷ or as a chronic aflatoxicosis which may cause cancer and mutations.^28,29 AFs, in particular AFB₁, are well known as carcinogens to human and animals.³⁰ AFB₁ is classified by The International Agency for Research on Cancer (IARC) as a group (A) carcinogen.³¹ AFs types are ordered according to their ability to induce toxicity, carcinogenicity, mutagenicity as AFB₁> AFG₁> AFB₂>AFG₂.³²

 

The toxicity of aflatoxins comes from their biological transformation process that undergoes in vivo.³³ After eating contaminated food, AFB₁ is absorbed via the gastrointestinal tract into the portal blood and then to the liver where it is metabolized.³⁴ AFB₁ has been shown to be converted primarily by cytochrome P450 in the liver to the highly reactive derivative AFB₁-exo-8,9-epoxide(AFBO).^30,35 AFBO has a high potency to bind to macromolecules like proteins and Deoxyribonucleic Acid (DNA);^30,36 this binding may cause suppression of the p53 tumor Gene explaining the formation of DNA mutations and carcinogenic effect of AFB₁.³⁷

 

In order to achieve a good food quality, food safety must be achieved,^38] The European Commission (EC) has set acceptable levels of AFs in many food, for peanuts the Maximum Tolerable Limit (MTL) is 4μg/kg for Total aflatoxins (Total AFs) and 2μg/kg for AFB₁ when they are intended for direct consumption.³⁹ While, according to Food and Drug Administration of the United States (FDA), total aflatoxins should not exceed 20μg/kg in all types of food.⁴⁰ Because of the health awareness of food contamination,⁴¹ many analytical techniques were used to detect the levels of aflatoxins in different foods.⁴² The most frequent techniques in scientific literatures are chromatographic methods;⁴³ like Thin Layer Chromatography (TLC),⁴⁴ High Performance Liquid Chromatography (HPLC)⁴⁵ and Enzyme Linked Immunosorbent Assay (ELISA).⁴⁶

 

In general, peanuts is one of the major sources for human exposure to aflatoxin;⁴⁷ This is due to their high possibility of contamination as well as their wide consumption.⁴⁸ High level of aflatoxins could be expected in peanut products when the initial level of aflatoxins in raw samples is high;⁴⁹ this may result in high risk for individuals who consume peanut products regularly,⁵⁰ which leading to serious problems due to their toxicological effects.⁵¹

 

The importance of this research comes from the absence of information on the presence of aflatoxins in foods marketed in Syria; and from the serious health consequences of eating foods contaminated with these toxins. Thus, the aim of the current study is to detect the levels of aflatoxins in raw peanuts' samples marketed in some local market in Latakia City, Syria.

 

2. MATERIALS AND METHODS:

2.1. Laboratory analysis:

The analysis of samples was conducted in the laboratories of the Syrian Ministry of Internal Trade and Consumer Protection.

 

 

 

2.2. Samples:

Twenty raw peanut samples (weight of each sample 500 g) were obtained from supermarkets located in five different areas in Latakia City which is one of the Mediterranean coastal regions of Syria. Samples were coded according to the area of sampling and the date of sampling from all the five regions. All samples were ground with blender and stored in well-sealed nylon plastic bags in a freezer till use. Before the analysis, the target sample was taken out of freezer till equilibrate with room temperature.

 

2.3. Chemicals and Reagents:

Mixed standard solution for total aflatoxins of 1,000 ng/ml (containing 250ng of aflatoxin B₁, 250ng of aflatoxin B₂, 250ng of aflatoxin G₁ and 250ng of aflatoxin G₂ in one milliliter of methanol) was purchased from Biopharm Rhône Ltd (Darmstadt, Germany). Tetra distilled water was made in the laboratories of the Syrian Ministry of Internal Trade and Consumer Protection. Acetonitrile-HPLC grade was obtained from Biosolve Chimie (Dieuze, France). Methanol-HPLC grade was purchased from (LiChrosolv®Merck Millipore, Darmstadt, Germany). Sodium Chloride NaCl was obtained from Sigma chemical Co. (St. Louis, MO, U.S.A.). Immunoaffinity Columns (IAC) AFLARHONE® WIDE were obtained from (R-Biopharm, Darmstadt, Germany).

 

2.4. Apparatus:

Blender (Hindico, BL-999-3 IN 1, China) was used for treating the samples. HPLC analysis was performed using a Shimadzu 20A Gradient LC System (Kyoto, Japan) consisted of a (LC-20AT) pump, (SIL-20A) auto sampler, (DGU 20A5) Prominence degasser, (CTO-20A) column oven, (CBM-20A) system controller and (RF-10AXL) fluorescence detector. On-line photochemical reactor for enhanced detection UVETM LC Tech GmbH (Dorfen, Germany) was used for Post Column Derivatization (PCD). Shimadzu LC solution software was used to carry out the integration and calculations.

 

2.5. Preparation of standard solutions:

The stock solution of total aflatoxins standard mixture (1,000 ng/ml) was diluted with methanol to obtain intermediate solution with a concentration of 100 ng/ml for total aflatoxins which corresponds to 25 ng/ml of AFB₁, as well as of AFB₂, AFG₁ and AFG₂. From this intermediate solution, a series of diluted standard solutions were prepared using appropriate volumes of HPLC mobile phase to obtain concentrations (20, 10, 5, 2.5, 1.25, 0.625,0.312 and 0.156ng/ml) for each type of the four AFs.

 

2.6. Extraction and Cleanup for aflatoxins:

Aflatoxins extraction and cleanup were done according to Association of the Official Analytical Chemistry (AOAC) method 991.31.⁵² The brief work steps included taking 25g of homogeneous ground sample, adding 5g of NaCl and mixing with 125ml of 70% methanol for few minutes in a blender. The extract was filtered using filter paper; 1ml of the filtrate was diluted with 30ml of distilled water and filtrated again. 15ml of this filtrate was passed through IAC at 1–2 drops per second. 10ml of deionized water was used to wash the IAC (this step was repeated twice). For elution the trapped aflatoxins in case of AFs contamination of the extracted sample, the column was flushed by 1ml of methanol and then collected into a vial. Finally, the volume of cleaned extract was adjusted to 2ml with distilled water and it was ready for injection into HPLC system.

 

2.7. Chromatographic Separation:

The chromatographic column was Brisa LC₂ C₁₈ (250 mm×4.6mm) with 5μm particle size. Mobile phase was consisted of water, methanol, and acetonitrile (3:1:1). The analysis was isocratically delivered at a flow rate of 1.2mL/min. The injection volume for both standard and samples was 50µl. The fluorescence detector was set at 360 nm as an λexc and 450nm as an λemi.

 

2.8. AFs Quantification:

The Limit of Detection (LOD), Limit of Quantification (LOQ) and linearity were estimated prior to the analysis of samples. LOD was calculated from the signal to noise ratios of 3:1 and LOQ was estimated from signal to noise ratio of 10:1.

 

Linearity was estimated by injection of aflatoxins standards solutions at concentrations of (20, 10, 5, 2.5, 1.25, 0.625, 0.312 and 0.156ng/ml) for each of AFB₁, AFB₂, AFG₁ and AFG₂. The equation of regression line (and calculation of factor of regression) (y = ax + b) for each aflatoxin was obtained.

 

3. RESULTS AND DISCUSSION:

Linearity, LOD and LOQ:

The linearity range and linear regression equation, the coefficient squared value (R²), LOD and LOQ for each individual aflatoxin are shown in (Table1).

 

Table 1: LOD, LOQ, the equation and range of linearity for AFB₁, AFB₂, AFG₁ and AFG₂.

Type of aflatoxin	Linearity range (ng/ml)	Linear regression equation	R2	LOD ng/ml	LOQ ng/ml
AFB1	0.156-20	Y1 = 7865.5x1 + 577.61	0.9995	0.02	0.06
AFB2	0.156-20	Y2 = 12376x2 - 144.71	0.9998	0.04	0.15
AFG1	0.156-20	Y3 = 10295x3 + 1085.5	0.9989	0.03	0.09
AFG2	0.156-20	Y4 = 8481.9x4 + 914.67	0.9997	0.07	0.12

 

Figure (2): (A) Chromatogram of AFs standard mixture 20 ng/ml containing 5ng/ml of each of AFG₂, AFG₁, AFB₂ and AFB₁, (B): Chromatogram of peanut sample was contaminated with AFB₂ and AFB₁, (C): Chromatogram of peanut sample was contaminated with AFG₂ and AFG₁, (D): Chromatogram of peanut sample was contaminated with AFG₂,AFG₁,AFB₂ and AFB₁, (E): Chromatogram of blank or negative sample (no Afs were detected).

 

The HPLC chromatograms of standard of Aflatoxin mixture (total AFs 20ng/ml), three positive samples and one negative sample are shown in (Figure 2). The chromatogram of Standard of Aflatoxins mixture shows that AFs have well-separated peaks and were eluted at time around 10.3, 11.9, 13.3 and 15.7 minutes for AFG₂, AFG₁, AFB₂ and AFB₁, respectively, (Figure 2, A).

 

Aflatoxins in tested peanuts samples:

In this study, the analysis of 20 raw peanut samples showed that (55%) of tested samples were positive for total AFs presence. As shown in Table 2, none of the four types of AFs was detected alone in all positive samples; at least two types of aflatoxins were shown together, and the four aflatoxins were found together only in one sample which was coded as (FR4).

 

The highest detectable level for total aflatoxins (149.96 ±0.31µg/kg) was recorded in sample (FR4) and in sample (FD4) for AFB₁ (37.10±0.14µg/kg), as shown in Table 2.

Table 2: Mean concentrations ±SD of AFB₁, AFB₂ , AFG₁ and AFG₂ and total aflatoxins in tested peanut samples (n=20).

Sample code	Mean AFB1 (µg/kg)±SD	Mean AFB2 (µg/kg)±SD	Mean AFG1 (µg/kg)±SD	Mean AFG2 (µg/kg)±SD	**Mean total AFs (µg/kg) ±SD
FR1	*ND	27.61±0.13	ND	8.35±0.02	35.96±0.15
FD1	3.58±0.05	1.89±0.07	ND	ND	5.42±0.01
FZ1	ND	ND	ND	ND	ND
FA1	ND	ND	ND	ND	ND
FS1	ND	2.28±0.1	1.56±0.18	ND	3.85±0.29
FR2	0.313±0.08	ND	11.96±0.19	ND	12.27±0.1
FZ2	ND	ND	ND	ND	ND
FD2	ND	ND	ND	ND	ND
FS2	ND	ND	12.54±0.11	12.38±0.16	24.93±0.07
FA2	ND	ND	ND	ND	ND
FS3	ND	ND	ND	ND	ND
FR3	ND	15.51±0.12	ND	4.91±0.05	20.42±0.17
FZ3	ND	ND	ND	ND	ND
FD3	ND	ND	6.66±0.1	2.57±0.07	9.24±0.17
FA3	ND	ND	ND	ND	ND
FS4	7.3±0.18	2.24±0.06	ND	ND	9.48±0.10
FD4	37.10±0.14	9.78±0.11	ND	ND	46.89±0.19
FZ4	ND	ND	ND	ND	ND
FA4	3.53±0.05	7.86±0.11	ND	ND	11.4±0.15
FR4	17.51±0.08	24±0.16	100.2±0.15	8.24±0.1	149.96±0.31
***Mean (µg/kg) ±SD	3.46±8.94	4.55±8.37	6.64±22.36	1.82±3.66	16.49±34.08

*ND: Not detected (below the quantification limit).

**Total aflatoxin was represented by the summation of aflatoxin B₁, B₂, G₁ and G₂ levels.

***The mean of individual aflatoxin in all tested samples was calculated by assuming that the level of each aflatoxin in samples below the detection limit was equal to zero.

 

Table 3. Number and percent of positive samples for total sample number (20) with the range and the mean ± SD of contamination for AFB₁, AFB₂, AFG₁ and AFG₂ in positive samples.

Type of aflatoxin	Number of positive samples (%)	(%) of positive samples	Range of contamination in positive samples (µg/kg)	Mean (µg/kg) ±SD of contamination in positive samples
AFB1	6	30%	(0.31-37.1)	11.6±13.82
AFB2	8	40%	(1.89-27.61)	11.38±10.08
AFG1	5	25%	(1.56-100.2)	26.58±41.39
AFG2	5	25	(2.57-12.38)	7.29±3.73
Total aflatoxins	11	55%	(3.58-149.96)	29.98±41.98

 

According to Table 3, (30%), (40%), (25%) and (25%) of the tested samples were contaminated with AFB₁, AFB₂, AFG₁ and AFG₂, respectively. In addition, the range of detectable concentrations was (0.31 -37.1µg\kg) for AFB₁ with a mean± SD of 11.6±12.62µg/kg and it was (3.85 - 149.96µg\kg) for total aflatoxins with a mean± SD of 29.98±40.03µg\kg.

 

The high AFs level in our outcomes can be explained by availability of several conditions that promote their production; like the weather and climate conditions of Latakia city which is characterized by a wet summer and rainy winter,⁵³or the bad storage practices in the markets.⁵⁴

 

The variant level of aflatoxins between the tested samples in our study could be interpreted by the different conditions that each sample was exposed to during the life cycle of peanuts until it reached the markets. At pre-harvest stage the most influencing factors are the different origin of cultivated peanuts and type of soil, agricultural practices, phytoalexin production.^6,51 While during the harvest period, the most crucial factor is the drying process.⁵¹ Sunlight drying is the most common method used, and there is no protocol for its implementation, but its application depends on the private opinion of farmers, such as the duration of the drying of peanuts.⁵⁴ On the other hand, the most influencing factors at post-harvest stage are the conditions of transportation and storage.⁵⁵ As, storing crops in conditions with high moisture content and temperature promotes production of AFs.⁵⁶

 

Many papers have estimated AFs levels in peanut samples; when we compare our results with those, we find that the present percent of total AFs contamination is in contrary with those reported in Sokoto State, Nigeria which is (82.5%);⁵⁷ and in Tabriz, Iran from two studies which are (17.3% and 33%).^58,59 On the other hand, this percent is somewhat compatible with those found in Lusaka, Zambia which was 55.4%⁶⁰ and in Karaman, Turkey which was (50%).⁶¹

 

Furthermore, in our study the highly detectable level of contamination for AFB₁ in raw peanut samples is close to that obtained by others which was 44μg/kg.⁶² But, it is different from what was recorded in Macedonia⁶³ and in Uganda⁵⁵, which were (289.2µg/kg) and (940µg/kg), respectively.

 

The incidence percent of each type of AFs in this study is incompatible with the finding of a Chinese study in which mentioned that AFB₁, AFB₂, AFG₁ and AFG₂ were appeared in 12.5%, 12.5%, 2.5% and 2.5% of studied peanut samples, respectively.⁶⁴ Furthermore, 57% of tested raw peanut samples were positive for AFB₁, 100% for AFB₂, 100% for AFG₁ and 0% for AFG₂ as mentioned in a Mexican study.⁶²

 

Compared to our results, other studies have shown differences in the range of AFB₁ and total AFs.^58,61,64 For instance, a Malaysian study showed that the mean concentration of AFB₁ and total AFs in contaminated peanut samples were 9.00 and 11.28µg/kg,⁶⁵ as well as they were 6.02 and 12.88μg/ kg in a Brazilian article,⁶⁶ while they were 11.6 and 29.98μg/ kg in our findings.

 

At least, the present data revealed that 5 samples (25%) and 10 samples (50%) of tested peanut samples exceeded the MTL set by the European Commission (EC) for AFB₁ and total AFs, respectively. These results disagree with a study in which the total AFs contamination level in all positive peanut samples was lower than MTL;^{59, 61} and they are different from that obtained in Egypt which indicated that 42% and 33% of peanut samples were higher than the permissible limits of EC for AFB₁ and total AFs, respectively.⁵⁴

 

Finally, the variation in the results between previous studies could be linked to the different climate and weather conditions and geographic location of the studied countries;^65,67,68 which create differences in the species of growing fungi and the strains of fungi producing toxins.⁶⁹

 

4. CONCLUSION:

Peanuts are one of the most traded nuts among the global, including Syria, but they are commonly contaminated by aflatoxins. Peanuts are widely used in food industries; therefore, this study was performed to determine AFs levels in raw peanut samples marketed in the Syrian City; Latakia. Although the limitations in this study, the results showed that the presence of aflatoxins was at levels exceeding the EC permissible limits for peanuts. We hope these findings to be considered by local authorities, especially in the absence of health awareness among consumers of high risk caused by aflatoxins on human health.

 

5. ACKNOWLEDGMENT:

I would like to thank everyone who contributed to achievement this work, and in particular I would like to mention Dr. Farah Youssef, for whom I am very grateful for her contribution to accomplishing this research in the best possible way.

 

6. REFERENCES:

1.      Sharmila, D. and L.J. Rebecca, A Study on The Influence of Vermicompost on The Growth Parameters of Arachis hypogaea. Research Journal of Pharmacy and Technology, 2019. 12(6): 2895-2901.

2.      Akerele, O. and S. Ejiko, Design and construction of groundnut oil expeller. International Journal of Engineering and Computer Science, 2015. 4(6): 12529-12538.

3.      Mahmoud, H., Z. Belel, and C. Nwakaire, Groundnut shell ash as a partial replacement of cement in sandcrete blocks production. International Journal of Development and sustainability, 2012. 1(3): 1026-1032.

4.      Yao, G., Peanut production and utilization in the People’s Republic of China. University of Georgia, 2004.

5.      David, S.B. and G. Raj, Development Studies of Biodegradable Pressure Sensitive Adhesives from Groundnut Oil and Butylmethacrylate. Asian Journal of Research in Chemistry, 2010. 3(1): 40-46.

6.      Lavkor, I. and I. Var, The Control of Aflatoxin Contamination at Harvest, Drying, Pre‐Storage and Storage Periods in Peanut: The New Approach. Aflatoxin-control, analysis, detection and health risks, 2017.

7.      Settaluri, V., et al., Peanuts and their nutritional aspects—a review. Food and nutrition Sciences, 2012. 3(12): 1644.

8.      Svitlana, A.S., Peanut Allergy-A Review. Research J. of Pharm. and Tech, 2016. 9(8): 1280-1286.

9.      Nyirahakizimana, H., et al., Occurrence of Aspergillus species and aflatoxin contamination in raw and roasted peanuts from formal and informal markets in Eldoret and Kericho towns, Kenya. Advances in Microbiology, 2013. 3(4): 333-342.

10.   Banu, N. and J.P. Muthumary, Screening of aflatoxigenic property of some Aspergillus flavus isolated from sunflower seeds and its products at sunflower oil refineries. Research Journal of Science and Technology. 2(5) 2010 : 102-107.

11.   Khan, P.A. and A. Ekka, Characterization of Phosphate solubilizing Microorganism isolated from hsoil. Research Journal of Pharmacy and Technology, 2019. 12 (3): p. 1353-1356.

12.   Sreenivasulu, V., K. Jayaveera, and P.M. Rao, Solid-state fermentation for the production of l-asparaginase by Aspergillus Sp. Research Journal of Pharmacognosy and Phytochemistry, 2009. 1 (1): 21-25.

13.   Chand, T., et al., Production of Industrially Significant Enzymes from Bio-Wastes Using Aspergillus niger by Solid State Fermentation. Research Journal of Pharmaceutical Dosage Forms and Technology, 2014. 6(1): 33-36.

14.   Pitt, J.I., Foodborne Infections and Intoxications: Chapter 30. Mycotoxins. 2013: Elsevier Inc. Chapters.

15.   Shekhar, M., et al., Effects of Climate Change on Occurrence of Aflatoxin and its Impacts on Maize in India. Int. J. Curr. Microbiol. App. Sci. 2018;7(6): 109-116.

16.   Medina, A., A. Rodriguez, and N. Magan, Effect of climate change on Aspergillus flavus and aflatoxin B1 production. Frontiers in Microbiology, 2014; 5: 348.

17.   Coppock, R.W., R.G. Christian, and B.J. Jacobsen, Aflatoxins, in Veterinary Toxicology. 2018; 983-994.

18.   Meena, A.K., et al., A Comparative Study of the Quality Assessment of Vitex negundo Linn Leaves Collected from Three Different Geographical Locations. Asian Journal of Research in Chemistry, 2011; 4(6): 986-989.

19.   Saleem, F., B. Sadia, and F.S. Awan, Control of aflatoxin production using herbal plant extract. Aflatoxin: Control, Analysis, Detection and Health Risks, 2017; 1.

20.   Yu, L., et al., Graphene oxide: an adsorbent for the extraction and quantification of aflatoxins in peanuts by high-performance liquid chromatography. Journal of Chromatography A, 2013. 1318: p. 27-34.

21.   Asghar, M.A., et al., Occurrence of aflatoxins contamination in brown rice from Pakistan. Iranian journal of public health, 2014; 43(3): 291.

22.   Martinez-Miranda, M.M., M. Rosero-Moreano, and G. Taborda-Ocampo, Occurrence, dietary exposure and risk assessment of aflatoxins in arepa, bread and rice. Food control, 2019; 98: 359-366.

23.   Juan, C., A. Ritieni, and J. Maٌes, Determination of trichothecenes and zearalenones in grain cereal, flour and bread by liquid chromatography tandem mass spectrometry. Food chemistry, 2012; 134(4): 2389-2397.

24.   Waliyar, F., et al., Prevalence and distribution of aflatoxin contamination in groundnut (Arachis hypogaea L.) in Mali, West Africa. Crop Protection, 2015. 70: 1-7.

25.   Dharumadurai, D., et al., Aflatoxins and Aflatoxicosis in Human and Animals. Aflatoxins-Biochemistry and Molecular Biology, InTech DOI, 2011; 10: 22717.

26.   Mousavi Khaneghah, A., et al., Aflatoxins in cereals: State of the art. Journal of food safety, 2018; 38(6): 12532.

27.   Guchi, E., Aflatoxin contamination in groundnut (Arachis hypogaea L.) caused by Aspergillus species in Ethiopia. Journal of Applied and Environmental Microbiology, 2015; 3(1): 11-19.

28.   Al-Hammadi, S., et al., The cytotoxicity of aflatoxin b1 in human lymphocytes. Sultan Qaboos University medical journal, 2014; 14(1): 65.

29.   Kumar, V., Aflatoxins: Properties, toxicity and detoxification. Nutrition and Food science International Journal, 2018; 6: 555696.

30.   Bbosa, G.S., et al., Aflatoxins metabolism, effects on epigenetic mechanisms and their role in carcinogenesis. Health, 2013b; 5(10):  720-726.

31.   Bakırdere, S., et al., Aflatoxin species: their health effects and determination methods in different foodstuffs. Central European Journal of Chemistry, 2012: 10(3): 675-685.

32.   Bbosa, G.S., et al., Review of the biological and health effects of aflatoxins on body organs and body systems. Aflatoxins-recent advances and future prospects, 2013a; 12: 239-265.

33.   Marchese, S., et al., Aflatoxin B1 and M1: Biological properties and their involvement in cancer development. Toxins, 2018; 10(6): 214.

34.   Agag, B., Mycotoxins in foods and feeds: 1-aflatoxins. Ass. Univ. Bull. Environ. Res, 2004; 7(1): 173-205.

35.   Santini, A. and A. Ritieni, Aflatoxins: risk, exposure and remediation, in Aflatoxins–Recent advances and future prospects. 2013, In Tech.

36.   Rushing, B.R. and M.I. Selim, Aflatoxin B1: A review on metabolism, toxicity, occurrence in food, occupational exposure, and detoxification methods. Food and chemical toxicology, 2018.

37.   Asemoloye, M.D., et al., Incidence and chemical implications of aflatoxin in street-vended foods. Aflatoxin: Control, Analysis, Detection and Health Risks, 2017: 153.

38.   Chatterjee, R. and P. Sharma, A comparative study regarding food adulteration and food safety in rural and urban area. Research Journal of Humanities and Social Sciences, 2017. 8(1): 65-68.

39.   Commission, E., Commission Regulation (EC) No. 401/2006 of 23 February 2006 laying down the methods of sampling and analysis for the official control of the levels of mycotoxins in foodstuffs. Off J Eur Union, 2006; 70: 12-34.

40.   Alshannaq, A. and J.-H. Yu, Occurrence, toxicity, and analysis of major mycotoxins in food. International journal of environmental research and public health, 2017; 14(6): 632.

41.   Kathirvel, S., et al., An Overview of Sample Preparation Techniques for Food Analysis. Asian Journal of Pharmaceutical Analysis, 2017; 7(1).

42.   Stroka, J., Determination of aflatoxins in food and feed with simple and optimised methods. 2001, Universitنt Wuppertal, Fakultنt für Mathematik und Naturwissenschaften.

43.   Chauhan, N.M., Aflatoxin: A Risky Menace for African’s Food Commodities. Aflatoxin: Control, Analys, Detection and Health Risks, 2017: 91.

44.   Braicu, C., et al., Screening and quantification of aflatoxins and ochratoxin A in different cereals cultivated in Romania using thin‐layer chromatography‐densitometry. Journal of Food Quality, 2008; 31(1)108-120.

45.   Namjoo, M., et al., Quantitative determination of aflatoxin by high performance liquid chromatography in wheat silos in Golestan province, north of Iran. Iranian journal of public health, 2016; 45(7):  905.

46.   Wacoo, A.P., et al., Methods for detection of aflatoxins in agricultural food crops. Journal of Applied Chemistry, 2014.

47.   Mahato, D.K., et al., Aflatoxins in food and feed: An overview on prevalence, detection and control strategies. Frontiers in Microbiology, 2019; 10:  2266.

48.   Lizلrraga-Paulيn, E.G., E. Moreno-Martيnez, and S.P. Miranda-Castro, Aflatoxins and their impact on human and animal health: an emerging problem, in Aflatoxins-Biochemistry and Molecular Biology. 2011, In tech Open.

49.   Bakhiet, S.E.A. and A.A.A. Musa, Survey and determination of aflatoxin levels in stored peanut in Sudan. Jordan Journal of Biological Sciences, 2011. 147(614): 1-16.

50.   Hong, L.S., N.I.M. Yusof, and H.M. Ling, Determination of aflatoxins B1 and B2 in peanuts and corn based products. Sains Malaysiana, 2010; 39(5): 731-735.

51.   Diao, E., et al., Factors influencing aflatoxin contamination in before and after harvest peanuts: a review. Journal of Food Research, 2015; 4(1): 148.

52.   Truckness, M., AOAC Official Method 991.31, Aflatoxin in corn, raw peanuts, and peanut butter. J. AOAC Int, 2000; 49: 22-24.

53.   Haydar, M., L. Benelli, and C. Brera, Occurrence of aflatoxins in Syrian foods and foodstuffs: a preliminary study. Food Chemistry, 1990; 37(4): 261-268.

54.   Abdel-Rahman, G.N., et al., Identify the natural levels of mycotoxins in Egyptian roasted peanuts and the DESTRUCTIVE effect of gamma radiation. The Journal of Microbiology, Biotechnology and Food Sciences, 2019; 8(5): 1174.

55.   Osuret, J., et al., Aflatoxin contamination of selected staple foods sold for human consumption in Kampala markets, Uganda. Journal of Biological Sciences, 2016; 16(1): 44.

56.   Adetunji, M.C., et al., Microbiological quality and risk assessment for aflatoxins in groundnuts and roasted cashew nuts meant for human consumption. Journal of toxicology, 2018.

57.   Salau, I., Aflatoxin Contamination of Stored Groundnut Kernel in Sokoto State, Nigeria. 2017.

58.   Ostadrahimi, A., et al., Aflatoxin in raw and salt-roasted nuts (pistachios, peanuts and walnuts) sold in markets of Tabriz, Iran. Jundishapur journal of microbiology, 2014. 7(1)

59.   Tahavori, A., F. Ghaderi, and M. Nemati, Determination of aflatoxins in nuts of Tabriz confectionaries by ELISA and HPLC methods. Advanced pharmaceutical bulletin, 2012; 2(1): 123.

60.   Bumbangi, N., et al., Occurrence and factors associated with aflatoxin contamination of raw peanuts from Lusaka district's markets, Zambia. Food Control, 2016; 68: 291-296.

61.   Kilicel, F., H.S. Karapinar, and A. Cimen, Quantitation of Aflatoxins in Food Materials Using HPLC-FLD Method. Science Journal of Analytical Chemistry, 2017; 5(6): 90.

62.   Alvarado-Hernلndez, J., et al., Aflatoxins in natural peanuts (Arachis hypogaea L.) of Mexico: Validation of the biochemical methods for extraction and quantification. J Plant Biochem Physiol, 2016; 4:  1000168.

63.   Stojanovska-Dimzoska, B., et al., Occurrence of aflatoxins in peanuts and peanut products determined by liquid chromatography with fluorescence detection. Matica Srpska Journal for Natural Sciences, 2013; (124): 27-35.

64.   Shi, Y.G., et al. Applied Technology in Using HPLC for Determining Aflatoxin Contents in Peanuts. In Advanced Materials Research. 2014. Trans Tech Publ.

65.   Arzandeh, S., J. Selamat, and H. Lioe, Aflatoxin in raw peanut kernels marketed in Malaysia. Journal of Food and Drug Analysis, 2010; 18(1)

66.   Oliveira, C., et al., Determination of aflatoxins in peanut products in the northeast region of Sمo Paulo, Brazil. International journal of molecular sciences, 2009; 10(1): 174-183.

67.   Akbas, M.Y. and M. Ozdemir, Effect of different ozone treatments on aflatoxin degradation and physicochemical properties of pistachios. Journal of the Science of Food and Agriculture, 2006; 86(13)2099-2104.

68.   Alhussaini, M.S., Mycobiota and mycotoxins of nuts and some dried fruits from Saudi Arabia. Journal of American Science, 2012; 8(12): 525-534.

69.   Mohammed, A., et al., Aspergillus and aflatoxin in groundnut (Arachis hypogaea L.) and groundnut cake in Eastern Ethiopia. Food Additives and Contaminants: Part B, 2016; 9(4): 290-298.

 

 

 

Accepted on 21.07.2020           © RJPT All right reserved

Research J. Pharm. and Tech 2021; 14(3):1431-1437.