A single step extraction-cleanup procedure using porous membrane-protected micro-solid phase extraction (μ-SPE) in conjunction with liquid chromatography-tandem mass spectrometry for the extraction and determination of aflatoxins (AFs) B1, B2, G1 and G2 from food was successfully developed. After the extraction, AFs were desorbed from the μ-SPE device by ultrasonication using acetonitrile. The optimum extraction conditions were: sorbent material, C8; sorbent mass, 20mg; extraction time, 90 min; stirring speed, 1,000 rpm; sample volume, 10 mL; desorption solvent, acetonitrile; solvent volume, 350 μL and ultrasonication period, 25 min without salt addition. Under the optimum conditions, enrichment factor of 11, 9, 9 and 10 for AFG2, AFG1, AFB2 and AFB1, respectively were achieved. Good linearity and correlation coefficient was obtained over the concentration range of 0.4-50 ng g(-1) (r(2) 0.9988-0.9999). Good recoveries for AFs ranging from 86.0-109% were obtained. The method was applied to 40 samples involving malt beverage (19) and canned coffee (21). No AFs were detected in the selected samples.
For application to the analysis of aflatoxins (AF) in commercial peanut and corn products, the ISOLUTE multimode column (IMC, solid phase multifunctional column) method was validated by comparing with the modified Florisil column (MFC) method. Twenty-two peanut and eight corn products from Malaysia and the Philippines were analysed for AFB1, AFB2, AFG1 and AFG2 firstly by the MFC method and then by the IMC method. For peanut products, 14 out of 22 samples were positive by the two methods in the range of 1-378 micrograms/kg of AF, and correlation coefficients (r) for AFB1 and AFB2 were 0.987 and 0.997, respectively. For corn and corn products, all the samples were positive in the range of 1-130 micrograms/kg, and r values were 0.992 and 0.805 for AFB1 and AFB2 respectively. Thus, the results were significantly (p < 0.01) in close agreement, particularly for lower range of 1-50 micrograms/kg of AF concentrations in all the samples. For the occurrence of AF, 11 (65%) of peanut products from Malaysia were contaminated with AF at a mean level of 50 micrograms/kg (maximum 180 micrograms/kg) and two (40% products from the Philippines were contaminated with as high as 375 micrograms/kg and 177 micrograms/kg of AF, respectively. All the corn products from the Philippines were contaminated with AF at a mean level of 44 micrograms/kg (maximum 130 micrograms/kg). Contamination of commercial foods with high levels of AF is a very important issue to both the countries since these foods are very popular among children.
Traditional herbal medicines, popularly known as 'jamu' and 'makjun' in Malaysia and Indonesia, are consumed regularly to promote health. In consideration of their frequent and prolonged consumption, the natural occurrence of aflatoxins (AF) in these products was determined using immunoaffinity column clean-up and high-performance liquid chromatography with pre-column derivatization. The evaluated method, which entails dilution of sample extracts with Tween 20-phosphate buffered saline (1:9, v/v) and a chromatographic system using isocratic mobile phase composed of water-methanol-acetonitrile (70:20:10, v/v/v), was effective in separating AFB1, AFG1 and AFG2 from interference at their retention times. Results were confirmed using post-column derivatization with photochemical reactor. For 23 commercial samples analyzed, mean levels (incidence) of AFB(1), AFB(2) and AFG1 in positive samples were 0.26 (70%), 0.07 (61%) and 0.10 (30%) microg/kg, respectively; one sample was positive for AFG2 at a level of 0.03 (4%) microg/kg. In contrast to the high levels of AF in crude herbal drugs and medicinal plants reported previously by other researchers, the low contamination levels reported in this study may be attributed to the higher selectivity to AF of the method applied. Based on the AFB1 levels and the daily consumption of positive samples, a mean probable daily intake of 0.022 ng/kg body weight was calculated.
Thirty milled rice samples were collected from retailers in 4 provinces of Malaysia. These samples were evaluated for Aspergillus spp. infection by direct plating on malt extract salt agar (MESA). All Aspergillus holomorphs were isolated and identified using nucleotide sequences of ITS 1 and ITS 2 of rDNA. Five anamorphs (Aspergillus flavus, A. oryzae, A. tamarii, A. fumigatus and A. niger) and 5 teleomorphs (Eurotium rubrum, E. amstelodami, E. chevalieri, E. cristatum and E. tonophilum) were identified. The PCR-sequencing based technique for sequences of ITS 1 and ITS 2 is a fast technique for identification of Aspergillus and Eurotium species, although it doesn't work flawlessly for differentiation of Eurotium species. All Aspergillus and Eurotium isolates were screened for their ability to produce aflatoxin and ochratoxin A (OTA) by HPLC and TLC techniques. Only A. flavus isolate UPM 89 was able to produce aflatoxins B1 and B2.