title: Development and validation of a high-performance liquid chromatography method with post-column derivatization for the detection of aflatoxins in cereals and grains.
abstract: A novel, reliable and rapid high-performance liquid chromatography (HPLC) method with post-column derivatization was developed and validated. The HPLC method was used for the simultaneous determination of aflatoxin B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) in various cereals and grains. Samples were extracted with 80:20 (v/v) methanol:water and purified using C18 (40-63 μm) solid-phase extraction cartridges. AFs were separated using a LiChroCART-RP-18 (5 μm, 250 × 4.0 mm(2)) column. The mobile phase consisted of methanol:acetonitrile:buffer (17.5:17.5:65 v/v) (pH 7.4) delivered at the flow rate of 1.0 mL min(-1) The fluorescence of each AF was detected at λex = 365 nm and λem = 435 nm. All four AFs were properly resolved within the total run time of 20 min. The established method was extensively validated as a final verification of the method development by the evaluation of selectivity (AFB1, AFB2, AFG1 and AFG2), linearity (R(2) ≥ 0.9994), precision (average SD ≤ 2.79), accuracy (relative mean error ≤ -5.51), robustness (p < 0.0080), ruggedness (p < 0.0100) and average recoveries (89.2-97.8%). The limits of quantification of AFB1, AFB2, AFG1 and AFG2 were 0.080, 0.073, 0.062 and 0.066 ng g(-1), respectively. Finally, the developed method was applied for the analysis of AFs in 45 samples comprising rice (n = 20), wheat (n = 15) and maize (n = 10). The results showed that 65% of rice, 20% of wheat and 80% of maize samples were found contaminated with AFs. Thus, according to the achieved results, it is suggested that the newly developed HPLC method could be effectively applied for the routine analysis of the AFs in different cereals and grains.
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan masif345@yahoo.com.
authors.forename: Muhammad Asif
authors.initials: MA
authors.lastname: Asghar
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.
authors.forename: Javed
authors.initials: J
authors.lastname: Iqbal
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.
authors.forename: Aftab
authors.initials: A
authors.lastname: Ahmed
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.
authors.forename: Mobeen Ahmed
authors.initials: MA
authors.lastname: Khan
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.
authors.forename: Zuzzer Ali
authors.initials: ZA
authors.lastname: Shamsuddin
authors.affiliation: Food and Feed Safety Laboratory, Food and Marine Resources Research Centre, PCSIR Laboratories Complex, Karachi, Pakistan.
authors.forename: Khalid
authors.initials: K
authors.lastname: Jamil
mesh_terms: D016604:Aflatoxin B1 / Q000032:analysis; D000348:Aflatoxins / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D002523:Edible Grain / Q000737:chemistry* / Q000382:microbiology; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D006863:Hydrogen-Ion Concentration; D057230:Limit of Detection; D016014:Linear Models; D012275:Oryza / Q000737:chemistry; D015203:Reproducibility of Results; D052616:Solid Phase Extraction; D000067348:Whole Grains / Q000737:chemistry* / Q000382:microbiology; D003313:Zea mays / Q000737:chemistry
chemical_list: D000348:Aflatoxins; C027955:aflatoxin G1; C029754:aflatoxin G2; C029753:aflatoxin B2; D016604:Aflatoxin B1
keywords: B2; G1 and G2; High-performance liquid chromatography; Kobra Cell™; aflatoxins B1; cereals and grains; method validation; post-column derivatization