To demonstrate improved resolution and rapid analysis times achieved for the UPLC separation of aflatoxins M1, G2, G1, B2, and B1, without derivatization, using the ternary mixing capabilities of the ACQUITY UPLC® H-Class System with the Waters Aflatoxin Analysis Kit.
Aflatoxins are a group of mycotoxins produced as metabolites by the fungi Aspergillus Flavus and Aspergillus Parasiticus. They can be found in various foodstuffs such as grains, nuts, spices, and dairy products. There are four naturally occurring aflatoxins: B1, B2, G1, and G2. The third subset, M1, arise as a metabolic by-product when dairy cattle eat B1-contaminated grains. This can result in contaminated dairy products such as milk.
These compounds are toxic and can be carcinogenic to humans and animals. B1 and G1 are the more potent of the four naturally-occurring aflatoxins. Due to their toxicity, government regulatory agencies impose strict limits on aflatoxins in food-stuffs. For this reason, the food industry needs sensitive, accurate, and reproducible analytical methods to measure these compounds. The methods are usually based on reversed-phase HPLC with fluorescence detection. However, since reversed-phase eluents quench the fluorescence of aflatoxins B1 and G1, derivatization is commonly used to enhance the response of these analytes. Typical choices are pre-column derivatization with trifluoroacetic acid (TFA) or post-column derivatization with iodine, electrochemically-generated bromine, or photochemical UV.
These approaches are time consuming and require the purchase of costly post-column reactors, or, in the case of electrochemically-generated bromine, an electrochemical cell. In particular, the use of TFA raises concerns of handling safety by technicians.
Samples were prepared using the Waters Aflatoxin Analysis Kit featuring the VICAM AflaTest® P methodology prior to analysis. Using the ACQUITY UPLC H-Class System with its UPLC®-optimized fluorescence (FLR) Detector, a baseline separation of aflatoxins M1, G2, G1, B2, and B1 was achieved with a run time of 4.5 minutes. No derivatization was necessary. The ACQUITY UPLC H-Class System features a Quaternary Solvent Manager (QSM) and Auto•Blend™ Technology enabling dynamic, programmable blending of solvents. Here, a simple ternary mixture of water, methanol, and acetonitrile was applied in conjunction with an ACQUITY UPLC BEH C18 Column to perform the separation.