High-performance liquid chromatography ultraviolet-photodiode array detection method for aflatoxin B1 in cattle feed supplements

Research (Published online: 17-08-2017)

17. High-performance liquid chromatography ultraviolet-photodiode array detection method for aflatoxin B1 in cattle feed supplements

Lazuardi Mochamad and Bambang Hermanto

Veterinary World, 10(8): 932-938

ABSTRACT

Aim: The objective of the current study is to determine the concentration of aflatoxin B1 using high-performance liquid chromatography (HPLC) with a photodiode array (PDA) detector.

Materials and Methods: Aflatoxin B1 certified reference grade from Trilogy Analytical Laboratory dissolved acetonitrile (ACN) at 10 μg/mL was used for standard assessment. HPLC instruments such as ultraviolet-PDA detector used a Shimadzu LC-6AD pump with DGU-20A5 degasser, communication module-20A, and PDA detector SPD-M20A with FRC-10A fraction collector. The HPLC was set isocratic method at 354 nm with a reverse-phase ODS C18 column (LiChrospher® 100 RP-18; diameter, 5 μm) under a 20°C controlled column chamber. Rheodyne® sample loops were performed in 20 μL capacities. The mobile phase was performed at fraction 63:26:11 H2O: methanol:ACN at pH 6.8. A total of 1 kg of feed contained 10% bread crumbs and 30% concentrated, 40% forage, and 20% soybean dregs were using commercials samples. Samples were extracted by ACN and separated with solid phase extraction ODS 1 mL than elution with mobile phase to collect at drying samples performed. The samples were ready to use after added 1 mL mobile phase than injected into the system of HPLC.

Results: We found that the retention time of aflatoxin B1 was approximately 10.858 min. Linearity of 0.01-0.08 μg/mL aflatoxin B1 dissolved in mobile phase was obtained at R2=0.9. These results demonstrate that these methods can be used to analyze aflatoxin B1 and gain 89-99% recovery. The limit of detection of this assay was obtained at 3.5 x 10-6 μg/mL.

Conclusion: This method was easy to apply and suitable to analyzing at small concentrations of aflatoxin B1 in formulated product of feed cattle.

Keywords: feeds supplements, high-performance liquid chromatography photodiode array detector, isocratic methods, mycotoxin.