Quantification and validation of enzyme immunoassay for urinary aflatoxin B1–N7-guanine adduct for biological monitoring of aflatoxins

Abstract

The aflatoxin B₁–N⁷-guanine (AFB₁–N⁷-guanine) adduct has been established as one of the relevant biomarkers of dietary aflatoxin (AFB₁) exposure. Measurement of this adduct is potentially a useful dosimeter in molecular epidemiological studies. This paper reports the application and evaluation of a sensitive indirect competitive enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of urinary AFB₁–N⁷-guanine adduct in high risk populations exposed to dietary aflatoxin. Earlier, we had reported a simple and rapid indirect ELISA method for AFB₁–N⁷-guanine adduct in the urine and liver tissues using polyclonal antibodies specific to AFB₁–N⁷-guanine adduct. The method was evaluated using a rodent model (Fischer 344), exposed to 1 mg kg⁻¹ body mass of AFB₁ and human urine samples obtained from a maize eating population, environmentally exposed to AFB₁ through their diet. The levels of AFB₁–N⁷-guanine adduct in rat and human urine ranged from 6.42 to 20.16 μg mg⁻¹ creatinine and from 9.30 to 13.43 ng mg⁻¹ creatinine, respectively. The level of AFB₁ in the diet as estimated by ELISA ranged from 1000 to 3600 ng d⁻¹. The interesting observation in these studies is that the females (in both rodents and human subjects) are more efficient than males at excreting the adduct. Total adduct (DNA bound adduct and guanine adduct excreted in urine) was found to be similar in male and female rats. However, 63% of the total adduct was accounted for in urine of female rats, whereas male rats excreted 47% of the total adduct in their urine. The present method may find wide application as a biochemical tool in molecular epidemiological studies with respect to human exposure to dietary aflatoxins.