Quantitative analysis of aflatoxin-albumin adducts

Quantitative exposure assessments made using biologically relevant markers will facilitate epidemiological studies of risk from environmental carcinogens. Blood proteins are readily accessible macromolecules that have been shown to be targets for activated chemical carcinogens. Serum albumin is quantitatively the most abundant target for aflatoxin B1 and the measurement of aflatoxin-serum albumin adducts has been used to detect exposed individuals. The goal of these experiments was to devise an analytical procedure that would increase the overall recovery of aflatoxin adducts in serum albumin, and thereby improve the accuracy of exposure monitoring. The method developed consisted of the following procedures. Proteins were precipitated from serum (< or = 100 microliters) with 80% ammonium sulfate, with incubation at 4 degrees C for 2 h. Following dialysis against phosphate-buffered saline (pH 7.0 for 3 h at 4 degrees C), the proteins were digested with protease (Pronase) (1:4.1 w/w enzyme:protein) for 15 h at 37 degrees C with shaking. Enzyme and other undigested proteins were precipitated with acetone (1:2 v/v, 40 min, 4 degrees C). After evaporation of the acetone under vacuum, levels of aflatoxin B1-albumin adducts were determined by radioimmunoassay carried out on 300 microliters fractions. This procedure obviated the isolation of albumin prior to analysis and reduced interference in the radioimmunoassay. High recoveries of aflatoxin B1 adducts were achieved together with a low limit of detection. The applicability of the procedure in epidemiological studies of human aflatoxin exposure was illustrated by results of analysis of aflatoxin-albumin adducts in serum samples from residents of Chongming Island, People's Republic of China.