Chemopreventive Effects of Certain Nutritive Acids on Aflatoxin-induced Cytotoxicity

Abstract

Many investigators have evaluated the effects of several plant products in modulating in vitro metabolism of a atoxin B 1 (AFB 1 ) by microsomal mixed function oxidase (MFO), thereby reducing its toxicity. However, the importance of these studies is of limited signi cance as MFO is not present in all cells. Our earlier study revealed a concentrationdependent increase in the rate of haemolysis, indicating a atoxin-induced cytotoxicity on red blood cells. It could be due to the direct action of a atoxin on the plasma membrane, as MFO required for bioactivation of AFB 1 is absent in red blood cells. The present study was designed to evaluate the ef cacy of some micronutrients, namely lactic, citric and folic acid, in ameliorating a atoxin-induced cytotoxicity on red blood cells in vitro.A atoxin was obtained by growing Aspergillus parasiticus (NRRL 3240) on sucrose-magnesium sulphate-potassium nitrate-yeast extract (SMKY) liquid medium at 28 6 2°C for 10 days [1]. Extraction and quanti cation of a atoxin were performed using the method of Nabney and Nesbitt [2].Blood samples were taken by venepuncture from healthy adult New Zealand strain rabbits (Oryctolagus cuniculus) and collected in heparinized tubes. A red blood cell (RBC) suspension in normal saline (0.9% NaCl) with a cell density of 2 3 10 4 cells ml 2 1 was prepared [3]. To assess the ef cacy of the test compound (folic acid, citric acid or lactic acid) in amelioration of a atoxin-induced cytotoxicity (haemolysis), 1 ml of the RBC suspension was added to either (a) the test compound (10 l g ml 2 1 ), (b) 2.5 l g ml 2 1 a atoxin [3], or (c) 2.5 l g ml 2 1 a atoxin and 5-100 l g ml 2 1 test compound. The volume of each tube was made up to 2 ml with additional saline in order to have the required concentrations of a atoxin and test compound which were also prepared in normal saline. The tubes were mixed gently and incubated at 37°C for 16 hours with intermittent shaking. Thereafter, the tubes were centrifuged at 1000g for 10 min and the colour density of the supernatant was measured spectrophotometrically at 540 nm. The percentage haemolysis was calculated using the formula: Percentage haemolysis 5Absorbance of the individual tubes Absorbance with 100% haemolysis 3 100.