BIOASSAY-DRIVEN ANALYSIS OF CHEWING TOBACCO EXTRACTS

The aims were to develop a method to simulate extraction of toxic comp onents into human saliva, to identify these components through bioassa y-driven analyses using the Microtox(R) test, and to quantify the toxi c compounds. Chewing tobacco extractions into artificial saliva simula ted oral chewing conditions. The Microtox bioassay-directed subfractio nation scheme identified acutely toxic components in chewing tobacco e xtracts. GC-MS identified and quantified the toxic components. The sub fractionation (pH 7 extraction, pH 2 extraction, then pH 10 extraction using organic solvents) scheme was shown to be comparable to biologic al digestive sequences and to simulate nicotine bioavailability. The l ow toxicity of artificial saliva allowed development of a sensitive bi oassay. Nicotine was the major acutely toxic agent detected. Classific ation by Draize ocular irritative categories showed that leached nicot ine concentrations were mildly to moderately irritating. Thus, the Mic rotox test can be used to detect irritative compounds and subfractions , and can be potentially applied to saliva biological monitoring in hu mans.