Assay to Measure Oral Bacterial Alcohol Dehydrogenase Activity

Background: Certain species of bacteria found in the oral cavity have been shown to metabolize alcohol into acetaldehyde, a potential carcinogen, via the enzyme alcohol dehydrogenase. It is hypothesized that measuring alcohol dehydrogenase activity can quantify exposure to acetaldehyde, which may correlate with risk of developing oral squamous cell carcinoma. Objectives: The purpose of this study was to design an assay to measure the propensity of a subject's oral flora to produce this carcinogen. Methods: Assay conditions began with a 50 ul bacterial sample or saliva sample treated to lyse bacteria, and diluted to 500 ul total volume with distilled water and 10 mM NAD. After 5 minutes, 5 ul of 100% ethanol was added and the reaction was monitored for 25 minutes. Conversion of NAD to NADH by the alcohol dehydrogenase reaction was monitored via change in absorbance at 340 nanometers using a dual beam UV spectrophotometer. Results: The assay was successful in detecting measurable alcohol dehydrogenase activity in a positive control at bacterial counts as low as 5.9 x 10 to the 7 live cells/ml. However, little activity was found in saliva samples from the subjects surveyed. Conclusions: A simple, quick assay for assessing alcohol dehydrogenase activity was optimized. The utility of such a test for using saliva to measure relative risk due to acetaldehyde synthesis will require testing on a broader range of subject specimens to determine if the assay can achieve appropriate sensitivity.

Acknowledgements: Funding provided by NIH/NIDCR grants T32 DEO14678-05 and DE10058. Thanks to Dr Philip Wertz for technical advice.