One of the tracer methods often employed to measure glucose turnover in the non-steady state uses the one compartment model of Steele (Ann. NY Acad Sci 1959). However, this model gives adequate results when it is assumed that only a fraction of the glucose pool takes part in rapid changes of glucose specific activity, thereby being necessary to use a correction factor called the "pool fraction." The aim of this study was to experimentally determine the best pool fraction needed in the rat for the calculation of glucose turnover using a one-compartment model. This is important as no data are available so far in this widely used species. For this purpose, glucose turnover was measured in anesthetized lean and genetically obese fa/fa rats, using two different experimental designs. In all conditions, the error in estimating the total rate of glucose appearance was lowest when 0.5 was used as the pool fraction. The error was greater with an increase and a decrease in the pool fraction value. It is concluded that in the rat the one-compartment model measures changes in glucose turnover with reasonable accuracy in non-steady-state conditions and that a pool fraction of 0.5 gives the best results.