The renal fractional extraction (Ex) is a parameter describing the ability of the kidneys to remove a substance x from the circulating plasma. Ex is calculated often as the ratio between the arteriovenous concentration difference and the arterial concentration. This method simplifies the calculations but it is associated with an underestimation of Ex. In the study described here, a theoretical analysis of the error is made, with a graphical presentation of its magnitude in different diuresis/renal plasma flow (RPF) ratios and at different levels of Ex. The error was analysed also in a clinical situation where the renal extraction of PAH (EPAH) and EDTA (EEDTA) were determined in six patients during different stages of cardiac surgery. The underestimation of EPAH was seldom more than 4%, while EEDTA was underestimated often with more than 20%. It is concluded that the simplified formula is accurate when calculating the renal extraction for substances like PAH, with a normally high extraction, even if the diuresis/RPF ratio is high. For substances with low extractions, e.g. filtration markers, in some clinical and experimental situations it is necessary to take the renal plasma flows into consideration or to arrange for low urine production to avoid significant errors.