During a 70-h fermentation of a lignocellulose hydrolysate, the ethanol produced was monitored on-line using a microdialysis probe as an in situ sampling device. The dialysate components were then separated in a column liquid chromatographic system and the ethanol was selectively detected by an amperometric alcohol biosensor. The result was compared with two off-line analysis methods : one chromatographic method with refractive index (RI) detection and one enzymatic method based on spectrophotometric detection. The two methods based on enzymes were shown to give lower values than the chromatographic method based on RI detection, which is discussed in terms of selectivity. The investigated on-line setup was found to be a flexible system for monitoring of fermentations, allowing a sampling frequency of at least 12 h(-1) and with a delay between sampling and detection of less than 5 min.