The xylitol production has become an important process to investigate given the diverse product applications, especially in the food industry. The biotechnological xylitol production has been assessed mainly through an experimental approach via batch and fed-batch operation. This study presents an analysis for optimal operation of the fed batch process following a systematic methodology. The model based approach includes the collection, validation and calibration of the mathematical model, followed by the proposal of some operation scenarios. The implementation of such methodology allowed to increase 20% the yield of xylitol compared with the obtained previously in some experiments. Moreover, an analysis for the diauxic phenomenon was also performed, allowing to determine an optimal feeding ratio between substrates.