Sustainability of biofuels is increasingly taken into account; therefore, sustainable production technologies are needed. There has been a long history of converting Jerusalem artichoke into ethanol. Jerusalem artichoke (Helianthus tuberosus L.) is a low-requirement crop, it has a high carbohydrates yield and, nowadays, it does not interfere with food chain. It is, then, a promising energy crop for sustainable bioethanol production. However, the main storage carbohydrate of Jerusalem artichoke, inulin, can not be directly fermented by classic fermentation yeasts, so, either a hydrolysis followed by fermentation with classical yeast or the use of yeasts with inulinase activity are required to obtain bioethanol. Therefore, it is needed to know not only total sugar content, but also their composition, for the bioethanol production optimization from Jerusalem artichoke tubers. Several methods have been used in literature for carbohydrates analysis present in Jerusalem artichoke tubers. However, for further development of carbohydrate analysis, faster and more reliable identification and peak confirmation, mass spectrometry (MS) detection is required. In this paper, liquid chromatography with electrospray ionization mass spectrometry (LC-ESI-MS) was used as an alternative technique to analyse sugars content and composition in tubers from Jerusalem artichoke. Two simple, rapid, sensitive and specific LC-ESI-MS methods were developed under the positive ionization mode. Glucose, fructose, sucrose, kestose and inulin were determined. Furthermore, inulin profile can be characterized. Analytical reversed phase LC columns were used using only water as eluent. These methods can be useful to optimize the whole bioethanol production chain from Jerusalem artichoke. (C) 2011 Elsevier Ltd. All rights reserved.