Aims To study the biotechnological production of lipids containing rich amounts of the medically and nutritionally important -linolenic acid (GLA), during cultivation of the Zygomycetes Thamnidium elegans, on mixtures of glucose and xylose, abundant sugars of lignocellulosic biomass. Methods and Results Glucose and xylose were utilized as carbon sources, solely or in mixtures, under nitrogen-limited conditions, in batch-flask or bioreactor cultures. On glucose, T.elegans produced 31 center dot 9gl1 of biomass containing 15 center dot 0gl1 lipid with significantly high GLA content (1014mgl1). Xylose was proved to be an adequate substrate for growth and lipid production. Additionally, xylitol secretion occurred when xylose was utilized as carbon source, solely or in mixtures with glucose. Batch-bioreactor trials on glucose yielded satisfactory lipid production, with rapid substrate consumption rates. Analysis of intracellular lipids showed that the highest GLA content was observed in early stationary growth phase, while the phospholipid fraction was the most unsaturated fraction of T.elegans. Conclusions Thamnidium elegans represents a promising fungus for the successful valorization of sugar-based lignocellulosic residues into microbial lipids of high nutritional and pharmaceutical interest. Significance and Impact of the Study Xylitol production and cultivation in bioreactor trials is reported for the first time for T.elegans, while cultivation on xylose-based media resulted in high GLA production by this fungus.