Production of biodiesel is currently limited due to lack of economically beneficial feedstocks. Suitability of municipal wastewater sludge and olive mill waste as feedstocks for biodiesel production was evaluated. The various bio-waste sources were analyzed for their oil content and fatty acid composition using conventional analyses complemented with time domain (TD)-NMR analysis. TD-NMR, a rapid non-destructive method newly applied in this field, yielded good correlations with conventional methods. Overall biodiesel yields obtained by TD-NMR analysis were 7.05% and 9.18% (dry wt) for olive mill pomace and liquid wastes, and 11.92%, 7.07%, and 4.65% (dry wt) for primary, secondary, and anaerobically stabilized sludge, respectively. Fatty acid analysis indicated fundamental suitability of these agro-industrial waste resources for biodiesel production. Evaluation of bio-waste materials by TD-NMR revealed the potential of this tool to identify waste-oil sources cost effectively and quickly, supporting expansion of a sustainable biodiesel industry in Israel and other regions.