A preliminary study of the bulk hydrogen, carbon and nitrogen stable isotope composition of 28 authentic saffron samples produced from Crocus sativus L. cultivated in the typical production areas of Western Macedonia in Greece (8), Khorasan Province in Iran (7), Sardinia in Italy (6) and Castilla-La Mancha in Spain (7) is described. A chemical characterisation of 16 key quality parameters was also completed on the same samples by UV–Vis, HPLC and GC analyses. Multivariate analysis of the data revealed that 60.7% of saffron samples could be correctly assigned to their respective production countries using the chemical parameters. However, the combined bio-element stable isotope data reliably classified 100% of the saffron samples according to their respective geographical origins using posterior cross validation. Further work is required to establish the long-term stability of these models with respect to different years of production and other major producers such as India and Morocco.Highlights► Geographical differentiation of saffron spice by multi-element isotope analysis. ► UV-Vis, HPLC and GC techniques were used for saffron chemical characterisation. ► Differentiation according to chemical composition was possible with 61%. ► The combined bio-element stable isotope data reliably classified 100% of these samples.