AIMS: To identify if culture conditions affect the chemical composition of exopolysaccharide (EPS) produced by Aureobasidium pullulans. METHODS AND RESULTS: In batch airlift and continuously stirred tank (CSTR) reactors the EPS produced with low (0.13 g l(-1) N) initial NaNO(3) or (NH(4))(2)SO(4) levels contained pullulan, with maltotriose as its major component, similar to that synthesized in the airlift reactor with high (0.78 g l(-1) N) initial NaNO(3) levels. EPS produced by CSTR grown cultures with high (NH(4))(2)SO(4) levels contained little pullulan, possibly because of a population shift from unicells to mycelium. This chemical difference may explain why total EPS yields did not fall as they did with cultures grown under identical conditions with high NaNO(3) levels, where the pullulan component of the EPS disappeared. EPS synthesized in N-limiting chemostat cultures of A. pullulans changed little with growth rate or N source, being predominantly pullulan consisting of maltotriose units. CONCLUSIONS: While the EPS chemical composition changed little under N-limiting conditions, high initial medium N levels determined maltotriose content and/or pullulan content possibly by dictating culture morphology. SIGNIFICANCE AND IMPACT OF THE STUDY: These results emphasize the requirement of all studies to determine EPS chemical composition when examining the influence of culture conditions on EPS yields.