Aims: To analyse the exopolysaccharide ( EPS) production by Lactobacillus helveticus ATCC 15807 in a chemically defined medium ( CDM) and the effect of nutrients and stress culture conditions on cell growth and EPS formation. Methods and Results: Cultures were conducted in CDM: ( i) containing essential and nonessential bases and vitamins; ( ii) without nonessential bases and vitamins [ Simplified CDM ( SCDM)]; ( iii) SCDM supplemented individually with vitamins and bases. The influence of carbohydrates, pH and osmotic culture conditions on growth and polymer formation was analysed. Adenine and lactose stimulated both growth and EPS production. Constant pH fermentations ( 4.5 and 6.2) did not improve EPS synthesis while NaCl and glycerol were detrimental for growth and polymer formation. In all media the EPS monomer composition was glucose and galactose ( 2.5 : 1). Conclusions: A SCDM containing adenine and lactose was optimal for cell growth and EPS formation by Lact. helveticus ATCC 15807. Controlled pH ( 6.2 and 4.5) and osmotic stress culture conditions did not improve polymer production. The EPS characteristics were identical in all media. Significance and Impact of the Study: This work provides a better knowledge on EPS synthesis by Lact. helveticus. A CDM to perform regulation studies on EPS production by Lact. helveticus species is now available.