The changes in content of endogenous hormones in stolons and tubers of Jerusalem artichoke (Helianthus tuberosus L.) regulate tuber growth, but the specific knowledge about the importance of balance among the endogenous hormones is lacking. Two varieties of Jerusalem artichoke (NY-1 and QY-2) were tested for the endogenous zeatin (ZT), auxins (IAA), gibberellins (GA3) and abscisic acid (ABA) in regulating sugar and dry matter accumulation in tubers. The dry matter content and sugar accumulation in tubers were correlated positively with endogenous ZT and negatively with GA3 content and GA3/ABA and IAA/ABA content ratios. Throughout the tuber formation, ZT content was higher in NY-1 than QY-2 tubers, whereas ABA content was higher in QY-2 than NY-1 tubers. The content ratios GA3/ABA and IAA/ABA were greater in NY-1 than QY-2 before tuber initiation, but QY-2 surpassed NY-1 during the tuber growth stage. The GA3/ABA and IAA/ABA content ratios declined during tuber growth. The results suggested that a dynamic balance of endogenous hormones played an important role in tuber development.