Axillary buds, collected from greenhouse-grown plants of Bambusa vulgaris Schrad. ex Wendl (B. vulgaris), were incubated on a static liquid culture medium, Murashige and Skoog (MS) medium with 2% (w/v) sucrose, and supplemented with 12.0 µM 6-benzyladenine (BA). They were transferred to a temporary immersion system (TIS) using liquid MS medium supplemented with 0 (CK-free medium), 6.0, 12.0, 18.0 µM BA. The morphological and anatomical indicators were measured. The BA influenced in vitro multiplication of B. vulgaris. The best results were achieved in the SIT with a concentration of 6.0 µM of BA, which increased the number of shoots (5.1 shoots/explant) in the absence of hyperhydric shoots. Results demonstrated that the water content in the sprouts increased with 12.0 and 18.0 µM BA every four hours. Furthermore, these high levels of BA contributed to a lower accumulation of phenolic compounds and lignin content. The total chlorophyll significantly increased when using 6.0 uM BA, but decreased both parameters with other treatments. These results favor to increase the number of shoots/explants during in vitro multiplication. They will also optimize the in vitro culture conditions, leading to an improvement of in vitro propagation methods for this species.