An efficient protocol for rooting and acclimation of in vitro propagated Ruppia maritima was the focus of this research. The effects of four dilutions of a nutrient medium (f) and four different concentrations of sodium bicarbonate on rooting production were investigated. The optimal rooting medium was determined to be f/2 with 0.25 mM sodium bicarbonate in 5 ppt synthetic seawater at a pH of 7.5. Cultures of R. maritima were found to be photosynthetically inactive as measured by oxygen evolution in sucrose-based multiplication medium but were physiologically capable of adjusting to autotrophism once transferred to a bicarbonate medium. This transition was immediate and comparable to cultures that had acclimated in bicarbonate-based medium for 21 days. Ruppia maritima rooted onto coconut fiber mats and unanchored plants were acclimated in outdoor aquaria during May 1995. The results of these studies showed that micropropagated R. maritima can be acclimated easily and effectively at a low cost in aquaria, making R. maritima a useful seagrass species for restoration and mitigation projects.