The aim of the present work was to study the applicability of rotating packed bed (RPB) for biodiesel through the biocatalytic method. In this research, the RPB facilitated a more homogeneous mixture of substrates due to its higher mass transfer efficiency and better micromixing environment. This was superior to the traditional continuous stirred tank reactor (CSTR) system. Candida sp. 99-125 lipase was used without any organic solvent or additive, and demonstrated a significant catalyst efficiency. The key factors, such as the high gravity factor (b), pattern of the catalyst and methanol-FFA molar ratio etc. were investigated. Under the optimal conditions, the hydrolysis yield of fatty acids was 97.0% after 24 h and the esterification yield of biodiesel was 96.0% 6 h later. The esterifying yield didn't have an obvious decline in the fifth batch. Consequently, the RPB is an attractive and effective reactor for enzymatic synthesis. (C) 2016 Elsevier Ltd. All rights reserved.