Three phase partitioning (TPP), a technique which is based on partitioning of hydrophilic constituents, proteins, and hydrophobic constituents in three phases comprising of water, ammonium sulphate and organic solvent, was explored for extraction of diterpene, forskolin from Coleus forskohlii roots. The process which consists of simultaneous addition of t-butanol and ammonium sulphate to the aqueous slurry of C. forskohlii roots was optimized with respect to the concentration of ammonium sulphate loading and the ratio of t-butanol to slurry. A maximum of 30.83% recovery of forskolin was obtained under the optimized conditions. Ultrasonication and enzyme pretreatment with commercial enzyme preparation of Stargen (R) 002 and Accellerase (R) 1500 followed by TPP gave 79.95% and 83.85% recovery when used individually within 4 h as compared to 12 h in Soxhlet extraction. A combination of the two pretreatments increased the yield marginally. Hence enzymatic pretreatment followed by TPP is recommended for extraction of forskolin. (C) 2012 Elsevier B.V. All rights reserved.