Incorporation of 2-hydroxyl fatty acids by Acinetobacter calcoaceticus RAG-1 to tailor emulsan structure

Acinetobacter calcoaceticus RAG-1 was cultured on different chain length saturated 2-hydroxyl fatty acid (2-HOFA) carbon sources as follows: C12:0 (2-OH), C14:0 (2-OH), C16:0 (2-OH) and C18:0 (2-OH). These 2-HOFAs were used as either sole carbon sources or cosubstrates with C14:0 (total 1% w/v) to form new emulsans (EMs) having controlled side chain FA structure and, therefore, unique emulsifier characteristics. EM yields and cell dry weights ranged from 0.6 to 1.8 g/l and 0.9 to 3.9 g/l, respectively, depending on the carbon source(s) and the cultivation conditions. The content of C12:0 (2-OH) EM substituents reached high levels (306 nmol/mg-EM, 64.4 mol% of total FAs) by selectively feeding this FA. Substantial quantities of 2-HOFAs with chain lengths > or = C14-up to 96 nmo1/mg-EM or 15.2 mol% for C16:0 (2-OH)-were also incorporated in EMs by providing the corresponding 2-HOFA carbon source in the medium. By increasing the medium 2-HOFA concentration large increases in EM total FA contents resulted. The EM FA content was as high as 955 nmol/mg-EM or 23 wt% for a culture containing 0.75 g/100 ml C18:0 (2-OH). An important metabolic pathway involved in EM side chain formation from C16:0 (2-OH) and C18:0 (2-OH) involves decarboxylation, oxidation of the alkanol to the corresponding n-1 FA-CoA intermediate and formation of odd chain length substituent side chain linkages by an EM acyl transferase. Addition of the enzyme alkylating agent iodoacetamide to cultures was used to: (i) enhance the incorporation into EMs of both C12:0 (2-OH) and C16:0 (2-OH) substituents; and (ii) increase by 1.3 to 1.8 fold (by wt.) the total EM FA content. Finally, it was concluded that enhanced emulsification activity of EMs is not necessarily achieved by forming products with relatively high 2- and 3-hydroxydodecanoic acid contents.