Exploring Design Space and Optimization of nutrient factors for maximizing lipid production in Metchnikowia pulcherrima with Design of Experiments

Due to the importance of unsaturated fatty acids for human health and the increasing global demand in the food and food crop area, oleaginous yeasts are promising alternative microorganisms for commercial lipid production due to the high volumetric productivity, with Metchnikowia pulcherrima being an underexplored oleaginous yeast with potential as a lipid producer. Critical to achieving high productivity lipid production are nutrient factors. A sensitivity test identified carbon and nitrogen sources as important factors in nitrogen limited broth (NLB) for lipid production in M. pulcherrima i.e. glucose, yeast extract and Ammonium sulphate. Response Surface Methodology (RSM) involving sets of 15 experimental runs of three-factor three-level Box-Behnken Design (BBD) was implemented for exploring the design of the carbon and nitrogen source in the growth media composition. Quadratic surfaces were least-square fitted and used to identify regions of optimal lipid yield. Multiple sets of runs were conducted with the parameter range progressively adapted and repeated until a clear optimum was identified. The highest total lipid production was in the low carbon concentration range (2.27-21.5 g/L) which suggests the process is more productive with the optimised media compared to NLB media (30.4 g/L of carbon). The optimal carbon concentration was 14.8 g/L whereas the dependence on nitrogen was not found to be significant. The yield of optimal point (YP/S) was 2.39 times higher than NLB media after validation because the depletion of glucose at the end of fermentation (72-104 h) contributes the high increase of lipid accumulation (mg Lipid/ 100 ml culture) of the optimal point (94.5%) which was higher than NLB media (34.5%) for 60%.