The production of fatty acids ethyl esters (FAEEs) to be used as biodiesel from oleaginous microalgae shows great opportunities as an attractive source for the production of renewable fuels without competing with human food. To ensure the economic viability and environmental sustainability of the microbial biomass as a raw material, the integration of its production and transformation into the biorefinery concept is required. In the present work, lipids from wet Isochrysis galbana microalga were extracted with ethyl acetate with and without drying the microalgal biomass (dry and wet extraction method, respectively). Then, FAEEs were produced by lipase-catalyzed transesterification and esterification of the extracted lipids with ethanol using lipase B from Candida antarctica (CALB) and Pseudomonas cepacia (PC) lipase supported on SBA-15 mesoporous silica functionalized with amino groups. The conversion to FAEEs with CALB (97 and 85.5 mol% for dry and wet extraction, respectively) and PS (91 and 87 mol%) biocatalysts reached higher values than those obtained with commercial Novozym 435 (75 and 69.5 mol%). Due to the heterogeneous nature of the composition of microalgae lipids, mixtures with different CALB:PC biocatalyst ratio were used to improve conversion of wet-extracted lipids. The results showed that a 25:75 combi-lipase produced a significantly higher conversion to FAEEs (97.2 mol%) than those produced by each biocatalyst independently from wet-extracted lipids and similar ones than those obtained by each lipase from the dry extraction method. Therefore, that optimised combi-lipase biocatalyst, along with achieving the highest conversion to FAEEs, would allow improving viability of a biorefinery since biodiesel production could be performed without the energy-intensive step of biomass drying.

Endopolygalacturonase (EndoPGase) is one of the crucial pectinases belonging to the class of carbohydrase. The catalytic action of EndoPGase captivate the attention for production of this extremely valuable catalyst of industrial sector. The main focus was to ascertain a potential bacterial candidate for endoPGase production. The isolated bacterial strain was further identified by 16S rDNA gene sequencing. A genomic library was constructed by using Lambda ZAP II vector system to investigate the pectinolytic potential of the expressed genes. The parameters for enzyme biosynthesis were optimized by single as well as multiple factor approach at a time. The results of our investigation led to the identification of a potent strain of Bacillus subtilis NR2. The strain was found active for pectic enzyme activity under shaking- flask fermentation at pH 5.0 and 50 °C temperature of incubation. Among all monomeric and polymeric substrates, citrus pectin followed by wheat bran was considered the best enzyme inducers at 1 % concentration. Moreover, an increasing trend in enzyme activity was observed with the increasing inducer concentration. The combined effect of three variables (pH, substrates, and substrate concentration) was explored by response surface methodology involving Box Benken Design (BBD). The study concluded that the soil isolated B. subtilis can be utilized for commercial scale pectinase enzyme production.

When bioprospecting for novel industrial enzymes, substrate promiscuity is a desirable property that increases the reusability of the enzyme. Among industrial enzymes, ester hydrolases have great relevance for which the demand has not ceased to increase. However, the search for new substrate promiscuous ester hydrolases is not trivial since the mechanism behind this property is greatly influenced by the active site's structural and physicochemical characteristics. These characteristics must be computed from the 3D structure, which is rarely available, and expensive to measure, hence the need for a method that can predict promiscuity from a sequence alone. Here we report such a method called EP-pred, an ensemble binary classifier, that combines three machine learning algorithms: SVM, KNN, and a Linear model. EP-pred has been evaluated against the Lipase Engineering Database together with a hidden Markov approach leading to a final set of 10 sequences predicted to encode promiscuous esterases. Experimental results confirmed the validity of our method since all ten proteins were found to exhibit a broad substrate ambiguity.