Zero-waste biorefinery of oleaginous microalgae as promising sources of biofuels and biochemicals through direct transesterification and acid hydrolysis

Oleaginous microalgae are considered as promising sources of biofuels and biochemicals due to their high lipid content and other high-value components such as pigments, carbohydrate and protein. This study aimed to develop an efficient biorefinery process for utilizing all of the components in oleaginous microalgae. Acetone extraction was used to recover microalgal pigments prior to processes for the other products. Microalgal lipids were converted into biodiesel (fatty acid methyl ester, FAME) through a conventional two-step process of lipid extraction followed by transesterification, and alternatively a one-step direct transesterification. The comparable FAME yields from both methods indicate the effectiveness of direct transesterification. The operating parameters for direct transesterification were optimized through response surface methodology (RSM). The maximum FAME yield of 256 g/kg-biomass was achieved when using chloroform:methanol as co-solvents for extracting and reacting reagents at 1.35:1 volumetric ratio, 70 °C reaction temperature, and 120 min reaction time. The carbohydrate content in lipid-free microalgal biomass residues (LMBRs) was subsequently acid hydrolyzed into sugars under optimized conditions from RSM. The maximum sugar yield obtained was 44.8 g/kg-LMBRs and the protein residues were recovered after hydrolysis. This biorefinery process may contribute greatly to zero-waste industrialization of microalgae based biofuels and biochemicals.     

DETERMINING BLUBBER FATTY ACID COMPOSITION: A COMPARISON OF IN SITU DIRECT AND TRADITIONAL METHODS

Fatty acids (FAs) are used to make inferences about the foraging behavior and diets of free-ranging marine mammals. However, several methods are currently available for determining the FA composition of blubber and these methods may produce different results. We compared in situ direct transesterification methods, where a small amount of tissue is sampled, with more traditional methods involving prior lipid extraction of the entire sample of interest. Using gray seal ( Halichoerus grypus ) and beluga whale ( Delphinapterus leucas ) blubber, we found that when the direct in situ method was used on a 2-mg sample of blubber, the resulting FA profile differed significantly from that produced when traditional full-extraction methods were employed. Regardless of where the small spot sample was taken within the blubber depth, it was not representative of the entire blubber FA composition, as blubber is non-homogeneous throughout its depth. We also modified the in situ direct method to allow analysis of the entire blubber layer. Results of this full-layer direct method compared quite favorably with traditional extraction methods and may provide a reasonable alternative for analyses. Although application of our full-layer direct method will require further verification in certain marine mammal blubber samples, we conclude that the large differences obtained when using the direct method are not a consequence of the chemical method itself. Rather, they arise from non-representative sampling of the blubber FA composition.     

Biodiesel fuel production via transesterification of oils using lipase biocatalyst

Biodiesel has gained widespread importance in recent years as an alternative, renewable liquid transportation fuel. It is derived from natural triglycerides in the presence of an alcohol and an alkali catalyst via a transesterification reaction. To date, transesterification based on the use of chemical catalysts has been predominant for biodiesel production at the industrial scale due to its high conversion efficiency at reasonable cost. Recently, biocatalytic transesterification has received considerable attention due to its favorable conversion rate and relatively simple downstream processing demands for the recovery of by-products and purification of biodiesel. Biocatalysis of the transesterification reaction using commercially purified lipase represents a major cost constraint. However, more cost-effective techniques based on the immobilization of both extracellular and intracellular lipases on support materials facilitate the reusability of the catalyst. Other variables, including the presence of alcohol, glycerol and the activity of water can profoundly affect lipase activity and stability during the reaction. This review evaluates the current status for lipase biocatalyst-mediated production of biodiesel, and identifies the key parameters affecting lipase activity and stability. Pioneer studies on reactor-based lipase conversion of triglycerides are presented.     

Chemoenzymatic synthesis of enantiopure 1,4-dihydropyridine derivatives

1,4-Dihydropyridines possess a broad range of biological activities, such as the ability to control the influx of calcium into cells, as well as neuroprotective, antineurodegenerative, cognition and memory enhancing, anti-inflammatory, antiviral and many other properties. Chirality plays an important role in the biological activity of 1,4-dihydropyridines. The chemoenzymatic synthesis of 1,4-dihydropyridine derivatives in enantiopure form as the key intermediates for the synthesis of enantiopure drugs and chiral analogues of symmetrical drugs has become an advantageous alternative to the other synthetic methods. Hydrolytic enzymes, as efficient chemo-, regio- and stereoselective biocatalysts have been successfully applied for the asymmetrisation or kinetic resolution of various 1,4-dihydropyridine derivatives. Several synthetic strategies to overcome the inactivity of hydrolytic enzymes towards 1,4-dihydropyridine carboxylic acids have been developed during the last decade, often based on the introduction of a spacer between an enzymatically labile group and the 1,4-DHP nucleus. Good to excellent enantioselectivities can be obtained by careful optimisation of the reaction temperature and the organic (co)solvent used in the enzymatic transformations.     

Chemical modification of alpha-chymotrypsin for obtaining high transesterification activity in low water organic media

Alpha-chymotrypsin was made more hydrophilic by modifying 11 (out of 16) ε-amino groups with pyromellitic dianhydride. The hydrophilic preparation was precipitated with n-propanol. This preparation gave significantly higher initial rates at the optimum a_w (127.51 nmol mg^?1 min^?1 in n-octane and 21.30 nmol mg^?1 min^?1 in acetonitrile at a_w=0.33) compared with the lyophilized preparation (53.50 nmol mg^?1 min^?1 in n-octane and 0.26 nmol mg^?1 min^?1 in acetonitrile at a_w=0.97). FT-IR showed that the precipitate of modified alpha-chymotrypsin has a higher content of alpha-helices and beta-sheets compared to the lyophilized powder.     

Synthesis of vinyl arabinose ester catalyzed by protease from Streptomyces sp.

The transesterification of 0.5 M divinyladipate with 0.25 M arabinose in dimethylformamide for 7 days was catalyzed by Streptomyces sp. alkaline protease to give 5-O-vinyladipoyl-d-arabinofuranose at ca. 50% yield. Only enzymatic transesterification of primary hydroxyl group of arabinofuranose proceeded without esterification of arabinopyranose.