Erika C.G. Aguieiras

From structure to catalysis: recent developments in the biotechnological applications of lipases.

Microbial lipases are highly appreciated as biocatalysts due to their peculiar characteristics such as the ability to utilize a wide range of substrates, high activity and stability in organic solvents, and regio- and/or enantioselectivity. These enzymes are currently being applied in a variety of biotechnological processes, including detergent preparation, cosmetics and paper production, food processing, biodiesel and biopolymer synthesis, and the biocatalytic resolution of pharmaceutical derivatives, esters, and amino acids. However, in certain segments of industry, the use of lipases is still limited by their high cost. Thus, there is a great interest in obtaining low-cost, highly active, and stable lipases that can be applied in several different industrial branches. Currently, the design of specific enzymes for each type of process has been used as an important tool to address the limitations of natural enzymes. Nowadays, it is possible to “order” a “customized” enzyme that has ideal properties for the development of the desired bioprocess. This review aims to compile recent advances in the biotechnological application of lipases focusing on various methods of enzyme improvement, such as protein engineering (directed evolution and rational design), as well as the use of structural data for rational modification of lipases in order to create higher active and selective biocatalysts.

Estolides Synthesis Catalyzed by Immobilized Lipases

Estolides are vegetable-oil-based lubricants obtained from oleic acid or any source of hydroxy fatty acids. In this work, the estolides synthesis from oleic acid and methyl ricinoleate (biodiesel from castor oil), using immobilized commercial lipases (Novozym 435, Lipozyme RM-IM, and Lipozyme TL-IM) in a solvent-free medium was investigated. Acid value was used to monitor the reaction progress by determining the consumption of acid present in the medium. Novozym 435 showed the best performance. Water removal improved the conversion. Novozym 435 was more active at atmospheric pressure. Novozym 435 was reused four times with conversion reaching 15% after the fourth reaction at 80°C. Estolides produced under the reaction conditions used in this work presented good properties, such as, low temperature properties as pour point (−24°C), viscosity (23.9 cSt at 40°C and 5.2 cSt at 100°C), and viscosity index (153).

Estudo do comportamento da lipase comercial Lipozyme RM IM em reações de esterificação para obtenção de biodiesel

The aim of this work was to study monoalkyl ester synthesis catalyzed by immobilized lipase Lipozyme RM IM via the esterification reaction. Yields of over 90% were obtained with butanol in esterification reactions with oleic acid. In the reactions with deodorizer distillates of vegetable oils and butanol, the conversion obtained was greater than 80% after 2.5 h. For the esterification reaction of palm fatty acid deodorizer distillate (PFAD) and butanol, seven reuse cycles of Lipozyme RM IM were carried out and the final conversion was 42% lower than the initial conversion.

Production of wax esters via microbial oil synthesis from food industry waste and by-product streams

The production of wax esters using microbial oils was demonstrated in this study. Microbial oils produced from food waste and by-product streams by three oleaginous yeasts were converted into wax esters via enzymatic catalysis. Palm oil was initially used to evaluate the influence of temperature and enzyme activity on wax ester synthesis catalysed by Novozyme 435 and Lipozyme lipases using cetyl, oleyl and behenyl alcohols. The highest conversion yields (up to 79.6%) were achieved using 4U/g of Novozyme 435 at 70°C. Transesterification of microbial oils to behenyl and cetyl esters was achieved at conversion yields up to 87.3% and 69.1%, respectively. Novozyme 435 was efficiently reused for six and three cycles during palm esters and microbial esters synthesis, respectively. The physicochemical properties of microbial oil derived behenyl esters were comparable to natural waxes. Wax esters from microbial oils have potential applications in cosmetics, chemical and food industries.

Chapter 8 – Solid-State Fermentation for the Production of Lipases for Environmental and Biodiesel Applications

Abstract This chapter reviews the current state-of-art and recent process developments in solid-state fermentation (SSF) for the production of lipases, focusing on its use for biodiesel purposes. Some important parameters for the production of lipase by SSF such as the choice of microorganisms and substrates are discussed, and several applications of the biocatalysts produced are outlined. The recent technology of simultaneous lipase production and immobilization by SSF is addressed, and the recent advances in the application of fermented solids with lipase activity for biodiesel synthesis are highlighted. Moreover, some variables that significantly affect the enzymatic processing biodiesel synthesis and the overall cost of biodiesel (i.e., raw material and alcohols, solvents, water content, and reactor configuration) are also pointed out. This chapter also addresses the application of lipase by SSF in the treatment of high-fat wastewater from the food industry. The generation of this type of wastewater is growing worldwide due to the increase in population and food production. The replacement of dissolved air flotation, a unit typically used in the conventional treatment of these wastewaters, by an enzymatic hydrolysis tank results in a lower fixed capital investment, eliminates a number of operational problems in the bioreactors, and allows greater energy recovery (of the fat contained in the wastewater) in the anaerobic biological treatment employed in the food industries.

Bioprocess development for biolubricant production using microbial oil derived via fermentation from confectionery industry wastes

Microbial oil produced from confectionery and wheat milling side streams has been evaluated as novel feedstock for biolubricant production. Nutrient-rich fermentation media were produced by a two-step bioprocess involving crude enzyme production by solid state fermentation followed by enzymatic hydrolysis of confectionery industry waste. Among 5 yeast strains and 2 fungal strains cultivated on the crude hydrolysate, Rhodosporidium toruloides and Cryptococcus curvatus were selected for further evaluation for biolubricant production based on fermentation efficiency and fatty acid composition. The extracted microbial oils were enzymatically hydrolysed and the free fatty acids were esterified by Lipomod 34-MDP in a solvent-free system with trimethylolpropane (TMP) and neopentyl glycol (NPG). The highest conversion yields were 88% and 82.7% for NPG esters of R. toruloides and C. curvatus, respectively. This study also demonstrates that NPG esters produced from microbial oil have promising physicochemical properties for bio-based lubricant formulations that could substitute for conventional lubricants.

Productive Chain of Biofuels and Industrial Biocatalysis: Two Important Opportunities for Brazilian Sustainable Development

Abstract The integration between biofuels and chemicals production from biomass stimulates the transition to the inevitable bioeconomy era; this era can be achieved by implementing new technologies in existing industrial units where waste streams and by-products can be used as a renewable source of raw materials for the production of commodities and other value-added chemicals. This synergistic approach requires less capital investment, creates new business and job opportunities, expands the market and reduces the environmental impact caused by the operation of industrial plants. This chapter depicts the current situation of the two main biofuels in Brazil, ethanol and biodiesel, and introduces the discussion of opportunities and bottlenecks in the exploitation of lignocellulosic and oleaginous materials, focusing on the important role of enzymatic and microbial processes to support a sustainable industry.