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Study of fungal enzymatic efficiency in transesterification and hydrolysis reactions from native vegetable oils for biotechnological applications

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Mariana de Souza Rocha
Total Authors: 1
Document type: Master's Dissertation
Press: Ribeirão Preto.
Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
Defense date:
Examining board members:
Maria de Lourdes Teixeira de Moraes Polizeli; Hamilton Cabral; Anizio Marcio de Faria; María Eugenia Guazzaroni
Advisor: Maria de Lourdes Teixeira de Moraes Polizeli; Ana Claudia Vici

The present work aimed to identify the best fungal producers of lipases with efficient capacity of transesterification and hydrolysis reactions. The fungi were selected from the Library of Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto/USP, located in the Laboratory of Microbiology and Cell Biology for the study of lipase activity in its free and immobilized form, with the objective of synthesizing products for biotechnological applications. The screening stage was concluded with the selection of the thermophilic fungi Scytalidium thermophilum and Humicola grisea and the mesophilic fungus Aspergillus phoenicis for presenting the best rates of enzymatic activity and production of extracellular protein. The best culture conditions were standardized as: SR culture medium (Segato-Rizzatti) added of 1% sunflower oil, buriti and soybean for A. phoenicis, S. thermophilum and H. grisea, respectively. After the verification of low thermical stability of the free lipase produced by A. phoenicis and S. thermophilum, it was used the immobilization technique on Octyl-sepharose, Butylsepharose, C-18 Sepabeads, PEI-agarose, Duolite and MANAE-agarose. For S. thermophilum lipases immobilized on Sepabeads C-18 (retention 98.59%) and Duolite (retention 98.73%) better stability was observed at 50 ° C and at pH 6 and 7. The Duolite derivative was very stable in the solvents cyclohexane, ethanol and acetone, maintaining residual activity above 70% after 24 hours of incubation. For A. phoenicis lipases, the Octyl-sepharose support (retention 98.83%) was satisfactorily stable at 50 ° C and pH 6, plus at cyclohexane and hexane solvents solutions. In addition, the immobilization of A. phoenicis lipase on Octyl-sepharose led to enzymatic hyperactivation. Through a screening for the transesterification reaction, the Duolite derivative for the S. thermophilum lipases was selected for the continuation of the studies for biodiesel production. Ethanol was used for the transesterification tests due to its sustainability characteristics compared to methanol. Following the methodology of ethanol:sunflower oil (1:2) added with 10% (w/v oil) of Duolite derivative and 82% of cyclohexane, the total ethyl ester content produced was 53.78%. The biodiesel produced was submitted to physic-chemical analysis and the following values were obtained: acid index of 0.56 mg NaOH/g and iodine content index of 42.3 g I2/100 g of biodiesel sample, demonstrating the aptness of the product in the current legislation that determines the biofuel quality. The yield of the transesterification using sunflower oil as substrate was 74%. The lipase of A. phoenicis immobilized on Octyl-sepharose was used for the hydrolysis reaction of soybean, buriti, açaí, pracaxi and ojon oils and tucumã butter. The concentration of fatty acids released was quantified by the sulphophosphorus-vanillin colorimetric method, resulting in the hydrolysis of buriti (3.4 mg/mL), açai (3.06 mg/mL) and ojon (2.27 mg/mL) oils in total fatty acids content. The derivatives developed in this work have interesting characteristics for the development of bioprocesses to be applied in the industry. (AU)

FAPESP's process: 17/25011-6 - Study of fungal enzymatic efficiency in transesterification and hydrolysis reactions from native vegetable oils aiming biotechnological applications
Grantee:Mariana de Souza Rocha
Support Opportunities: Scholarships in Brazil - Master