Síntese de biodiesel catalisada por lipase B de Candida antarctica imobilizada em esferas de estireno divinil-benzeno

Trabajo presentado en el XI Seminário Brasileiro de Tecnologia Enzimática (Enzitec), celebrado en Río de Janeiro (Brasil) del 14 al 16 de abril de 2014.O processo de transesterificação de óleos vegetais para a síntese de biodiesel catalisada por substâncias básicas tem sido utilizado largamente em escala industrial e altas conversões são obtidas. Entretanto, uma grande quantidade de água é necessária para a purificação dos ésteres, gerando grandes quantidades de rejeitos aquosos inadequados para descarte, e dessa forma, a utilização de lipases imobilizadas destaca-se como uma alternativa ao método alcalino. Assim, o presente trabalho teve por objetivo estudar a síntese de biodiesel, a partir do óleo de soja, catalisada pela lipase do tipo B de Candida antarctica imobilizada em esferas de estireno divinil-benzeno (MCI GEL CHP20P), denominada CALB-MCI. Um delineamento composto central rotacional (DCCR) e a metodologia de superfície de resposta (MSR) foram utilizados para otimizar os parâmetros razão molar metanol:óleo de soja, teor de enzima imobilizada, e teor de água adicionada nas reações. Após os resultados da otimização, foram realizados testes de estabilidade operacional de CALB-MCI em oitos ciclos de batelada e, além disso, foi avaliado a melhor maneira de recuperar o biocatalizador após cada ciclo testando duas alternativas de tratamento: lavagens com hexano e sem lavagens. Na condição ótima foi possível obter um rendimento de 36,73 %, o que representa uma taxa inicial de reação de 57 mmol L-1 h-. Com relação a estabilidade operacional, as lavagens com hexano demonstraram ser um excelente tratamento, onde o derivado CALB-MCI manteve 75 % de rendimento de conversão relativa após oito bateladas, sugerindo sua aplicabilidade na produção de biodiesel.Peer Reviewe

Enzymatic reactors for biodiesel synthesis: Present status and future prospects

Lipases are being extensively researched for the production of biodiesel as a “silver bullet” in order to avoid the drawbacks of the traditional alkaline transesterification. In this review, we analyzed the main factors involved in the enzymatic synthesis of biodiesel, focusing in the choice of the immobilization protocol, and the parameters involved in the choice and configuration of the reactors. An extensive discussion is presented about the advantages and disadvantages of each type of reactor and their mode of operation. The current scenario of the market for enzymatic biodiesel and some future prospects and necessary developments are also briefly presented.This work was supported by grants from Brazilian Coordenação de Aperfoiçoamento de Pessoal de Nível Superior (CAPES).Peer Reviewe

Combi-lipase for heterogeneus substrates: a new approach for hydrolysis and trasnesterification of vegetable oils using mixtures of biocatalysts

Trabajo presentado en el 7th International Congress on Biocatalysis, celebrado en Hamburgo (Alemania) del 31 de agosto al 04 de septiembre de 2014.Combination of different enzymes is mainly used for cascade or sequent ial reactions, however, in the present work , is being proposed the design of a combi-lipase biocatalyst strategy far the simultaneous hydrolysis ar transesterification of a mixture of different substrates. lt is based on the combination of different lipases as biocatalysts in reactions using heterogeneous substrates , such vegetable oils. The hydrolysis and the transesterification of different vegetable oils (soybean oil, olive oil and palm oil) were evaluated, and Novozym 435 (CALB), Lipozyme TL-IM (TLL), and Lipozyme RM-IM (RML) were used as biocatalysts. For each reaction and oil, it was evaluated the best mixture of lipase, and thereafter the reaction parameters (substrate molar ratio, temperature and biocatalyst content) were optimized using an experimental design. Results showed that, although individually TLL was the most active enzyme, the combination of different lipases presented better effects. For hydrolysis of soybean oil, a mixture of 80 % of RML and 20 % of CALB was the best biocatalyst. Reaction parameters were optimized, allowing to obtain more than 80 % of hydrolysis in 24 h using the combi-lipase, up from less than 50 % when any individual lipase. In the transesterification, using olive oil and ethanol the best mixture was composed by 30 % TLL, 12.5 % of RML and 57.5 % of CALB, reaching to 98 % of conversion after 18 h, while using palm oil and ethanol the best mixture was 52.5 % of TLL and 47.5 % of RML, yielding 80 % after 18 h. Reusability of the combi-lipase showed that it could be used for at least 15 cycles without any significant decrease in hydrolysis, and for 7 cycles in transesterificat ion. The concept of combí-biocatalyst might be a useful technology for reactions including full modification of heterogeneous substrates.Peer Reviewe

Combi-lipase for heterogeneous substrates- a new approach for hydrolysis and transesterification using mixtures of biocatalysts

Trabajo presentado en el VII Workshop on Biocatalysis and Biotransformations - 1º Simposio Latinoamericano de Biocatalisis y Biotransformaciones, celebrado en Búzios (Brasil) del 23 al 26 de septiembre de 2014.[Introduction]: Combination of different enzymes is mainly used for cascade or sequential reactions, however, in the present work, is being proposed the design of a combi-lipase biocatalyst strategy for the simultaneous hydrolysis or transesterification of a mixture of different substrates.1,2 It is based on the combination of different lipases as biocatalysts in reactions using heterogeneous substrates, such vegetable oils.[Results and discussion]: The hydrolysis and the transesterification of different vegetable oils (soybean oil, olive oil and palm oil) were evaluated, and Novozym 435 (CALB), Lipozyme TL-IM (TLL), and Lipozyme RM-IM (RML) were used as biocatalysts. For each reaction and oil, it was evaluated the best mixture of lipase, and thereafter the reaction parameters (substrate molar ratio, temperature and biocatalyst content) were optimized using an experimental design. For hydrolysis of soybean oil, a mixture of 80 % of RML and 20 % of CALB was the best biocatalyst. Reaction parameters were optimized, allowing to obtain more than 80 % of hydrolysis in 24 h using the combi-lipase, up from less than 50 % when any individual lipase, as shown in Figure 1. In the transesterification, using olive oil and ethanol the best mixture was composed by 30 % TLL, 12.5 % of RML and 57.5 % of CALB, reaching to 98 % of conversion after 18 h, while using palm oil and ethanol the best mixture was 52.5 % of TLL and 47.5 % of RML, yielding 80 % after 18 h. Reusability of the combi-lipase showed that it could be used for at least 15 cycles without any significant decrease in hydrolysis, and for 7 cycles in transesterification.[Conclusion]: The concept of combi-biocatalyst might be a useful technology for reactions including full modification of heterogeneous substrates. Further studies are granted to test these preparations in continuous flow reactors.This work was supported by grants from FAPERGS and CNPq (Brazil) and CTQ2013-41507-R from Spanish MINECO. The authors would like to thank Mr. Ramiro Martínez (Novozymes, Spain) for kindly supplying the enzymes used in this research.Peer Reviewe

Optimization of synthesis of fatty acid methyl esters catalyzed by lipase B from Candida antarctica immobilized on hydrophobic supports

In this work two immobilized preparations of lipase (EC 3.1.1.3) B from Candida antarctica (CALB) were compared for the synthesis of fatty acid methyl esters (FAME) using soybean oil. Commercial Novozym 435 (CALB-435) and CALB immobilized on styrene-divinylbenzene beads (CALB–MCI) were tested for the transesterification reactions. Central composite design (CCD) and response surface methodology (RSM) were used to optimize the reaction parameters, substrate molar ratio, enzyme content, and the added amount of water, on the initial reaction rate as response. The biocatalysts showed different optimal conditions for the production of FAME. For CALB-435, optima conditions were 5.6:1 molar ratio methanol:oil, 25% enzyme, and 5.44% of added water, while for MCI–CALB, these optima were 3:1 molar ratio methanol:oil, 25% enzyme, and 1.18% of added water, resulting in initial reaction rates of 51.47 mmol L−1 h−1, and 57 mmol L−1 h−1 of FAME, respectively. Conversions of 93.38% using CALB-435, and 99.03% using CALB–MCI were obtained after 72 h of reaction under the optimized conditions. Repeated batches of reaction were carried out to test the operational stability of biocatalysts, with both preparations keeping around 70% of their initial activity after eight batches.This work was supported by grants from Brazilian Coordenação de Aperfoiçoamento de Pessoal de Nível Superior (CAPES) and CTQ2009-07568 from Spanish Ministerio de Ciencia e Innovación.Peer Reviewe

Optimization of ethyl ester production from olive and palm oils using mixtures of immobilized lipases

Although reactions of transesterification are generally catalyzed by one specific lipase preparation, the concept of “combi-lipase” could be better explored for the production of biodiesel, since oils are heterogeneous substrates. In this research, we tested this concept by evaluating the enzymatic transesterification of olive and palm oils, two diverse fatty acid compositions, using standalone or mixtures of three immobilized lipases as biocatalysts: Novozym 435 (CALB), Lipozyme TL-IM (TLL), and Lipozyme RM-IM (RML). For olive oil, the combination of 29.0% of TLL, 12.5% of RML, and 58.5% of CALB was the best, allowing for 95% conversion efficiency in 18 h of reaction, up from 50% for the best individual lipase (CALB). For palm oil, the best enzyme combination was 52.5% of TLL and 47.5% of RML, resulting in 80% of conversion of ethyl esters in 18 h, compared to only 44% when standalone TLL was used. Repeated batches of reaction were carried out in order to test the operational stability of the combi-lipase systems, with results showing that they could be used for at least seven cycles keeping higher than 80% of their initial activities.This work was supported by grants from Brazilian Coordenação de Aperfoiçoamento de Pessoal de Nível Superior (CAPES) and CTQ2013-41507-R from Spanish MINECO.Peer Reviewe