Process development for bioethanol production using wheat straw biomass

Tese de doutoramento em Engenharia Química e BiológicaThis thesis is focused on the process development for bioethanol production based in the biorefinery concept. Nowadays, the role of biorefineries is centred on integration of traditional and modem processes for utilization of biological raw material in the production of energy and chemicals. Lignocellulosic materials such as wheat straw are renewable biomass than can be used as substrate in fermentation processes. The use of wheat straw includes the following steps: milling, pretreatment, enzymatic saccharification and bioconversion of the obtained sugars in bioethanol. In a first stage, autohydrolysis pretreatment was used for solubilization of hemicellulose. In this part, the evaluation of autohydrolysis conditions and determination of their mono saccharide composition in the liquid phase was studied. The results showed the importance of the variation of the particle size distribution on the extraction of total sugars from hemicellulose (glucose, xylose and arabinose), demonstrating that the use of a blend with defined percentages of the various particle sizes is an important parameter to establish before carrying out a pretreatment. In a second stage, the pretreated solids obtained after autohydrolysis were subjected to delignification using organosolv process. A high purity lignin was obtained after the sequential autohydrolysis-organosoiv process. Following the biorefinery philosophy, the autohydrolysis hemicellulose extracted by autohydrolysis optimum conditions was used as reinforcements in a k-car/ LBG polymeric matrix for biocomposite manufacturing. Biocomposite from extracted hemicellulose showed to be a good material to be used in the reinforcement of edible films. The susceptibility to enzymatic saccharifiction of pretreated solids obtained by autohydrolysis and by sequential autohydrolysis-organosolv was studied. According to the obtained results, enzymatic saccharification of the autohydrolysis pretreated solids proved to be more effective than when the organosolv pretreated solids were used. The maximum extent of the enzymatic conversion of cellulose to glucose was 90.88 % and 64.04 %, respectively. These results demonstrate that the autohydrolysis pretreated solids are a good substrate for simultaneous saccharification and fermentation. The simultaneous saccharification and fermentation (SSF) of lignocellulosic materials requires the utilization of mcroorganisms capable of working at high temperatures. In this part of the work, a flocculant Saccharomyces cerevisiae CA1 1 was evaluated for its ability to grow and ferment glucose in the temperature range of 40 - 50 °C. The highest ethanol concentrations obtained were 24.12 and 24.38 gfL at 40°C and 45 °C, respectively, for an initial glucose concentration of 50 gfL. The results indicate that the S. cerevisiae CA1 1 was able to produce ethanol, showing a good performance at high temperatures. In order to evaluate the effects of temperature, substrate concentration (autohydrolysis pretreated wheat straw) and enzyme loading on: 1) ethanol conversion yield, 2) ethanol concentration, and 3) CO2 concentration, a central composite design (CCD) was used. Results showed that the ethanol conversion yield was mainly affected by enzyme loading, whereas for ethanol and CO2 concentration, enzyme loading and substrate concentration were found to be the most significant parameters. The maximum ethanol concentrations (14.84 gfL) were obtained at 45 °C, 3 % (w/v) substrate and 30 FPU of enzyme loading, corresponding to an ethanol yield of 82.4 %. The scale-up of SSF was performed evaluating the effect of stirring on ethanol yield. When the SSF was conducted at higher stirring speed (250 rpm), 15.09 gfL of ethanol was obtained. This corresponds to an overall ethanol yield of 84.49 %. Overall, results presented in this work, describe the development of an integrated process based on the biorefinery concept for the production of bioethanol from wheat straw. It is shown that wheat straw pretreated with autohydrolysis process can be used as a substrate for bioethanol production using a flocculent S. cerevisiae strain. Moreover, extracted hemicellulose is applied as reinforcement for edible films and the obtained lignin is a high purity product.Esta tese trata do desenvolvimento de um processo para produção de bioetanol com base no conceito de biorefinaria. Hoje em dia o papel das biorefinarias centra-se na integração de processos tradicionais e modernos para utilização de material biológico bruto na produção de energia e reagentes. Materiais lenhocelulôsicos como a palha de trigo são uma biomassa renovável que pode ser usada corno substrato em processos de fermentação. Este processo consiste em moagem da palha de trigo, pré-tratamento da palha de trigo, sacarificaçäo enzimática e bioconversão dos açúcares obtidos em bioetanol. Inicialmente o tratamento de auto-hidrólise foi usado para solubilizar a hemicelulose. Nesta parte do trabalho, avaliaram-se as condições de auto-hidrólise e determinaram-se os monossacáridos presentes na fase liquida. Os resultados demonstraram a importância da variação da distribuição do tamanho das partículas na extração dos açúcares totais a partir da hemicelulose (glucose, xilose e arabinose), e a necessidade de estabelecer a composição da mistura a utilizar antes de proceder a um pré-tratarnento. Os sólidos pré-tratados por auto¬hidrólise foram recuperados e deslenhificados através do processo organosolv, tendo sido obtida urna lenhina de elevada pureza. Corn base no conceito de biorefinaria, a hemicelulose extraída por auto-hidrólise sob condições ótimas foi incorporada como reforço para uma matriz polirnénca de k-carl LBG, sendo obtido um biocompósito que demonstrou ser um bom material para ser usado no reforço de filmes edíveis. A suscetibilidade à sacarificação enzimática dos sólidos pré-tratados obtidos por auto¬-hidrólise e urna sequência de auto-hidrólise-organosolv foi estudada. Os resultados obtidos demonstram que a sacarificação enzimática é mais eficiente quando se usaram sólidos pré-tratados sé por auto-hidrólise. Os valores máximos de conversão enzimática de celulose em glucose foram de 90.88 % e de 64.04 %, para sólidos tratados por auto¬-hidrólise e pela sequência auto-hidrólise-organosolv, respectivamente. O processo de sacarificação e fermentação simultânea (SFS) de materiais lenhocelulâsicos requer o uso de microrganismos capazes de trabalhar a elevadas temperaturas. Nesta parte do trabalho, uma estirpe floculante de Saccharomyces cerevisiae CA11 foi utilizada pela sua capacidade de crescer e fermentar glucose numa gama de temperaturas de 40 - 50 °C. As produções máximas de etanol obtidas a partir de 50 g/L glucose foram de 24.12 e 24.38 g/L a 40 °C e 45 °C, respectivamente. Os resultados indicam que as células de S. cerevisiae CA1 i forem capazes de produzir etanol, demonstrando um bom desempenho a altas temperaturas. Para avaliar os efeitos da temperatura, concentração de substrato (palha de trigo pré tratada com auto-hidrólise) e carga de enzima em: 1) rendimento de conversão em etanol, 2) concentração de etanol, e 3) concentração de CO2, usou-se um desenho experimental do tipo "central composite design" (DCC). Os resultados demonstraram que o rendimento de conversão em bioetanol foi principalmente afetado pela carga de enzima, enquanto a concentração de etanol e de CO2, carga de enzima e concentração de substrato foram considerados os parâmetros mais significativos. A concentração máxima de etanol (14.84 giL) foi obtida a 45 °C, 3 % (piv) de substrato e 30 FPU de carga enzimática, correspondendo a um rendimento em etanol de 2.4 %. O aumento de escala do SFS foi realizado avaliando o efeito da agitação no rendimento de etanol. Quando o SFS foi efetuado a velocidade de agitação mais elevada (250 rpm), obtiveram-se 15.09 g/L de etanol. Este valor corresponde a um rendimento global em etanol de 84.49 %. Globalmente, os resultados apresentados neste trabalho descrevem o desenvolvimento de um processo integrado baseado no conceito de biorefinaria para a produção de etanol a partir da palha de trigo. Demonstrou-se que a palha de trigo tratada por auto-hidrólise pode ser usada como substrato para a produção de bioetanol utilizando uma estirpe floculante de Saccharomyces cerevisiae. Demonstrou-se também a possibilidade de utilizar a hemicelulose extraída como reforço em filmes edíveis e a alta pureza da lenhina obtida

A new approach on brewer's spent grains treatment and potential use as lignocellulosic yeast cells carriers

The major objective of this work is to improve the pretreatments of brewer’s spent grains (BSG) aiming at their use as a source for lignocellulosic yeast carriers (LCYC) production. Therefore, several pretreatments of BSG have been designed aiming at obtaining various yeast carriers, differing on their physicochemical composition. Cellulose, hemicellulose, lignin, fat, protein, and ash content were determined for crude BSG and the LCYCs. The long chain fatty acids profile for the crude BSG was also analyzed. Chemical treatments successfully produced several different LCYC based on BSG. The highest cellulose content in LCYC was achieved upon application of caustic (NaOH) treatment during 40 min. Either caustic or combined acid−caustic treatments predominately generated hydrophobic, negatively charged LCYC. The feasibility of using BSG for LCYC production is strengthened by the fact that added-value byproduct can be extracted before the chemical treatments are applied.The brewer’s spent grain supplied by UNICER Bebidas de Portugal, S.A. (S. Mamede de Infesta, Portugal) is gratefully acknowledged

Industrial robust yeast isolates with great potential for fermentation of lignocellulosic biomass

The search of robust microorganisms is essential to design sustainable processes of second generation bioethanol. Yeast strains isolated from industrial environments are generally recognised to present an increased stress tolerance but no specific information is available on their tolerance towards inhibitors that come from the pretreatment of lignocellulosic materials. In this work, a strategy for the selection of different yeasts using hydrothermal hydrolysate from Eucalyptus globulus wood, containing different concentrations of inhibitors, was developed. Ten Saccharomyces cerevisiae and four Kluyveromyces marxianus strains isolated from industrial environments and four laboratory background strains were evaluated. Interestingly, a correlation between final ethanol titer and percentage of furfural detoxification was observed. The results presented here highlight industrial distillery environments as a remarkable source of efficient yeast strains for lignocellulosic fermentation processes. Selected strains were able to resourcefully degrade furfural and HMF inhibitors, producing 0.8 g ethanol/Lh corresponding to 94% of the theoretical yield.The authors thank Rosane Schwan (Federal University of Lavras, Brazil) for kindly providing the yeast strains (isolated from Brazilian ''cachaca'' and ''bio-ethanol'' fermentations) and Juan Carlos Parajo and Gil Garrote (University of Vigo, Spain) for assistance in the pre-treatment of lignocellulose biomass. Research described in this article was financially supported by FEDER funds of EU and ''Fundacao para a Ciencia e a Tecnologia'' (FCT), Portugal: Contract PTDC/BIO/66151/2006, Strategic Project PEst-OE/EQB/LA0023/2013, Project ''BioInd - Biotechnology and Bioengineering for improved Industrial and Agro-Food processes, REF. NORTE-07-0124FEDER-000028'' Co-funded by the Programa Operacional Regional do Norte (ON.2 - O Novo Norte), QREN, FEDER and the PhD grant to FP and Pos-Doc grant to AR

Evaluation of autohydrolysis pretreatment using microwave heating for enzymatic saccharification of corn residues

Pretreatment of lignocellulosic materials (LCMs) is one of the most critical stages in the production of 2G bioethanol, this stage allows to maximize the production of fermentable sugars in the enzymatic saccharification process (ESP). Recently the microwave heating (MH) have been studied for enhanced the LCMs pretreatment, this technology reduces the energy requirements in the process, due to the fast heat transfer and it has allowed to redefine a lot of reactions which the thermal factor plays an essential role in the process. In this work were evaluated the effects of autohydrolysis pretreatment from corn residues using microwave heating and the pretreated solids as substrate in the enzymatic saccharification. The autohydrolysis pretreatment was performed using water as catalyst, the time (10, 30 and 50) and temperature (160, 180 and 200 ºC) were evaluated and the pretreated solids were used in the ESP. The enzymatic saccharification were performed with a working volume of 50 mL, 50 mM citrate buffer (pH 4.8), 2% (w/v) sodium with a cellulose concentration of 1 % (w/v) and incubated at 50 °C. The CellicCTec2 - cellulase was used with a loading of 20 FPU/g. This work showed that microwave autohydrolysis processing is an efficient pretreatment producing a solid enriched with cellulose (63.67±0.91) . The solid pretreated at 200 °C for 10 min was the best condition for saccharification yield (96.95% ± 0.79). This autohydrolysis pretreatment using microwave heating and enzymatic saccharification is a good alternative to obtain fermentable sugars for bioethanol production

Multi-step approach to add value to corncob: production of biomass-degrading enzymes, lignin and fermentable sugars

This work presents an integrated and multi-step approach for the recovery and/or application of the lignocellulosic fractions from corncob in the production of high value added compounds as xylo-oligosaccharides, enzymes, fermentable sugars, and lignin in terms of biorefinery concept. For that, liquid hot water followed by enzymatic hydrolysis were used. Liquid hot water was performed using different residence times (1050 minutes) and holding temperature (180200 °C), corresponding to severities (log(R0)) of 3.364.64. The most severe conditions showed higher xylo-oligosaccharides extraction (maximum of 93%) into the hydrolysates and higher recovery of cellulose on pretreated solids (maximum of 65%). Subsequently, hydrolysates and solids were used in the production of xylanases and cellulases, respectively, as well as, pretreated solids were also subjected to enzymatic hydrolysis for the recovery of lignin and fermentable sugars from cellulose. Maximum glucose yield (100%) was achieved for solids pretreated at log(R0) of 4.42 and 5% solid loading.Michele Michelin is a recipient of a FCT fellowship (SFRH/BPD/ 100786/2014). ThisstudywassupportedbythePortugueseFoundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-010145-FEDER-006684) and BioTecNorte operation (NORTE-01-0145FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 – Programa Operacional Regional do Norte. Héctor Ruiz would like to thank the financial support to the Mexican Science and Technology Council (CONACYT, Mexico) for the Basic Science Project-2015-01 (Ref. 254808) and the Energy Sustainability Fund 2014-05 (CONACYT-SENER), Mexican Centre for Innovation in Bioenergy (Cemie-Bio), and Cluster of Bioalcohols (Ref. 249564). We thank Dr. Nelson Lima from MUM (Micoteca da Universidade do Minho, PT) that gently provided the Trichoderma reesei fungi.info:eu-repo/semantics/publishedVersio

Hydrothermal processing, as an alternative for upgrading agriculture residues and marine biomass according to the biorefinery concept : a review

The concept of a biorefinery that integrates processes and technologies for biomass conversion demands efficient utilization of all components. Hydrothermal processing is a potential clean technology to convert raw materials such as lignocellulosic materials and aquatic biomass into bioenergy and high added-value chemicals. In this technology, water at high temperatures and pressures is applied for hydrolysis, extraction and structural modification of materials. This review is focused on providing an updated overview on the fundamentals, modelling, separation and applications of the main components of lignocellulosic materials and conversion of aquatic biomass (macro- and micro- algae) into value-added products.The authors Hector A. Ruiz and Bruno D. Fernandes thank to the Portuguese Foundation for Science and Technology (FCT, Portugal) for their fellowships (grant number: SFRH/BPD/77361/2011 and SFRH/BD/44724/2008, respectively) and Rosa M. Rodriguez-Jasso thanks to MexicanScience and Technology Council (CONACYT, Mexico) for PhD fellowship support (grant number: 206607/230415)

Enhancement and modeling of enzymatic hydrolysis on cellulose from Agave bagasse hydrothermally pretreated in a horizontal bioreactor

One of the major challenges in biofuels production from lignocellulosic biomass is the generation of high glucose titers from cellulose in the enzymatic hydrolysis stage of pretreated biomass to guarantee a cost-effective process. Therefore, the enzymatic saccharification on cellulose at high solid loading is an alternative. In this work, the agave bagasse was hydrothermally pretreated and optimized at 194°C/30min, obtaining a pretreated solid rich in cellulose content (>46.46%), and subjected to enzymatic hydrolysis at high solid levels. A horizontal bioreactor was designed for enzyme saccharification at high solid loadings [25% (w/v)]. The bioreactor improved mixing efficiency, with cellulose conversions up to 98% (195.6g/L at 72h). Moreover, mathematical modeling of cellulase deactivation demonstrated that cellulases lose most of their initial activity in the first hours of the reaction. Also, cellulose was characterized by X-ray diffraction, and the pretreated solids were visualized using scanning electron microscopy.This project was funded by the Secretary of Public Education of Mexico - Mexican Science and Technology Council (SEP-CONACYT) with the Basic Science Project-2015-01 (Ref. 254808). Marcela Sofía Pino also thanks the National Council for Science and Technology (CONACYT, Mexico) for her Master Fellowship support (grant number: 611312/452636), and Dr. Michele Michelin thanks the Portuguese Foundation for Science and Technology (FCT) for her postdoctoral fellowship (SFRH/BPD/100786/2014).info:eu-repo/semantics/publishedVersio

Clinical audit of multidisciplinary care at a medium-sized hospital in Spain

Background. Multidisciplinary care is a key enabler in the provision of high quality care for cancer patients. Despite compelling evidence supporting their benefit to patients and for providers, multidisciplinary cancer conferences (MCC) are not universally occurring. Team composition of MCC reflects the multidisciplinary nature of the body. Lack of nursing input can have a negative impact on team decision making. The objective of this study was to evaluate multidisciplinary care and adherence to national recommendations at a medium-sized hospital through a clinical audit of cancer conferences and clinical records. Methods. A total of 77 multidisciplinary cancer conferences were visited and 496 electronic health records were reviewed. The regularity of meetings and multidisciplinary attendance were evaluated. Each electronic health record was checked to verify documented prospective discussion before any treatment was started. Results. Nine multidisciplinary teams meet on a weekly or biweekly basis at the hospital with an average number of ten people and six different specialties represented. Average duration of meetings was 46.8 min. Though most patients (64.5%) were discussed at some point at the relevant cancer conference, only 40% had a documented multidisciplinary team discussion prior to the first treatment. Pathological stage (pTNM) was documented in 53.6% of clinical records. Conclusions. Nursing representatives should be included as usual attendees at cancer conferences. Prospective discussion of all cancer cases should be encouraged. Use of checklists and systematic collection of key information, specifically cancer staging, could improve clinical documentation in the electronic clinical record