Camila Florencio

Bio: Camila Florencio is an academic researcher from Empresa Brasileira de Pesquisa Agropecuária. The author has contributed to research in topics: Bagasse & Trichoderma reesei. The author has an hindex of 9, co-authored 17 publications receiving 327 citations. Previous affiliations of Camila Florencio include Federal University of São Carlos.

Correlation between agar plate screening and solid-state fermentation for the prediction of cellulase production by Trichoderma strains.

Abstract: The viability of converting biomass into biofuels and chemicals still requires further development towards the reduction of the enzyme production costs. Thus, there is a growing demand for the development of efficient procedures for selection of cellulase-producing microorganisms. This work correlates qualitative screening using agar plate assays with quantitative measurements of cellulase production during cultivation under solid-state fermentation (SSF). The initial screening step consisted of observation of the growth of 78 preselected strains of the genus Trichoderma on plates, using microcrystalline cellulose as carbon source. The 49 strains that were able to grow on this substrate were then subjected to a second screening step using the Congo red test. From this test it was possible to select 10 strains that presented the highest enzymatic indices (EI), with values ranging from 1.51 to 1.90. SSF cultivations using sugarcane bagasse and wheat bran as substrates were performed using selected strains. The CG 104NH strain presented the highest EGase activity (25.93 UI·g−1). The EI results obtained in the screening procedure using plates were compared with cellulase production under SSF. A correlation coefficient () of 0.977 was obtained between the Congo red test and SSF, demonstrating that the two methodologies were in good agreement.

Addition of Soybean Protein Improves Saccharification and Ethanol Production from Hydrothermally Pretreated Sugarcane Bagasse

Abstract: The bioconversion yield of ethanol from lignocellulosic feedstocks is negatively affected by the unproductive adsorption of cellulolytic enzymes onto lignin. In this work, soybean protein was used as a lignin-blocking additive, with the aim of improving the production of ethanol from enzymatic hydrolysates of pretreated sugarcane bagasse. Investigation was made of the effects of the type of hydrothermal pretreatment process—steam explosion (SE) or liquid hot water (LHW), loadings of solids and enzymes, and bioreactor type. The addition of soybean protein led to a exceptional 76% increase of glucose released using the LHW pretreated bagasse, after 24 h of reaction, employing a high-solids loading (15%, w/v) and a low enzyme dosage (5 FPU/g dry biomass). A significant improvement was also achieved for industrial-like mixing conditions in a bench-scale stirred tank reactor, increasing the glucose released by 61 and 42% for the LHW and SE processes, respectively. Ethanol production was also positively affected by the presence of soybean protein, with increases of up to 86 and 65% for the LHW and SE hydrolysates, compared to the control experiment. Characterization of the sugarcane bagasse after the adsorption of soybean protein, using Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM), corroborated the higher affinity of the additive for the LHW bagasse. These findings suggest that soybean protein supplementation during enzymatic hydrolysis by commercially available enzymes is an effective strategy for achieving higher saccharification yields from hydrothermally pretreated biomass, hence improving ethanol production.

Validation of a Novel Sequential Cultivation Method for the Production of Enzymatic Cocktails from Trichoderma Strains

Abstract: The development of new cost-effective bioprocesses for the production of cellulolytic enzymes is needed in order to ensure that the conversion of biomass becomes economically viable. The aim of this study was to determine whether a novel sequential solid-state and submerged fermentation method (SF) could be validated for different strains of the Trichoderma genus. Cultivation of the Trichoderma reesei Rut-C30 reference strain under SF using sugarcane bagasse as substrate was shown to be favorable for endoglucanase (EGase) production, resulting in up to 4.2-fold improvement compared with conventional submerged fermentation. Characterization of the enzymes in terms of the optimum pH and temperature for EGase activity and comparison of the hydrolysis profiles obtained using a synthetic substrate did not reveal any qualitative differences among the different cultivation conditions investigated. However, the thermostability of the EGase was influenced by the type of carbon source and cultivation system. All three strains of Trichoderma tested (T. reesei Rut-C30, Trichoderma harzianum, and Trichoderma sp INPA 666) achieved higher enzymatic productivity when cultivated under SF, hence validating the proposed SF method for use with different Trichoderma strains. The results suggest that this bioprocess configuration is a very promising development for the cellulosic biofuels industry.

Insights from enzymatic degradation of cellulose and hemicellulose to fermentable sugars– a review

Abstract: Lignocellulose, the most abundant and renewable resource on Earth is an important raw material, which can be converted into high value products. However, to this end, it needs to be pretreated physically, chemically, or biologically. Its complex structure and recalcitrance against physical, chemical, or biological degradation render its breakdown an important target of study. The understanding of the enzymatic processes of lignocellulose breakdown and the changes in its chemistry are thus essential. Here, we review the current analytical challenges in the analysis of lignocellulose composition, lignocelluloytic pretreatment, analysis of enzymatic hydrolysis catalyzed by cellulases or hemicellulases and their biotechnological applications. Complex techniques including biochemical, genomic, and metagenomics methods such as high performance anion exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD), Respiration Activity Monitoring System (RAMOS), and next-generation sequencing are described. HPAEC-PAD is a promising, rapid, and reliable analytical technique for sugar quantification following lignocellulose breakdown. RAMOS is an effective technique for monitoring the growth of microorganisms during the different phases of enzyme production, enzymatic hydrolysis, and fermentation. The emergence of high throughput, next-generation sequencing techniques has enriched the databases of genes encoding glycoside hydrolase classes commonly involved in lignocellulose decomposition, and this knowledge can be readily used to analyse the involved processes. Still, novel analytical methods are highly welcome to understand the complete process of lignocelluloytic breakdown. In order to decrease environmental pollution and to save energy, lignocellulose conversion needs to be promoted in order to effectively compete with fossil resources on a global scale in future.

Lignin-enzyme interaction: A roadblock for efficient enzymatic hydrolysis of lignocellulosics

Abstract: Efficiently producing second-generation biofuels from biomass is of strategic significance and meets sustainability targets, but it remains a long-term challenge due to the existence of biomass recalcitrance. Lignin contributes significantly to biomass recalcitrance by physically limiting the access of enzymes to carbohydrates, and this could be partially overcome by applying a pretreatment step to directly target lignin. However, lignin typically cannot be completely removed, and its structure is also significantly altered during the pretreatment. As a result, lignin residue in the pretreated materials still significantly hindered a complete conversion of carbohydrate to its monosugars by interacting with cellulase enzymes. The non-productive adsorption driven by hydrophobic, electrostatic, and/or hydrogen bonding interactions is widely considered as the major mechanism of action governing the unfavored lignin-enzyme interaction. One could argue this type of interaction between lignin residue and the activated enzymes is the major roadblock for efficient enzymatic hydrolysis of pretreated lignocellulosics. To alleviate the negative effects of lignin on enzyme performance, a deep understanding of lignin structural transformation upon different types of pretreatments as well as how and where does lignin bind to enzymes are prerequisites. In the last decade, the progress toward a fundamental understanding of lignin-enzyme interaction, structural characterization of lignin during pretreatment and/or conformation change of enzyme during hydrolysis is resulting in advances in the development of methodologies to mitigate the negative effect of lignin. Here in this review, the lignin structural transformation upon different types of pretreatments and the inhibition mechanism of lignin in the bioconversion of lignocellulose to bioethanol are summarized. Some technologies to minimize the adverse impact of lignin on the enzymatic hydrolysis, including chemical modification of lignin, adding blocking additives, and post-treatment to remove lignin were also introduced. The production of liquid biofuels from lignocellulosic biomass has shown great environmental benefits such as reducing greenhouse gas emissions and mitigate climate change. By addressing the root causes of lignin-enzyme interaction and how to retard this interaction, it is our hope that this comprehensive review will pave the way for significantly reducing the high cost associated with the enzymatic hydrolysis process, and ultimately achieving a cost-effective and sustainable biorefinery system.

Lignin-enzyme interaction: A roadblock for efficient enzymatic hydrolysis of lignocellulosics

Abstract: Efficiently producing second-generation biofuels from biomass is of strategic significance and meets sustainability targets, but it remains a long-term challenge due to the existence of biomass recalcitrance. Lignin contributes significantly to biomass recalcitrance by physically limiting the access of enzymes to carbohydrates, and this could be partially overcome by applying a pretreatment step to directly target lignin. However, lignin typically cannot be completely removed, and its structure is also significantly altered during the pretreatment. As a result, lignin residue in the pretreated materials still significantly hindered a complete conversion of carbohydrate to its monosugars by interacting with cellulase enzymes. The non-productive adsorption driven by hydrophobic, electrostatic, and/or hydrogen bonding interactions is widely considered as the major mechanism of action governing the unfavored lignin-enzyme interaction. One could argue this type of interaction between lignin residue and the activated enzymes is the major roadblock for efficient enzymatic hydrolysis of pretreated lignocellulosics. To alleviate the negative effects of lignin on enzyme performance, a deep understanding of lignin structural transformation upon different types of pretreatments as well as how and where does lignin bind to enzymes are prerequisites. In the last decade, the progress toward a fundamental understanding of lignin-enzyme interaction, structural characterization of lignin during pretreatment and/or conformation change of enzyme during hydrolysis is resulting in advances in the development of methodologies to mitigate the negative effect of lignin. Here in this review, the lignin structural transformation upon different types of pretreatments and the inhibition mechanism of lignin in the bioconversion of lignocellulose to bioethanol are summarized. Some technologies to minimize the adverse impact of lignin on the enzymatic hydrolysis, including chemical modification of lignin, adding blocking additives, and post-treatment to remove lignin were also introduced. The production of liquid biofuels from lignocellulosic biomass has shown great environmental benefits such as reducing greenhouse gas emissions and mitigate climate change. By addressing the root causes of lignin-enzyme interaction and how to retard this interaction, it is our hope that this comprehensive review will pave the way for significantly reducing the high cost associated with the enzymatic hydrolysis process, and ultimately achieving a cost-effective and sustainable biorefinery system.