Pachira aquatica fruits shells valorization: renewables phenolics through analytical pyrolysis study (Py-GC/MS) / Valorização das cascas dos frutos da Pachira aquática: fenólicos renováveis através de estudo analítico de pirólise (Py-GC/MS)

This research valorized Pachira aquatica Aubl.'s fruit shells (PAS) through its energetic characterization and flash pyrolysis for biofuels or chemicals production. The characterization was performed through proximate and ultimate analysis, bulk density, higher heating value (HHV), hemicellulose, cellulose and lignin content, thermogravimetric analysis and absorption spectra in the infrared region obtained by Fourier-transform infrared spectroscopy technique (FTIR). The analytical flash pyrolysis was performed at 500°C in a Py-5200 HP-R coupled to a gas chromatograph (Py-GC/MS). The PAS biomass presents potential for thermochemical energy conversion processes due to its low moisture and ash content, 76.90% of volatile matter, bulk density of 252.6 kg/m3 and HHV of 16.24 MJ/kg. Flash pyrolysis products are mostly phenols or light organic acids derived from the decomposition of polysaccharides. Results confirmed the potential of PAS to produce bio-phenolics, such as 4-methoxyphenol which is an important active ingredient for skin depigmentation used in drugs and cosmetics, and as phenolic extract that can be used as a precursor to resins, applications that convert this forest waste into bio products for industry into a green circular economy.

Este trabalho teve como objetivo a valorização das cascas dos frutos da Pachira aquatica Aubl. (PAC) através da sua caracterização energética e pirólise flash para produção de biocombustíveis ou produtos químicos. A caracterização foi realizada através de análises imediata e final, densidade aparente, poder calorífico superior (PCS), conteúdos de hemicelulose, celulose e lignina, análise termogravimétrica e espectros de absorção na região do infravermelho obtidos pela técnica de espectroscopia no infravermelho com transformada de Fourier (FTIR). A pirólise flash analítica foi realizada a 500 °C em equipamento Py-5200 HP-R acoplado a um cromatógrafo à gás (Py-GC/MS). A biomassa das PAC apresenta potencial para processos de conversão termoquímica de energia devido ao seu baixo teor de umidade e cinzas, além de 76,90% de materiais voláteis, densidade aparente de 252,6 kg/m3 e PCS igual a 16,24 MJ / kg. Os produtos da pirólise rápida são principalmente fenóis ou ácidos orgânicos leves derivados da decomposição de polissacarídeos. Os resultados confirmam o potencial das PAC para produzir bio-fenólicos, como o 4-metoxifenol que é um importante ingrediente ativo para despigmentação da pele usado em medicamentos e cosméticos, e como extrato fenólico que pode ser usado como precursor de resinas. Estas aplicações convertem esses resíduos florestais em produtos biológicos para a indústria em uma economia circular verde.

Overexpression of a leucine transfer RNA gene tL(CAA)K improves the acetic acid tolerance of Saccharomyces cerevisiae / 生物工程学报

Acetic acid is a common inhibitor present in lignocellulosic hydrolysate. Development of acetic acid tolerant strains may improve the production of biofuels and bio-based chemicals using lignocellulosic biomass as raw materials. Current studies on stress tolerance of yeast Saccharomyces cerevisiae have mainly focused on transcription control, but the role of transfer RNA (tRNA) was rarely investigated. We found that some tRNA genes showed elevated transcription levels in a stress tolerant yeast strain. In this study, we further investigated the effects of overexpressing an arginine transfer RNA gene tR(ACG)D and a leucine transfer RNA gene tL(CAA)K on cell growth and ethanol production of S. cerevisiae BY4741 under acetic acid stress. The tL(CAA)K overexpression strain showed a better growth and a 29.41% higher ethanol productivity than that of the control strain. However, overexpression of tR(ACG)D showed negative influence on cell growth and ethanol production. Further studies revealed that the transcriptional levels of HAA1, MSN2, and MSN4, which encode transcription regulators related to stress tolerance, were up-regulated in tL(CAA)K overexpressed strain. This study provides an alternative strategy to develop robust yeast strains for cellulosic biorefinery, and also provides a basis for investigating how yeast stress tolerance is regulated by tRNA genes.

Single cell oil production by a novel yeast Trichosporon mycotoxinivorans for complete and ecofriendly valorization of paddy straw

Background: Oleaginous yeasts can be grown on different carbon sources, including lignocellulosic hydrolysate containing a mixture of glucose and xylose. However, not all yeast strains can utilize both the sugars for lipogenesis. Therefore, in this study, efforts were made to isolate dual sugar-utilizing oleaginous yeasts from different sources. Results: A total of eleven isolates were obtained, which were screened for their ability to utilize various carbohydrates for lipogenesis. One promising yeast isolate Trichosporon mycotoxinivorans S2 was selected based on its capability to use a mixture of glucose and xylose and produce 44.86 ± 4.03% lipids, as well as its tolerance to fermentation inhibitors. In order to identify an inexpensive source of sugars, nondetoxified paddy straw hydrolysate (saccharified with cellulase), supplemented with 0.05% yeast extract, 0.18% peptone, and 0.04% MgSO4 was used for growth of the yeast, resulting in a yield of 5.17 g L−1 lipids with conversion productivity of 0.06 g L−1 h−1 . Optimization of the levels of yeast extract, peptone, and MgSO4 for maximizing lipid production using Box­Behnken design led to an increase in lipid yield by 41.59%. FAME analysis of single cell oil revealed oleic acid (30.84%), palmitic acid (18.28%), and stearic acid (17.64%) as the major fatty acids. Conclusion: The fatty acid profile illustrates the potential of T. mycotoxinivorans S2 to produce single cell oil as a feedstock for biodiesel. Therefore, the present study also indicated the potential of selected yeast to develop a zero-waste process for the complete valorization of paddy straw hydrolysate without detoxification

Improvement of Delignification, Desilication and Cellulosic Content Availability in Paddy Straw via Physico-chemical Pretreatments

Aim: Paddy straw consists of cellulose and hemicellulose as their plant materials leading to their potential to produce bioethanol through several processes such as pretreatment, enzymatic hydrolysis and ethanol fermentation. Among these processes, pretreatment of paddy straw is particularly important for enzymatic hydrolysis process as they are being limited by the presence of ash and silica content. This study was set to observe the effect of different pretreatments on cellulose, hemicellulose, lignin and ash content of paddy straw. Place and Duration of Study: This study was conducted in Department of Biology, Faculty of Science, Universiti Putra Malaysia, between October 2015 and June 2016. Methodology: Pretreatments comprises the combination of physical (mechanical) and chemical treatments to modify the lignocellulosic structure while reduce lignin and separate silica content in paddy straw fibre. Paddy straw was prepared into three different sizes (2mm, 5mm and 8 mm) for physical treatment. Autoclave, boiled and four different concentrations (0.5%, 1%, 2% and 5% (v/v) and (w/v) respectively) of nitric acid and sodium hydroxide, respectively for chemical treatment were used on paddy straw. Results: Size five millimeter paddy straw showed the highest cellulose content (35.61%) compared to the other sizes and when the paddy pretreated with 2% (w/v) sodium hydroxide (NaOH), the percentage of cellulose content escalated to 72.47%. Pretreatment of 2% (w/v) NaOH have performed the most efficient delignification and desilication process (1.02% lignin; 5.44 ash content); and the performance was supported with SEM images on surface area of the paddy straw with large distortion caused by the treatment. Conclusion: Therefore, a physico-chemical pretreatment of size 5 mm and 2% (w/v) NaOH was found to be the most suitable condition to break the cellulose-lignin complex and make the paddy straw becomes feasible for biofuel production.

Isolation of thermotolerant xylose-utilizing yeasts for ethanol and xylitol production

Aims@#The implementation of simultaneous saccharification and co-fermentation (SScF) and consolidated bioprocessing (CBP) is highly anticipated for industrial bioethanol applications. Thus, microorganisms capable of utilizing hexose and pentose sugars, as well as thermotolerant, are considered advantageous for optimum ethanol production.@*Methodology and results@#Thermotolerant yeast strains were isolated from wastewater ponds of ethanol-producing facility as well as empty fruit bunch composting area and screened for xylose- and glucose-fermenting ability. Five out of 24 total isolates were able to grow at 40 ºC and were found positive for ethanol production from xylose. Based on their high efficiency of xylose and glucose utilization, two isolates were chosen for further characterization. They were identified as Kluyveromyces marxianus UniMAP 1-1 and Schwanniomyces etchellsii UniMAP 1-7 based on the D1/D2 region of the large subunit ribosomal DNA. The growth kinetics of each isolate on xylose and glucose at 40 °C were determined. The two isolates were able to ferment xylose to ethanol at a maximum concentration between 0.533  0.415 and 1.243  0.246 g/L with concomitant xylitol production between 9.932  0.303 and 12.933  0.505 g/L. Fermentation of glucose to ethanol was also tested for these isolates and the yields were and 0.361 and 0.118 g/g for UniMAP 1-1 and UniMAP 1-7, respectively.@*Conclusion, significance and impact of study@#The potential of these thermotolerant microbes to be used for xylitol and bioethanol production from lignocelluloses are evident from this study.

Thermotolerant fermenting yeasts for simultaneous saccharification fermentation of lignocellulosic biomass

Lignocellulosic biomass is the most abundant renewable source of energy that has been widely explored as second-generation biofuel feedstock. Despite more than four decades of research, the process of ethanol production from lignocellulosic (LC) biomass remains economically unfeasible. This is due to the high cost of enzymes, end-product inhibition of enzymes, and the need for cost-intensive inputs associated with a separate hydrolysis and fermentation (SHF) process. Thermotolerant yeast strains that can undergo fermentation at temperatures above 40°C are suitable alternatives for developing the simultaneous saccharification and fermentation (SSF) process to overcome the limitations of SHF. This review describes the various approaches to screen and develop thermotolerant yeasts via genetic and metabolic engineering. The advantages and limitations of SSF at high temperatures are also discussed. A critical insight into the effect of high temperatures on yeast morphology and physiology is also included. This can improve our understanding of the development of thermotolerant yeast amenable to the SSF process to make LC ethanol production commercially viable.

Production, purification and characterization of an ionic liquid tolerant cellulase from Bacillus sp. isolated from rice paddy field soil

Background: Lignocellulosic biomass is a renewable, abundant, and inexpensive resource for biorefining process to produce biofuel and valuable chemicals. To make the process become feasible, it requires the use of both efficient pretreatment and hydrolysis enzymes to generate fermentable sugars. Ionic liquid (IL) pretreatment has been demonstrated to be a promising method to enhance the saccharification of biomass by cellulase enzyme; however, the remaining IL in the hydrolysis buffer strongly inhibits the function of cellulase. This study aimed to isolate a potential IL-tolerant cellulase producing bacterium to be applied in biorefining process. Result: One Bacillus sp., MSL2 strain, obtained from rice paddy field soil was isolated based on screening of cellulase assay. Its cellulase enzyme was purified and fractionated using a size exclusion chromatography. The molecular weight of purified cellulose was 48 kDa as revealed by SDS-PAGE and zymogram analysis. In the presence of the IL, 1-ethyl-3-methylimidazolium acetate ([C2mim][OAc]) concentration of 1 M, the cellulase activity retained 77.7% of non-IL condition. In addition, the optimum temperature and pH of the enzyme is 50°C and pH 6.0, respectively. However, this cellulase retained its activity more than 90% at 55°C, and pH 4.0. Kinetic analysis of purified enzyme showed that the Km and Vmax were 0.8 mg/mL and 1000 μM/min, respectively. Conclusion: The characterization of cellulase produced from MSL2 strain was described here. These properties of cellulase made this bacterial strain become potential to be used in the biorefining process.

Enhancement of cellulolytic enzymes and xylanase production via classical mutational techniques under solid-state fermentation condition

Aims: High cost of cellulases remains the most significant barrier to the economical production of bio-ethanol from lignocellulosic biomass (LB). The present study aims at developing a local cellulolytic fungal strain through random mutagenesis coupled with the feasibility of solid-state fermentation (SSF) by utilizing agricultural wastes such as oil palm frond (OPF) as the substrate. Methodology and results: Out of 95 wild isolates tested, native fungal strain Aspergillus niger, designated DWA8 was isolated as the top enzymatic secretor. For quantitative enzyme analysis, SSF was conducted using 1x106 spore/mL inoculated onto 5 g of ground OPF, incubated at room temperature for 7 days, with 70% moisture content and an initial medium pH of 7. Random mutagenesis has always been tempting in the enhancement of enzyme production. In this work, the compounded treatment of microwave, ultraviolet (UVC) and Ethyl Methanesulfonate (EMS) have generated an Aspergillus niger MUE3.06 mutant with an overall increase of 114% in CMCase activity, approximately 70% in FPase and Xylanase activity respectively compared with the parental DWA8 strain. Thus this finding is capable to be fully developed as an established mutational scheme to create highly productive filamentous fungus in a cheap, simple and sustainable way. Conclusion, significance and impact of study: It was the first attempt to explore the combine effect of the three popular mutagens upon cellulases and xylanases. It is believed that more diversified of mutagen types induce more diversified mutation pattern (with instructive planning), which is very desirable in creating new enzymes with novel abilities.

Biomass Pre-treatment Methods and Their Economic Viability for Efficient Production of Biofuel.

Development of sustainable technology for bioenergy/biofuel generation using lignocellulosic biomass is of prime importance in the present day research. An economically viable technology for production of ethanol from lignocellulosic biomass will certainly provide opportunity to the non-oil producing countries to refrain from deprivation of fuel and explore the concept of bio-refineries for complete utilization of the feedstock. Biomass feedstock is abundant in the form of network of cellulose, hemicelluloses and lignin and hence the name lignocellulose. The structure and composition of lignocellulosic biomass is a major hindrance in its complete digestibility into fermentable monomer sugars. Reports are in the literature indicating that pre-treatment of biomass from wood and grasses can increase the yield of sugars upto 90%. Here, we have reviewed various technologies and methods for the pre-treatment of biomass with emphasis on their advantages and disadvantages with respect to the production of biofuel. Also, capital expense (CAPEX) and operational expense (OPEX) is considered to assess the economic viability of the technology.

ESTUDIO DE MODIFICACIÓN QUÍMICA Y FÍSICA DE BIOMASA (Citrus sinensis Y Musa paradisiaca) PARA LA ADSORCIÓN DE METALES PESADOS EN SOLUCIÓN / STUDY OF PHYSICAL AND CHEMICAL MODIFICATION OF BIOMASS (Citrus sinensis AND Musa paradisiaca) FOR THE ADSORPTION OF HEAVY METALS IN SOLUTION

Se estudia el efecto de las modificaciones a carbón activado y recubrimiento con quitosano de biomasa lignocelulósica obtenida de cáscaras de plátano y naranja, para la adsorción de Cr (VI). La caracterización de los grupos funcionales en las biomasas aptos para la adsorción se verificó mediante un análisis elemental (CHON) y espectroscopia de infrarrojo (IR), mientras que para los carbones activados se determinó su área superficial por medio de un análisis BET. El contenido de Cr (VI) en solución se midió mediante espectrofotometría UV-vis, usando el método de la difenilcarbazida. Los resultados mostraron una remoción de los iones de Cr (VI) de 66,6 y 93 ppm para las cáscaras de naranja y plátano respectivamente, los carbones activados removieron 85 y 95 ppm, mientras que las biomasas modificadas con quitosano presentaron una adsorción 61,24 y 88,2 ppm. Se observa que la cinética de adsorción fue mejor descrita por la ecuación de Pseudo Segundo Orden, y el efecto de competitividad bimetálica se vio afectada de mayor forma por iones de níquel, y en menor proporción por iones de plomo.

The effect of changes to activated charcoal and chitosan coating of lignocellulosic biomass obtained from banana and orange peels for the absorption of Cr (VI) was studied. Characterization of the functional groups in the biomass suitable for the adsorption was monitored by elemental analysis (CHON) and infrared spectroscopy (IR), while for activated carbon surface area was determined by BET analysis. The Cr (VI) content in solution was measured by UV-vis spectrophotometer, using the diphenylcarbazide method. The results showed a removal of Cr (VI) ions of 66.6 and 93 ppm for orange peels and banana peels respectively; the activated carbons removed 85 and 95 ppm, while the modified biomasses with chitosan showed an adsorption of 61.24 and 88.2 ppm. It was observed that the adsorption kinetics was best described by the Pseudo Second Order equation, and the bimetallic competitiveness effect was affected more by nickel ions and to a lesser extent by lead ions.

Isolation and Optimization of Cellulolytic and Xylanolytic Microbes from Nepal and their Utilization for Lignocellulosic Biomass Degradation.

Microorganisms harbored by nature and guts of herbivorous animals can degrade different plant related biomass. One of the in-between steps for conversion of lignocellulosic biomass to ethanol entails isolation and identification of microorganisms that could convert pretreated biomass into a suitable form, which could then be fermented into bioethanol. We isolated 36 different microorganisms from hot spring, 6 from ruminant’s (goat) gut, 2 sample from hay spray on the basis of their ability to secrete enzymes that hydrolyzed different plant constituents. Similarly 3 microorganisms were isolated from the rotten wood available around Kathmandu University on the basis of utilization of xylose and glucose. 3 varieties of thermophiles, gut microorganism and microorganism from hay spray that showed the highest cellulolytic and xylanolytic activities by saccharification of cellulose and xylan into their monosaccharide glucose and xylose units respectively were then applied on different biomass (rice straw, corn stover and sugarcane bagasse). Before Saccharification biomass was made accessible for the digestion by enzymes through 3 different pretreatment strategies (3.35% H2SO4, NaOH and H2O2 with 1:10 substrate: chemical ratio) following thermal strategy of steam explosion. Also, different conditions like incubation time, pH and temperature for saccharification were assessed with the highest liberation of reducing sugar at pH 5, temperature of 5̊ C and incubation time of 4 days. Microorganism from rotten wood was able to utilize both xylose and glucose and yielded highest amount (5.567 mg/ml) of bioethanol.

Evaluation of polymeric adsorbent resins for efficient detoxification of liquor generated during acid pretreatment of lignocellulosic biomass.

Production of fuel ethanol from lignocellulosic biomass conventionally includes biomass pretreatment, hydrolysis, and fermentation. The liquor generated during dilute acid pretreatment of biomass contains considerable quantities of pentose sugars as well as various degradation products of sugars and lignin, like furfural, hydroxymethyl furfural (HMF), organic acids, aldehydes and others, which are known to be inhibitory for microbial growth. This pentose rich liquor is a potent resource which can be used to produce alcohol or other value added metabolites by microbial fermentation. However, the presence of these inhibitory compounds is a major hindrance and their removal is essential for efficient utilization of this byproduct stream. In the present work, the polymeric adsorbent resins, XAD-4, XAD-7 and XAD-16 were evaluated for their ability to adsorb fermentation inhibitors like furfural and HMF from the acid pretreated liquor. These resins could remove 55-75% of furfural and 100% of HMF and more than 90% sugar remained un-adsorbed in the pretreated liquor. Desorption of furfural from stationary phase was evaluated by using ethanol and hot water. The results suggest that these polymeric resins may be used for detoxification of acid pretreatment liquor with selective removal of sugar degradation products without affecting the sugar content in the solution.

Simultaneous saccharification and fermentation process of different cellulosic substrates using a recombinant Saccharomyces cerevisiae harbouring the beta-glucosidase gene

In Brazil, the production of ethanol from sugarcane produces large amounts of lignocellulosic residues (bagasse and straw), which have been driving research and development for the production of second generation ethanol. In the present work, a recombinant Saccharomyces cerevisiae strain expressing the beta-glucosidase gene from Humicola grisea was used for ethanol production from three different cellulosic sources by simultaneous saccharification and fermentation. Initially, a enzymatic pre-hydrolysis step was done with a solid:liquid ratio of 1:4, and an enzymatic load of 25 filter paper activity (FPU).g-1 of cellulosic substrate. Using sugarcane bagasse pretreated cellulignin, crystalline cellulose and carboxymethyl cellulose, 51.7 g L-1, 41.7 g L-1 and 13.8 g L-1 of ethanol was obtained, respectively, at the end of 55 hrs of fermentation. The highest ethanol productivity (0.94 g L-1 hrs-1) was achieved using sugarcane bagasse pretreated cellulignin. The use of a recombinant S. cerevisiae led to extremely low glucose concentrations when compared to other works reported in literature.