Separate Hydrolysis And Fermentation Research Articles

Separate hydrolysis and fermentation (SHF) of pretreated sugar beet pulp (SBP) into ethanol

Separate hydrolysis and fermentation (SHF) of pretreated sugar beet pulp (SBP) into ethanol

Biobutanol Production from Rice Husk

Abstract: This research aimed to study biobutanol production using Lignocellulosic renewable substrate Rice Husk, carried out using Clostridium acetobutylicum MTCC 11274 via the Separate hydrolysis and fermentation (SHF) process. The simple sugars were exposed using a delignification process with different concentrations of Ammonia solution and some were hydrolyzed using conc. H2SO4 after getting treated with ammonia. The sugars were estimated using DNSA Method after which samples were fermented. The concentration of Biobutanol was estimated using GC-MS showing 2.2 gm/L in one of the tested samples.

Bioprospecting marine microalgae and cyanobacteria as alternative feedstocks for bioethanol production

In view of fossil fuel depletion at an alarming rate, bioethanol is considered a sustainable alternative fuel to meet the global insatiable energy needs. In this regard, the present study focused on bioprospecting eight marine cyanobacteria and four marine green microalgae to select suitable feedstock for bioethanol production based on their biomass yield and carbohydrate accumulation. Before fermentation, hydrolysis is enhanced by the standardization of sulphuric acid and Lewis acid concentrations at a specific biomass-to-liquid ratio. Based on comprehensive carbohydrate component analysis, Leptolyngbya valderiana BDU 41001 with maximum fermentable sugar content (27% dry cell weight) was selected as a potential third-generation feedstock from eight cyanobacteria with high carbohydrate content (>25% dcw) for bioethanol production. The synergistic impact of 2% sulphuric acid (v/v) and 2% magnesium sulfate (w/v) at a biomass-liquid ratio of 1:15 yielded the highest hydrolysis yield of 88%. Scanning Electron Microscopy examination also evidently demonstrated the morphological changes that occurred during the hydrolyzation of cyanobacterial filaments. Separate Hydrolysis and Fermentation (SHF) were then used to produce bioethanol from the acquired hydrolysate by the conventional yeast Saccharomyces cerevisiae at an ideal pH of 5.5, yielding 0.16 g g-1 bioethanol yield with 45% bioethanol conversion. This study demonstrated the selection criteria of bioethanol feedstock and establishes an effective method for manufacturing bioethanol from marine cyanobacterium L. valderiana. Furthermore, this research contributes to the potential of Lewis acid as a cost-effective and environment-friendly source in the biomass conversion process.

Biohydrogen and Methane Production from Sugarcane Leaves Pretreated by Deep Eutectic Solvents and Enzymatic Hydrolysis by Cellulolytic Consortia

This study determined the optimal conditions for the deep eutectic solvent (DES) pretreatment of sugarcane leaves and the best fermentation mode for hydrogen and methane production from DES-pretreated sugarcane leaves. Choline chloride (ChCl):monoethanolamine (MEA) is the most effective solvent for removing lignin from sugarcane leaves. The optimum conditions were a ChCl: MEA molar ratio of 1:6, 120 °C, 3 h, and substrate-to-DES solution ratio of 1:12. Under these conditions, 86.37 ± 0.36% lignin removal and 73.98 ± 0.42% hemicellulose removal were achieved, whereas 84.13 ± 0.77% cellulose was recovered. At a substrate loading of 4 g volatile solids (VS), the simultaneous saccharification and fermentation (SSF) and separate hydrolysis and fermentation (SHF) processes yielded maximum hydrogen productions of 3187 ± 202 and 2135 ± 315 mL H2/L, respectively. In the second stage, methane was produced using the hydrogenic effluent. SSF produced 5923 ± 251 mL CH4/L, whereas SHF produced 3583 ± 128 mL CH4/L. In a one-stage methane production process, a maximum methane production of 4067 ± 320 mL CH4/L with a substrate loading of 4 g VS was achieved from the SSF process. SSF proved to be more efficient than SHF for producing hydrogen from DES-pretreated sugarcane leaves in a two-stage hydrogen and methane production process as well as a one-stage methane production process.

Valorization of Bombax ceiba Waste into Bioethanol Production through Separate Hydrolysis and Fermentation and Simultaneous Saccharification and Fermentation

In this study, Seed pods of B. ceiba were used as a novel, cheap, and sustainable feedstock for second-generation bioethanol production. B. ceiba waste was pretreated with NaOH under different conditions using a Box–Behnken design (BBD) with three factors and three levels. Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and scanning electron microscopy (SEM) were used to investigate the chemical, structural, and morphological modifications made by pretreatment. NaOH pretreatment followed by steam was more effective as it offered 60% cellulose and 9% lignin at 10% substrate loading, 5% NaOH conc., and 4 h residence time. Samples with maximum cellulose were employed for ethanol production by separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) using indigenously produced cellulase as well as commercial cellulase. HPLC analysis revealed the best saccharification (50.9%) at 24 h and the best ethanol yield (54.51 g/L) at 96 h of fermentation in SSF using commercial cellulose by Saccharomyces cerevisiae. SSF offered a better production of bioethanol from seed pods than SHF. The implications of the work support the notion that B. ceiba waste could be utilized for large-scale bioethanol production.

Ethanol Production from the Mixture of Waste French Fries and Municipal Wastewater via Separate Hydrolysis and Fermentation.

The potential of bioethanol generation using the mixture of waste French fries (WFF) and municipal wastewater (MWW) via separate hydrolysis and fermentation (SHF) was evaluated in this study. The effect of WFF substrate loading (SL, 10%, 16%, and 20%, w/v) on the SHF was also examined. Both glucose production and hydrolysis efficiency increased with increasing of SL from 10 to 16% and the maximum glucose yield of 0.236g glucose/g WFF and hydrolysis efficiency of 91.9% were obtained at SL of 16%. However, the glucose production and hydrolysis efficiency decreased when the SL further increased to 20% due to the inhibition on enzyme caused by higher glucose production. The mixture hydrolysate was then used as feedstock for ethanol fermentation. The maximum ethanol production of 22.69g/L was obtained from SL of 16%. The highest rate of glucose conversion to ethanol was 84.2%. The results demonstrated that the mixture of WFF and MWW could be used for ethanol production by the SHF.

Biofuels Production and Processing Technology

The negative global warming impact and global environmental pollution due to fossil fuels mean that the main challenge of modern society is finding alternatives to conventional fuels. In this scenario, biofuels derived from renewable biomass represent the most promising renewable energy sources. Depending on the biomass used by the fermentation technologies, it is possible obtain first-generation biofuels produced from food crops, second-generation biofuels produced from non-food feedstock, mainly starting from renewable lignocellulosic biomasses, and third-generation biofuels, represented by algae or food waste biomass. Although biofuels appear to be the closest alternative to fossil fuels, it is necessary for them to be produced in competitive quantities and costs, requiring both improvements to production technologies and diversification of feedstock. This Special Issue is focused on technological innovations, which include but are not limited to the utilization of different feedstock; different biomass pretreatments; fermentation strategies, such as simultaneous saccharification and fermentation (SSF) or separate hydrolysis and fermentation (SHF); different applied microorganisms used as monoculture or co-culture; and different setups for biofuel fermentation processes.

The use of commercial wine yeast Saccharomyces cerevisiae EC1118 for cassava ethanol production at high solids loading

Cassava is one of the most commonly imported raw materials for ethanol fermentation for the manufacture of food-grade distilled spirits in Korea. In cassava-producing countries, such as the Lao PDR, cassava can be considered low-price biomass for the production of bioethanol. In this study, the commercial wine yeast Saccharomyces cerevisiae EC1118 was tested for ethanol fermentation using cassava powder at a high solids loading (30%, w/v). α-Amylase and glucoamylase were used for the hydrolysis of cassava starch into glucose. To identify a suitable fermentation process for cassava, separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) were compared. From the complete enzyme hydrolysis of cassava, 254.1 g/L of glucose was obtained. SSF showed higher ethanol titer during the first 12 h of fermentation, but SHF showed a higher ethanol titer after 24 h of fermentation. Finally, there was no significant difference between SHF and SSF in the final ethanol titer after 48 h fermentation (133.6 and 130.6 g/L, respectively). In summary, both SHF and SSF are applicable for ethanol production with high solid cassava using wine yeast EC1118 under the test conditions.

Influence of nanoparticles inclusion on the production of bioethanol from corn stalks and leaves

Bioethanol is a viable alternative to petroleum-derived fossil fuels. It is renewable, low-cost, and the preferred fuel for most developing countries worldwide. Although it is possible to make bioethanol from corn stalks and leaves wastes, second-generation biofuels made from agricultural waste feedstocks represent a significant step forward. In the present research, nickel oxide (NiO) nanoparticles (NPs) were used as a biocatalyst to achieve maximum ethanol output. The pretreatment of 2% NaOH with NiO NPs, 15-min autoclave condition, showed the highest total and reducing sugar yield was 162.69 g/L and 43.75 g/L. After hydrolysis, the suitable total and reducing sugar yield of 185.43 g/L and 125.42 g/L was chosen for further fermentation with the expansion of Saccharomyces cerevisiae cells. Separate hydrolysis and fermentation (SHF) using corn stalk and leaf waste was significantly assisted by incorporating a nanocatalyst; ethanol concentration was increased to 15.8 g/L at 24 hours incubation period. The study revealed critical information regarding ways NiO NPs could be employed to improve the efficiency of the ethanol production bioprocess.

Valorisation of source-separated food waste to bioethanol: pilot-scale demonstration

Food waste is a waste stream that is produced globally in huge amounts and therefore constitutes a major environmental concern. Additionally, the growing consumption of fossil fuels sets the need for alternative energy sources. To this end, in this paper, an holistic approach towards bioethanol production from source-separated food waste was studied as an effective strategy to cope with both issues. Source-separated food waste collected from a Greek Municipality was used as raw material. Two fermentation modes, separate hydrolysis and fermentation and simultaneous saccharification and fermentation, were examined in laboratory and pilot scales with varying solids loadings. For separate hydrolysis and fermentation (SHF) trials, the solids loading increase led to a significant ethanol yield reduction from 79 to 55 g/kg food waste, whereas for simultaneous saccharification and fermentation (SSF), the ethanol yield was increased by 77% (from 62 to 110 g/kg food waste) as the solids loading was increased. This is also related to greater ethanol concentrations, which are beneficial in terms of technoeconomics. The lowest bioethanol production cost, 1.57 €/kg ethanol, was estimated for the scenario of SSF with 20% solids loading while for SHF the lowest production cost was achieved (4.40 €/kg ethanol) when 15% solids loading is applied. In most cases, the energy and enzyme costs presented the most pronounced impact on the total bioethanol cost. In conclusion, it was proved that the food waste valorisation towards bioethanol production is technically feasible on a pilot scale. However, further techno-economic factors of the whole value chain must also be taken into consideration while aiming to assess the viability of the process.

Bioethanol Production Optimization from KOH-Pretreated Bombax ceiba Using Saccharomyces cerevisiae through Response Surface Methodology

The present study was based on the production of bioethanol from alkali-pretreated seed pods of Bombax ceiba. Pretreatment is necessary to properly utilize seed pods for bioethanol production via fermentation. This process assures the accessibility of cellulase to the cellulose found in seedpods by removing lignin. Untreated, KOH-pretreated, and KOH-steam-pretreated substrates were characterized for morphological, thermal, and chemical changes by scanning electron microscopy (SEM), thermogravimetric analysis (TGA), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR). Hydrolysis of biomass was performed using both commercial and indigenous cellulase. Two different fermentation approaches were used, i.e., separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF). Findings of the study show that the maximum saccharification (58.6% after 24 h) and highest ethanol titer (57.34 g/L after 96 h) were observed in the KOH-steam-treated substrate in SSF. This SSF using the KOH-steam-treated substrate was further optimized for physical and nutritional parameters by one factor at a time (OFAT) and central composite design (CCD). The optimum fermentation parameters for maximum ethanol production (72.0 g/L) were 0.25 g/L yeast extract, 0.1 g/L K2HPO4, 0.25 g/L (NH4)2SO4, 0.09 g/L MgSO4, 8% substrate, 40 IU/g commercial cellulase, 1% Saccharomyces cerevisiae inoculum, and pH 5.

High-Solid Loading Enzymatic Hydrolysis of Waste Office Paper for poly-3-hydroxybutyrate Production Through Simultaneous Saccharification and Fermentation

Waste paper holds great potential as a substrate for the microbial production of bioplastic (Poly-3-hydroxybutyrate (PHB)). This study aimed to produce PHB by utilizing office paper as a substrate using Cupriavidus necator through batch and fed-batch simultaneous saccharification and fermentation (SSF) approach. For the batch experiment, different loadings of shredded office paper (3, 5 and 10%) with two different pretreatments H2O2 (OPH) and H2O2 and Triton X-100 (OPTH) were carried out. For the fed-batch experiment, paper loading started with 3% and two more additions were made at 36 and 84 h. Both experiments were conducted at 30 °C, 200 rpm and pH 7 using 55.5 FPU/g of cellulase and 37.5 CBU/g of β-glucosidase with a fixed amount of nitrogen source. High PHB yield was observed with OPH in all loadings, though the OPHT showed a better hydrolysis. Maximum PHB yield (4.27 g/L) was achieved with 10% OP on the sixth day of fermentation in batch SSF. Whereas, maximum PHB yield (4.19 g/L) was obtained within a shorter time (66 h)with OPH in the fed-batch experiment. The extracted PHB showed well-matched characteristic features to the standard PHB. Finally, this study proves the feasibility of employing the SSF process for PHB production using waste paper as an alternative approach to overcome the shortcoming of the separate hydrolysis and fermentation (SHF) process.

Novel system design for high solid lignocellulosic biomass conversion

A novel system (Oregon State University High Solids Reactor; OSU-HSR) was designed and constructed for enzymatic hydrolysis at ultrahigh solids content (40%) by promoting better mixing using low energy consumption in a horizontal reactor with a new impeller design and a controllable feeding unit. System performance was evaluated using separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) methodologies. Using the dilute acid pretreated wheat straw as the substrate in the OSU-HSR system, the highest glucose (219.7 g/L) and ethanol (127.1 g/L) concentrations were achieved with the use of the SHF method while the highest ethanol concentration using SSF method was 134.5 g/L. The SSF method increased the return on investment to 12.21% with an estimated global warming potential of 54.5 g CO2 eq/MJ Ethanol. The OSU-HSR successfully provided effective mixing and different fed-batch schemes, and can be used for efficient biochemical conversion of lignocellulosic biomass into bio-chemicals and biofuels.

Consolidated bioprocessing of cassava starch into microbial lipid for biodiesel production by the amylolytic yeast Lipomyces starkeyi

Direct conversion of starch materials into lipid has been rarely reported as most oleaginous species are unable to efficiently produce amylolytic enzymes. Herein, consolidated bioprocessing (CBP) of cassava starch was described efficiently for lipogenesis by Lipomyces starkeyi. Lipid concentration and yield reached 13.99 g/L and 0.187 g/g, respectively, from 75 g/L cassava starch under optimal conditions. The CBP strategy featured zero enzyme cost, simpler process, and higher oleaginicity compared with the separate hydrolysis and fermentation (SHF) strategy. Amylolytic enzymes with α-amylase, maltase and glucoamylase activities were secreted into extracellular space and the amylase activity reached 3.65 U/mL as maximum. The genome of L. starkeyi was sequenced and a small-scale metabolic model was established. The mechanism of starch hydrolysis for lipogenesis by L. starkeyi was estimated accordingly. The estimated biodiesel parameters of the CBP-derived lipid samples indicated high-quality fuel properties. The CBP strategy provides a highly integrated and techno-economic route for starch-to-lipid conversion.

Direct production of astaxanthin from food waste by Phaffia rhodozyma

Bioconversion of food waste (FW) into value-added products is a promising approach to solve FW management problem and achieve the global goal of carbon peak and carbon neutralization. In this study, FW could be used as a complete medium to compare the effects of four different fermentation strategies on direct astaxanthin fermentation by Phaffia rhodozyma. The carotenoid production by direct fermentation (DF) reached 129.5 mg/L, increasing by 61.3 %, compared with separate hydrolysis and fermentation (SHF), and carotenoid production by simultaneous saccharification fermentation (SSF) and 1 h pre-hydrolysis SSF (PSSF) were only 20.2 % and 21.5 % higher than DF, respectively. The mechanism of astaxanthin production in the DF could be attributed to the utilization of rice starch due to the production of amylase, and the stimulation of cell growth resulted from the metal elements and the lipid of soybean oil in FW. Remarkably, a 70.3 % increase of carotenoid production was observed when substrate loading increased from 10 % to 20 %. The present study provides a novel process for astaxanthin production from direct bioconversion of FW by P. rhodozyma, which shows potential for low-cost industrial production of astaxanthin.

Sequential fermentation of organosolv pretreated corn stover for improved cellulose hydrolysis and bioethanol yield

In this study, organosolv pretreatment of corn stover using 40% acetic acid was investigated for its amenability for enzymatic saccharification and bioethanol production. The pretreated corn stover resulted in 63.4±1.7% solids, 46.4±2.5% hemicellulose and 53.7±2.1% cellulose recoveries with 84.6% delignification. The reducing sugars of 0.107g/gds (107/Kg) released during pretreatment can be fermented to produce 0.125ml/gds (125ml/Kg) bioethanol. The saccharification of pretreated biomass under optimized conditions released 0.395g/gds reducing sugars, which upon subsequent fermentation by Saccharomyces cerevisiae and Pachysolen tannophilus produced 0.112g/gds (140ml/Kg) ethanol, with 66.54% fermentation efficiency and 33.9% ethanol yield (YPS), under separate hydrolysis and fermentation (SHF) conditions. The net price for the bioconversion of 1Kg corn stover for bioethanol (265 ml) production was calculated to be Rs. 144.3/L of ethanol. This price is high being calculated on laboratory scale (1Kg) and will decrease substantially upon upscaling.

Bioethanol from Napier grass employing different fermentation strategies to evaluate a suitable operation for batch bioethanol production

• Variations in substrate and enzyme additions in SSF of Napier grass were compared. • SHF and SSF resulted in 30.6 g/L and 28.5 g/L ethanol from 15% Napier grass. • Alleviate mixing problem by portioning addition did not improve ethanol production. • Pre-incubating the grass before SSF negatively impacts ethanol production. • Extra loading of substrate during SSF resulted in higher ethanol production. Napier grass has been used as animal feed especially in countries with tropical climates. As interest in alternative feedstocks for fuel bioethanol increases, Napier grass has been considered as a potential feedstock for lignocellulosic bioethanol production. Two main fermentation strategies, including separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF), were investigated with various substrate and enzyme feeding patterns in SSF. Total substrate loading of 15% and enzyme dosage of 40 FPU/g substrate were controlled to compare the performance of each fermentation variation. SHF and SSF resulted in similar ethanol production of 30.6 ± 0.4 g/L and 28.5 ± 2.3 g/L. Preincubation of grass and enzyme resulted in significant lower ethanol at 20.3 ± 1.6 g/L. Portioning the addition of grass and enzyme did not change the ethanol production significantly. Increasing the grass loading to 30% via portioning approach resulted in an increase in ethanol production in batch fermentation, while the addition of extra enzyme did not help with increasing ethanol production.

Strategies for Improvement of Lipid Production by Yeast Trichosporon oleaginosus from Lignocellulosic Biomass.

Microbial lipids have similar fatty acid composition to plant oils, and therefore, are considered as an alternative feedstock for biodiesel production. Oleaginous yeasts accumulate considerable amounts of lipids intracellularly during growth on low-cost renewable feedstocks such as lignocellulosic biomass. In this study, we cultivated yeast Trichosporon oleaginosus on hydrolysate of alkaline pretreated corn cobs. Different process configurations were evaluated and compared, including separate hydrolysis and fermentation (SHF) with cellulase recycle and simultaneous saccharification and fermentation (SSF) in batch and fed-batch mode. At low enzyme loading, the highest lipid concentration of 26.74 g L−1 was reached in fed-batch SSF fed with 2.5% (g g−1) substrate. Batch SHF was conducted for four rounds with recycling the cellulase adsorbed on unhydrolyzed lignocellulosic biomass. Thirty percent of cellulase saving was achieved for rounds 2–4 without compromising productivity and lipid yield. The addition of Tween 80 to lignocellulosic slurry improved the hydrolysis rate of structural carbohydrates in pretreated lignocellulosic biomass. Furthermore, supplementing the growth medium with Tween 80 improved lipid yield and productivity without affecting yeast growth. Oleaginous yeast T. oleaginosus is a promising strain for the sustainable and efficient production of lipids from renewable lignocellulosic feedstock.

Isolation and Identification of Cellulase Producing and Sugar Fermenting Bacteria for Second-Generation Bioethanol Production

Over the last decades, the negative impacts of fossil fuel on the environment and increasing demand for energy due to the unavoidable depletion of fossil fuels, has transformed the world’s interests towards alternative fuels. In particular, bioethanol production from cellulosic biomass for the transportation sector has been incrementing since the last decade. The bacterial pathway for bioethanol production is a relatively novel concept and the present study focused on the isolation of potential “cellulase-producing” bacteria from cow dung, compost soil, and termite gut and isolating sugar fermenting bacteria from palm wine. To select potential candidates for cellulase enzyme production, primary and secondary assays were conducted using the Gram’s iodine stain in Carboxy Methyl Cellulose (CMC) medium and the Dinitrosalicylic acid (DNS) assays, respectively. Durham tube assay and Solid-Phase Micro-Extraction (SPME) coupled with Gas Chromatography-Mass Spectrometry (GC-MS) was used to evaluate the sugar fermenting efficiency of the isolated bacteria. Out of 48 bacterial isolates, 27 showed cellulase activity where Nocardiopsis sp. (S-6) demonstrated the highest extracellular crude enzyme activity of endoglucanase (1.56±0.021 U) and total cellulase activity (0.93±0.012 U). The second-highest extracellular crude enzyme activity of endoglucanase (0.21±0.021 U) and total cellulase activity (0.35±0.021 U) was recorded by Bacillus sp. (T-4). Out of a total of 8 bacterial isolates, Achromobacter sp. (PW-7) was positive for sugar fermentation resulting in 3.07% of ethanol in broth medium at 48 h incubation. The results of the study revealed that Nocardiopsis sp. (S-6) had the highest cellulase enzyme activity. However, the highest ethanol percentage was achieved with by having both Bacillus sp. (T-4) and Achromobacter sp. (PW-7) for the simultaneous saccharification and fermentation (SSF) method, as compared to separate hydrolysis and fermentation (SHF) methodologies.

Polish Varieties of Industrial Hemp and Their Utilisation in the Efficient Production of Lignocellulosic Ethanol.

Nowadays, more and more attention is paid to the development and the intensification of the use of renewable energy sources. Hemp might be an alternative plant for bioenergy production. In this paper, four varieties of Polish industrial hemp (Białobrzeskie, Tygra, Henola, and Rajan) were investigated in order to determine which of them are the most advantageous raw materials for the effective production of bioethanol. At the beginning, physical and chemical pretreatment of hemp biomass was carried out. It was found that the most effective is the alkaline treatment with 2% NaOH, and the biomasses of the two varieties were selected for next stages of research: Tygra and Rajan. Hemp biomass before and after pretreatment was analyzed by FTIR and SEM, which confirmed the effectiveness of the pretreatment. Next, an enzymatic hydrolysis process was carried out on the previously selected parameters using the response surface methodology. Subsequently, the two approaches were analyzed: separated hydrolysis and fermentation (SHF) and a simultaneous saccharification and fermentation (SSF) process. For Tygra biomass in the SHF process, the ethanol concentration was 10.5 g∙L−1 (3.04 m3·ha−1), and for Rajan biomass at the SSF process, the ethanol concentration was 7.5 g∙L−1 (2.23 m3·ha−1). In conclusion, the biomass of Polish varieties of hemp, i.e., Tygra and Rajan, was found to be an interesting and promising raw material for bioethanol production.