Ravi P. Gupta

Pilot scale study on steam explosion and mass balance for higher sugar recovery from rice straw.

Pretreatment of rice straw on pilot scale steam explosion has been attempted to achieve maximum sugar recovery. Three different reaction media viz. water, sulfuric acid and phosphoric acid (0.5%, w/w) were explored for pretreatment by varying operating temperature (160, 180 and 200°C) and reaction time (5 and 10min). Using water and 0.5% SA showed almost similar sugar recovery (∼87%) at 200 and 180°C respectively. However, detailed studies showed that the former caused higher production of oligomeric sugars (13.56g/L) than the later (3.34g/L). Monomeric sugar, followed the reverse trend (7.83 and 11.62g/L respectively). Higher oligomers have a pronounced effect in reducing enzymatic sugar yield as observed in case of water. Mass balance studies for water and SA assisted SE gave total saccharification yield as 81.8% and 77.1% respectively. However, techno-economical viability will have a trade-off between these advantages and disadvantages offered by the pretreatment medium.

Structural features of dilute acid, steam exploded, and alkali pretreated mustard stalk and their impact on enzymatic hydrolysis

To overcome the recalcitrant nature of biomass several pretreatment methodologies have been explored to make it amenable to enzymatic hydrolysis. These methodologies alter cell wall structure primarily by removing/altering hemicelluloses and lignin. In this work, alkali, dilute acid, steam explosion pretreatment are systematically studied for mustard stalk. To assess the structural variability after pretreatment, chemical analysis, surface area, crystallinity index, accessibility of cellulose, FT-IR and thermal analysis are conducted. Although the extent of enzymatic hydrolysis varies upon the methodologies used, nevertheless, cellulose conversion increases from <10% to 81% after pretreatment. Glucose yield at 2 and 72h are well correlated with surface area and maximum adsorption capacity. However, no such relationship is observed for xylose yield. Mass balance of the process is also studied. Dilute acid pretreatment is the best methodology in terms of maximum sugar yield at lower enzyme loading.

Bioethanol production from wheat straw via enzymatic route employing Penicillium janthinellum cellulases.

This study concerns in-house development of cellulases from a mutant Penicillium janthinellum EMS-UV-8 and its application in separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) processes for bioethanol production from pre-treated wheat straw. In a 5L fermentor, the above strain could produce cellulases having activity of 3.1 FPU/mL and a specific activity of 0.83 FPU/mg of protein. In-house developed cellulase worked more efficiently in case of SSF as ethanol concentration of 21.6g/L and yield of 54.4% were obtained which were higher in comparison to SHF (ethanol concentration 12 g/L and 30.2% yield). This enzyme preparation when compared with commercial cellulase for hydrolysis of pre-treated wheat straw was found competitive. This study demonstrates that P. janthinellum EMS-UV-8 is a potential fungus for future large-scale production of cellulases.

Blending of cellulolytic enzyme preparations from different fungal sources for improved cellulose hydrolysis by increasing synergism

Cellulolytic enzymes were produced from the three fungal strains [P. janthinellum EMS-UV-8 (E), T. reesei Rut C-30 (R) and A. tubingenesis (A)] and used to prepare blends for the hydrolysis of avicel and acid treated wheat straw (A-WS). An enzyme blend prepared from three different crude preparations (E + R + A) on the basis of equivalent FPU or protein was found to be more synergistic and gave better hydrolysis of avicel or A-WS than the blend of two enzyme preparations (E + R, E + A and R + A) or individual enzyme preparations (E, R, and A). The triple blend gave two times higher hydrolysis of avicel or A-WS than the individual enzyme preparations at the same enzyme dosages. In all cases the individual or cumulative FPU or protein in blends was equal (10 FPU or 20 mg protein per g of substrate). Increased enzyme activities (CMCase and FPU) were found in the blends compared to the sum of individual enzyme activities added for the blend preparation. This reveals that the increased hydrolysis of cellulose by blends was a result of increased synergism between the same (endoglucanase) and/or different types of cellulases from different preparations. Enzyme blending is thus a facile, cost effective and sustainable approach for biomass saccharification for biofuels.

Biohydrogen production from a novel alkalophilic isolate Clostridium sp. IODB-O3

Hydrogen producing bacteria IODB-O3 was isolated from sludge and identified as Clostridium sp. by 16S rDNA gene analysis. In this study, biohydrogen production process was developed using low-cost agro-waste. Maximum H2 was produced at 37°C and pH 8.5. Maximum H2 yield was obtained 2.54±0.2mol-H2/mol-reducing sugar from wheat straw pre-hydrolysate (WSPH) and 2.61±0.1mol-H2/mol-reducing sugar from pre-treated wheat straw enzymatic-hydrolysate (WSEH). The cumulative H2 production (ml/L), 3680±105 and 3270±100, H2 production rate (ml/L/h), 153±5 and 136±5, and specific H2 production (ml/g/h), 511±5 and 681±10 with WSPH and WSEH were obtained, respectively. Biomass pre-treatment via steam-explosion generates ample amount of WSPH which remains unutilized for bioethanol production due to non-availability of efficient C5-fermenting microorganisms. This study shows that Clostridium sp. IODB-O3 is capable of utilizing WSPH efficiently for biohydrogen production. This would lead to reduced economic constrain on the overall cellulosic ethanol process and also establish a sustainable biohydrogen production process.

Ionic liquid pretreatment of biomass for sugars production: Driving factors with a plausible mechanism for higher enzymatic digestibility

In this study, five ionic liquids (ILs) have been explored for biomass pretreatment for the production of fermentable sugar. We also investigated the driving factors responsible for improved enzymatic digestibility of various ILs treated biomass along with postulating the plausible mechanism thereof. Post pretreatment, mainly two factors impacted the enzymatic digestibility (i) structural deformation (cellulose I to II) along with xylan/lignin removal and (ii) properties of ILs; wherein, K-T parameters, viscosity and surface tension had a direct influence on pretreatment. A systematic investigation of these parameters and their impact on enzymatic digestibility is drawn. [C2mim][OAc] with β-value 1.32 resulted 97.7% of glucose yield using 10 FPU/g of biomass. A closer insight into the cellulose structural transformation has prompted a plausible mechanism explaining the better digestibility. The impact of these parameters on the digestibility can pave the way to customize the process to make biomass vulnerable to enzymatic attack.

Improved saccharification of pilot-scale acid pretreated wheat straw by exploiting the synergistic behavior of lignocellulose degrading enzymes

Requirement of high enzyme dosage for lignocellulosic biomass hydrolysis is one of the challenges for the viability of the second generation bioethanol technology. Here, an optimal enzyme mixture was developed by partially replacing the cellulase proportion with accessory enzymes (β-glucosidase, xylanase, pectinase, laccase) and its hydrolytic performance was compared with different commercial counterparts for the saccharification of pretreated wheat straw (PWS) using a 250 kg per day continuous pilot plant. Maximum degree of synergism was observed with xylanase followed by pectinase, laccase, and β-glucosidase. The statistically optimized enzyme mixture enhanced hydrolysis by 51.23% and 40.66% in 6 h and 24 h, respectively. This study elucidates that presence of even small amount of oligomers and cellobiose pose a strong inhibition for the enzymes. Therefore, development of an optimal enzyme formulation is a sustainable approach to reduce overall enzyme loading for biomass saccharification.

Pilot scale dilute acid pretreatment of rice straw and fermentable sugar recovery at high solid loadings

The aim of this work was to study the dilute acid pretreatment of rice straw (RS) and fermentable sugar recovery at high solid loadings at pilot scale. A series of pretreatment experiments were performed on RS resulting in >25wt% solids followed by enzymatic hydrolysis without solid-liquid separation at 20 and 25wt% using 10FPU/g of the pretreated residue. The overall sugar recovery including the sugars released in pretreatment and enzymatic hydrolysis was calculated along with a mass balance. Accordingly, the optimized conditions, i.e. 0.35wt% acid, 162°C and 10min were identified. The final glucose and xylose concentrations obtained were 83.3 and 31.9g/L respectively resulting in total concentration of 115.2g/L, with a potential to produce >50g/L of ethanol. This is the first report on pilot scale study on acid pretreatment of RS in a screw feeder horizontal reactor followed by enzymatic hydrolysis at high solid loadings.

Life cycle assessment of rice straw utilization practices in India

The aim of this study is to find potential utilization practice of rice straw in India from an environmental perspective. Life cycle assessment (LCA) is conducted for four most realistic utilization practices of straw including: (1) incorporation into the field as fertilizer (2) animal fodder (3) electricity (4) biogas. The results show that processing of 1 ton straw to electricity and biogas resulted in net reduction of 1471 and 1023kg CO2 eq., 15.0 and 3.4kg SO2 eq. and 6.7 and 7.1kg C2H6 eq. emissions in global warming, acidification and photochemical oxidation creation potential respectively. Electricity production from straw replaces the coal based electricity and resulted in benefits in most of the environmental impacts whereas use as an animal fodder resulted in eutrophication benefits. The burning of straw is a harmful practice of managing straw in India which can be avoided by utilizing straw for bioenergy.

Evaluation of recalcitrant features impacting enzymatic saccharification of diverse agricultural residues treated by steam explosion and dilute acid

Exploring agricultural biomass for biofuel production necessitates pretreatment as a prerequisite step. However, due the variability in recalcitrance among biomasses, choosing an optimum pretreatment methodology suitable for multiple feedstocks is challenging. To assess which parameters of pretreated biomass may serve as useful indicators of potential subsequent enzymatic saccharification, an insight into the structural alteration during pretreatment and its impact on the downstream process is essential. In this study, two pretreatment methods, dilute acid (DA) and steam explosion (SE) have been employed on three different biomasses viz. rice straw (RS), cotton stalk (CS) and mustard stalk (MS). The alteration in recalcitrant features of the pretreated residues was measured by chemical analysis, XRD, BET and FT-IR. FT-IR proved useful to measure the cellulose related properties viz. lateral order index (LOI) and hydrogen bond index (HBI) besides lignin related features, i.e. cross-linked lignin (CLL), lignin/cellulose (L/C) and syringyl/guaiacyl (S/G) ratio. The results show that S/G ratio, specific surface area and HBI of the pretreated residues had a positive correlation with enzymatic saccharification across different biomasses and pretreatment methodologies employed. On the other hand, lignin content, CLL, L/C ratio and LOI showed a negative correlation. However, the extent of xylan removal showed a positive correlation with the enzymatic saccharification only when a single pretreatment method was applied to different biomasses. The structure-activity correlation presented here would help to assess and predict the enzymatic saccharification while applying DA or SE pretreatment methods on different biomasses. This correlation could provide assistance in designing an optimum technology.