Microbe assisted depolymerization of lignin rich waste and its conversion to gaseous biofuel

https://doi.org/10.1016/j.jenvman.2021.113684Get rights and content

Highlights

  • Potential lignin depolymerizing consortia (LDC) enriched through forced adaption.

  • LDC showed lignin breakdown ability irrespective of lignin source and nature.

  • Lignin rich residue yielded higher biogas (424 ml/g VS) after hydrolysis using LDC.

  • LDC is also effective on commercial lignins yielding biogas of 302–324 ml/g VS.

  • LRR from bioethanol plants found to be potential feedstock for biogas production.

Abstract

Biomethanation potential of lignin rich residue (LRR) obtained from lignocellulosic ethanol fermentation was evaluated after subjecting to microbe assisted pretreatment using selectively enriched lignin depolymerizing consortia (LDC). The efficiency of LDC in lignin depolymerization was established using alkali and dealkali lignins (AL and DL) along with LRR as feedstocks. Microbial growth on media having lignin as sole carbon source, activity of lignin depolymerizing enzymes, viz., lignin peroxidase and laccase, ability of culture to decolorize the lignin mimicking dyes like methylene blue and ramezol brilliant blue, were considered to confirm the efficiency of enriched mixed culture. Microbial treatment using LDC showed significant positive impact on lignin breakdown irrespective of the substrate (LRR, 46.33%; AL, 31.37%; DL, 34.20%). The hydrolysate of LRR obtained from microbial pretreatment showed higher biogas yield (424 ml/g VS) owing to the efficiency of lignin depolymerization and availability of readily available biodegradable components in residual lignin from previous processing. Depolymerization of commercial lignins also produced a good amount of biogas (302–324 ml/g VS) after pretreatment with LDC. Overall, an additional energy conversion efficiency of about 11.75 kJ/g VS was obtained by valorizing the residual lignin through integrating biomethanation technology to ethanol fermentation. Outcome of this study indicated the feasibility of using lignin rich residue generated from the second generation cellulosic bioethanol plants as a potential feedstock to meet the current gaseous fuel demands. This integration also helps in closing the biomass based biorefinery loop and also promotes the circular economy.

Introduction

Increasing concern over depleting fossil fuel resources combined with the escalating focus towards scientific battle against the climate change as well as to meet the sustainability goals has led researchers towards biofuels development using biomass, especially agricultural residues, as feedstock (Khosravanipour et al., 2018; Liu et al., 2017; Shrestha et al., 2017). Though, many ways have been identified for the conversion of cellulose and hemicelluloses fractions of biomass into bioenergy/biofuels, still effective utilization of residual lignin remains a major challenge for the economic viability and commercial success of biomass based biofuel technologies (Lee, 1997; Gutiérrez et al., 2020; Diego-Díaz et al., 2021). Lignin makes about 15–30% of the biomass fractions by weight and 40% by energy. About 800 to 1000 million MTPA rice straw is produced globally in which about 80% is produced in Asia alone, while wheat straw has an annual global production of 529 million MTPA (Govumoni et al., 2013; Kapoor et al., 2016; Karimi et al., 2006; Vasiliki et al., 2017). Ethanol fermentation of rice straw produces about 0.55–0.58 MT residue/MT of straw and is rich in carbon and nutrient (Ishizaki and Hasumi, 2014). The intended development of 79 billion liters of second generation biofuels (lignocellulosic ethanol) per annum in the USA was estimated to generate about 62 million MT of lignin (Langholtz et al., 2014). A typical bioethanol plant based on corn stover with a production capacity of 2000 TPA, will produce about 70,000 TPA lignin rich residue (Davis et al., 2013), which shows the left over energy potential of biomass. The maize cob is one of the highest available lignocellulosic waste feedstock worldwide and it can be potential option for anaerobic digestion as such or along with other low nitrogen bearing lignocellulosic wastes (Surra et al., 2019). Though, the lignocellulosic ethanol production has been established long back at pilot scale, commercial success of these plants is hindered mainly due to leaving behind the energy rich lignin residue as waste. Integration of novel approaches that can valorize the lignin rich residues to the lignocellulosic ethanol plant will contribute significantly for the commercial success of these plants and also help in establishing circular biobased economy (Lee, 1997; Xu et al., 2019). This will also redefine the biorefinery based energy landscape through balancing between the energy demand and environmental catastrophe (Beckham et al., 2016). Among various approaches, anaerobic digestion is being considered as potential option to valorize the lignin rich residue that can significantly improve the economic viability of the biomass (agricultural residues) (Davis et al., 2013; de Gonzalo et al., 2016; Zhang et al., 2009).

Anaerobic digestion, also named as biomethanation, is a proven technology for waste management where the organic content of waste is converted to biogas through microbial action. Biomethanation/anaerobic digestion is carried out by synergistic association of a group of microbes in 4 different stages, viz., hydrolysis, acidogenesis, acetogenesis and methanogenesis (Fitzgerald, 2018; Kreuger et al., 2011; Yuan et al., 2019; Zhou et al., 2019). Anaerobic digestion has been studied well using wide range of waste/wastewater but reports on the biomethanation of lignin rich waste are limited (DeAngelis et al., 2011; Ko et al., 2009; Mulat et al., 2018). Depolymerization of recalcitrant, irregular, complex, and highly heterogeneous aromatic structured lignin is the major challenge for establishment of full scale process for anaerobic digestion of lignin waste (Pu et al., 2013; Ragauskas et al., 2014).

Several methods were reported for lignin depolymerization including thermo-chemical, hydrothermal (Belkheiri et al., 2018), acid hydrolysis (Liu et al., 2018), pyrolysis (Liu et al., 2008; Windt et al., 2009), hydrogenolysis (Huber et al., 2006), gasification (Huber et al., 2006), etc (Duan et al., 2016; Shi et al., 2013; Tamminen et al., 2010; Verma and Ekka, 2018). Each of these methods have their own limitations like cost and energy intensive nature, not environmentally benign and generating secondary waste which is again a major issue of disposal (Khan and Ahring, 2020; Mulat et al., 2018). Alternatively, biological methods based on fungi and enzymes were also attempted for lignin breakdown that are sustainable, eco-friendly as well as energy and cost effective in nature, and were extensively studied (Kong et al., 2017; Reiter et al., 2013; Vicuña et al., 1993; Wesenberg et al., 2003). But both the methods are lagging behind due to some limitations like requirement of reducing agents and recyclability in case of enzyme, slow reaction rates and difficulty in withstanding the extreme conditions in case of fungi. On contrary, the bacterial metabolic machinery was identified as robust and can withstand the extreme changes (pH variation, redox imbalance, concentrations of phenolics, etc.) in the media during breakdown of lignin residues and its metabolites due to the environmental adaptability and biochemical versatility of bacteria (Mulat et al., 2018). There are very limited studies in this direction of lignin breakdown and valorization. Understanding of bacterial metabolism for lignin breakdown followed by anaerobic digestion under controlled conditions can open potential applications beyond combustion of the lignin rich residue (Taherzadeh and Karimi, 2008). Some bacterial species that were reported for their efficiency of lignin depolymerization include, Streptomyces, Pseudomonas, Athrobacter, Acinetobacter, Rhodococcus etc., through extracellular enzymes (Beckham et al., 2016; Duan et al., 2016; Mulat et al., 2018; Susmel and Stefanon, 1993). Rhodococcus jostii RHA1 reported to act on β-O-4 linkages of lignin by secreting specific peroxidases (Sainsbury et al., 2015). R.jostii was reported for its robustness as it can withstand adverse conditions such as low nutrient, high cell density and has ability to tolerate toxic metabolites concentrations. Pseudomonas putida is another potential candidate for lignin degradation to low molecular weight chemicals (Lieder et al., 2015; Martínez-García et al., 2014). Other bacterial species like Bacillus sp. (Abd-Elsalam, 2009) Novosphingobium sp. (Chen et al., 2012), and Citrobacter freundii (Chandra and Bharagava, 2013), are also exhibited the lignin degradation abilities. However, use of enriched selective consortia for lignin depolymerization is need of the hour considering the valorization of lignin rich residue from bioethanol plant. The lignin rich streams obtained after ethanol production are heterogeneous and contain leftover enzymes, organic acids, fermentation intermediates, along with lignin based compounds which are resisting its direct use in many applications. A meritorious way for funnelling these lignin residues into target chemicals through microbial depolymerization for complete utilization of lignin is essential. This further provides an opportunity to completely utilize the biomass except its ash content for biofuel production. In this context, an attempt was made in this study with the following objectives:

  • (i)

    To evaluate the efficiency of selectively enriched microbial consortia for their ability to depolymerise lignin

  • (ii)

    Pretreatment of lignin rich residue with the enriched microbial consortia in comparison with hydrothermal and thermochemical methods was studied.

  • (iii)

    To establish the lignin depolymerization ability of selectively enriched consortia using commercial lignins, viz., alkali and de-alkali lignins.

  • (iv)

    To convert the depolymerized lignin rich residue into methane via anaerobic digestion

Section snippets

Substrate selection

Lignin rich residue (LRR) was obtained from operating bioreactor after ethanol fermentation of rice straw, through mild acid pretreatment followed by enzymatic hydrolysis. The LRR obtained as slurry and used after removing the residual alcohol content, if any, by heating on stirrer plate at 80 °C for 2 h. Alkali lignin (AL) and de-alkali lignin (DL) were procured from M/s TCI chemicals. All other chemicals used in this study were analytical grade and used as received.

Biocatalyst

Two different microbial

Characterization of substrates

The detailed characterization of each selected substrate was carried out prior to experimentation (Table 2). LRR showed high moisture (87.98%) and low TS (15.44%) content while AL and DL, showed significantly low moisture content (<15%) and high TS (80–90%). Hence, experiments with AL and DL were carried out at high dilution factors but on contrary, LRR based experiments conducted at 50% dilution only. Overall, the TS content for experiments was maintained about 4.0 ± 0.4 g/l. In addition to

Conclusions

Selectively enriched LDC depicted higher lignin depolymerization over corresponding hydro-thermal, acid and peroxide pretreatments, with commercial lignins as well as the lignin rich residue obtained from bioethanol plant. Peroxide treatment was more effective than hydro-thermal and acid pretreatment, while LDC dominated all other pretreatment methods in lignin depolymerization. All the substrates resulted in higher biogas yields after pretreatment with LDC along with significantly higher

Credit author statement

Ms. N.L. Radhika: Involved in conducting experiments, data generation & compilation, manuscript preparation and corrections. Prof. Sarita Sachdeva: Involved in manuscript corrections and finalization. Dr. Manoj Kumar: Involved in experimental planning, data interpretation, manuscript corrections and finalization.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

NLR thank Indian Oil R&D for providing the lab space and facility to carryout the research.

References (76)

  • M. Kaur et al.

    An integrated approach for efficient conversion of Lemna minor to biogas

    Energy Convers. Manag.

    (2019)
  • J.J. Ko et al.

    Biodegradation of high molecular weight lignin under sulfate reducing conditions: lignin degradability and degradation by-products

    Bioresour. Technol.

    (2009)
  • E. Kreuger et al.

    Bioconversion of industrial hemp to ethanol and methane: the benefits of steam pretreatment and co-production

    Bioresour. Technol.

    (2011)
  • J. Lee

    Biological conversion of lignocellulosic biomass to ethanol

    J. Biotechnol.

    (1997)
  • Q. Liu et al.

    Mechanism study of wood lignin pyrolysis by using TG–FTIR analysis

    J. Anal. Appl. Pyrolysis

    (2008)
  • X. Liu et al.

    Anaerobic digestion of lignocellulosic biomasses pretreated with Ceriporiopsis subvermispora

    J. Environ. Manag.

    (2017)
  • G. Palmieri et al.

    Remazol Brilliant Blue R decolourisation by the fungus Pleurotus ostreatus and its oxidative enzymatic system

    Enzym. Microb. Technol.

    (2005)
  • S. Rabodonirina et al.

    Degradation of fluorene and phenanthrene in PAHs-contaminated soil using Pseudomonas and Bacillus strains isolated from oil spill sites

    J. Environ. Manag.

    (2019)
  • P.C. Sahoo et al.

    Accelerated CO2 capture in hybrid solvent using co-immobilized enzyme/complex on a hetero-functionalized support

    J. CO2 Util.

    (2017)
  • P.D. Sainsbury et al.

    Chemical intervention in bacterial lignin degradation pathways: development of selective inhibitors for intradiol and extradiol catechol dioxygenases

    Bioorg. Chem.

    (2015)
  • S. Shrestha et al.

    Biological strategies for enhanced hydrolysis of lignocellulosic biomass during anaerobic digestion: current status and future perspectives

    Bioresour. Technol.

    (2017)
  • S. Srikanth et al.

    Regulating feedback inhibition caused by the accumulated acid intermediates during acidogenic hydrogen production through feed replacement

    Int. J. Hydrogen Energy

    (2014)
  • E. Surra et al.

    Biomethane production through anaerobic co-digestion with Maize Cob Waste based on a biorefinery concept: a review

    J. Environ. Manag.

    (2019)
  • P. Susmel et al.

    Aspects of lignin degradation by rumen microorganisms

    J. Biotechnol.

    (1993)
  • S. Tayibi et al.

    Coupling anaerobic digestion and pyrolysis processes for maximizing energy recovery and soil preservation according to the circular economy concept

    J. Environ. Manag.

    (2021)
  • R. Vicuña et al.

    Ability of natural bacterial isolates to metabolize high and low molecular weight lignin-derived molecules

    J. Biotechnol.

    (1993)
  • D. Wesenberg et al.

    White-rot fungi and their enzymes for the treatment of industrial dye effluents

    Biotechnology Advances

    (2003)
  • M. Windt et al.

    Micro-pyrolysis of technical lignins in a new modular rig and product analysis by GC-MS/FID and GC × GC-TOFMS/FID

    J. Anal. Appl. Pyrolysis

    (2009)
  • Y.R. Wu et al.

    Characterization of anaerobic consortia coupled lignin depolymerization with biomethane generation

    Bioresour. Technol.

    (2013)
  • Z. Xu et al.

    Recent advances in lignin valorization with bacterial cultures: microorganisms, metabolic pathways, and bio-products

    Biotechnol. Biofuels

    (2019)
  • H.-D. Youn et al.

    Role of laccase in lignin degradation by white-rot fungi

    FEMS Microbiol. Lett.

    (1995)
  • H. Yuan et al.

    Effect of low severity hydrothermal pretreatment on anaerobic digestion performance of corn stover

    Bioresour. Technol.

    (2019)
  • H. Zhou et al.

    Anaerobic digestion of aqueous phase from pyrolysis of biomass: reducing toxicity and improving microbial tolerance

    Bioresour. Technol.

    (2019)
  • H. Abd-Elsalam

    Lignin biodegradation with ligninolytic bacterial strain and comparison of Bacillus subtilis and Bacillus sp isolated from Egyptian soil American-eurasian

    J. Agric. & Environ. Sci.

    (2009)
  • F.S. Archibald

    A new assay for lignin-type peroxidases employing the dye

    Azure B. Appl. Environ. Microbiol.

    (1992)
  • G.T. Beckham et al.

    Opportunities and challenges in biological lignin valorization

    Curr. Opin. Biotechnol.

    (2016)
  • T. Belkheiri et al.

    Hydrothermal liquefaction of kraft lignin in subcritical water: influence of phenol as capping agent

    Energy Fuels

    (2018)
  • A.F. Billings et al.

    Genome sequence and description of the anaerobic lignin-degrading bacterium Tolumonas lignolytica sp. nov

    Stand. Genomic Sci.

    (2015)
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