Enhancement of methane production in anaerobic digestion process: A review

Applied Energy 240 (2019) 120–137

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Enhancement of methane production in anaerobic digestion process: A review

T

Yue Lia, Yinguang Chena,b, , Jiang Wuc, ⁎

⁎

a

State Key Laboratory of Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Tongji University, 1239 Siping Road, Shanghai 200092, China b Shanghai Institute of Pollution Control and Ecological Security, Shanghai 200092, China c College of Architecture and Urban Planning, Tongji University, 1239 Siping Road, Shanghai 200092, China

HIGHLIGHTS

GRAPHICAL ABSTRACT

advances in increasing hydro• Recent lysis are introduced. for enhancing enzyme ac• Strategies tivity relevant with VFA generation are reviewed.

of direct interspecies • Acceleration electron transfer facilitates methanogenesis.

methane production • Improving through special pathway is beneficial to anaerobic digestion.

assessment of ap• Techno-economic proaches in the lab-scale is presented.

ARTICLE INFO

ABSTRACT

Keywords: Anaerobic digestion Methane Hydrolysis Acidification Microorganism Interspecies electron transfer

With the increase of energy consumption and wastes generation due to human activities, anaerobic digestion (AD), a technology which turns wastes into bio-energy, is receiving more and more attention in the world. It is well known that there are at least three stages involved in anaerobic digestion, i.e., hydrolysis, acidification, and methanogenesis. Until now, however, the advances in enhancing acidification and methanogenesis have not been reviewed. This review provides a comprehensive overview of the methods reported to enhance each step involved in anaerobic digestion. More important, enzymes are the key to anaerobic digestion, and the strategies for improving enzyme activity are summarized. As electron transfer has been reported to play an important role in anaerobic digestion, the research progress of the approaches for the acceleration of direct interspecies electron transfer in methane production is also introduced. In addition, the recent advances in increasing the reduction of carbon dioxide to methane, which has been widely observed in methanogenesis step, are reviewed. Furthermore, the techno-economic assessment of anaerobic digestion is made, and the key points for future studies are proposed.

⁎ Corresponding authors at: State Key Laboratory of Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Tongji University, 1239 Siping Road, Shanghai 200092, China (Y. Chen). E-mail address: [email protected] (Y. Chen).

https://doi.org/10.1016/j.apenergy.2019.01.243 Received 30 June 2018; Received in revised form 24 January 2019; Accepted 30 January 2019 0306-2619/ © 2019 Elsevier Ltd. All rights reserved.

Applied Energy 240 (2019) 120–137

Y. Li, et al.

1. Introduction

volatile fatty acid (VFA), accompanied by cellular materials generation. In the third step, hydrogen-producing acetogens metabolize hydrolytic products to acetic acid with the hydrogen and carbohydrate yield. A small number of homo-acetogenic bacteria utilize CO2/H2 as substrates to form acetic acid. Finally, the acidification products (such as acetic acid, formic acid, CO2/H2, etc.) are turned into methane by strictly anaerobic methanogens (see Eqs. (1)–(5)). Methanogenesis is a complex phenomenon accomplished by the synergistic action of various mesophilic bacterial species.

Along with the development of industry, a large amount of fossil fuels is consumed which caused climate change and deficiency of energy, thus human begins to quest for renewable energy sources that lessen greenhouse gases emission [1]. The strong dependency of fossil fuels contributes to a significant change in energy cost due to the variation of fuel cost, which in turn may result in economic and political challenges [2]. Therefore, Bioenergy with low greenhouse gas emission and lower price which meets growing energy demand plays a central role in promoting renewable alternatives. As renewable bioenergy, methane can be generated under anaerobic conditions from various substrates, such as sewage sludge, food waste, forestry resource, living stock manure, and wastewater. The quantity of waste production is amazing due to daily human activities and the progress of the industry. For example, the municipal organic waste produced has reached more than 150 million tons in China each year [3] and was estimated by World Bank to increase to 2.2 billion tons all over the world by 2025 [4]. Rice straw in China was reported to produce annually ranging between 180 and 270 million tons [5]. The National Development and Reform Commission has estimated that about 3.8 billion tons of livestock manure are produced annually in China [6]. Over 11.2 million tons of dry sludge is generated annually in China and almost 80% of it has not been stabilized [7]. Thus, municipal solid waste and domestic sewage should have reasonable disposal to cut down the environmental implication. It is widely known that anaerobic digestion is the most sustainable, cost-effective technology for waste treatment and energy recovery in the form of biofuel [8] which not only minimizes the amount of waste but also transforms it into bioenergy. As clean energy, biogas can replace fossil fuels which generate greenhouse gases via combustion in the household and commercial activities [9]. What’s more, the digestate of anaerobic digestion are rich in nutrients and it can be served as a fertilizer to the crop cultivation. Therefore, enhancing methane production from various waste can obtain more energy to compensate for the deficiency of non-regenerated energy with consumption the same quantity of the substrate. Anaerobic digestion (AD) contains hydrolysis, acidogenic fermentation, hydrogen-producing acetogenesis, and methanogenesis (Fig. 1). In the first step, complex organic substances that can’t be directly utilized by bacteria are decomposed into soluble monomers with the effect of an extracellular hydrolytic enzyme of acidogens. Acidification is also called fermentation which serves intermediate from substrate metabolism as an electron acceptor. In this process, acidogenic fermentation bacteria convert soluble monomers into terminal products, such as

CH3COO–+H2O → CH4 + HCO3– +

–

(1)

HCO3 +H →CH4 + 3H2O

(2)

4CH3OH → CO2 + 2H2O

(3)

-

+

4HCOO +2H →CH4 +

CO2 + 2HCO3–

4H2 + CO2 → CH4 + 2H2O

(4) (5)

In anaerobic digestion, due to the limitation of oxygen, a lot of microbes oxidize organic substances in a limited condition and then creates an equilibrium of the oxidative and reductive reaction. The major species for methane production is methanogens, so maintaining their activity is prerequisite for methane enhancement. Methanogens should be maintained in a stable condition due to the poor adaption to the variation of pH and the optimum pH for them is 6.5–7.5. An anaerobic environment is one of the basic condition for strictly anaerobic methanogens growth using oxidation–reduction potential (ORP) as the basis of judgment [11]. Organic loading rate directly reflects the equilibrium between substrate and microorganisms which also needs to be regulated to maintain the balance between acidification and methanogenesis [12]. The range of suitable temperature is wide because methanogens in the rate-limiting methanogenic stage are mainly divided into a mesophilic bacterium (Mb. Bryantii, Ms. Barkeri and Mc. vanniielii) and thermophilic bacteria (Mt. thermophila and Ms. thermophila) [13–15]. In the biological treatment, due to the close relationship between substrate and microorganisms, remarkable biological treatment efficiency has been reported to accompany by the high sludge concentration. Due to the mass transfer in the substrate, its accessibility for methanogens is a key factor for methanogenesis. In addition, enough nutrients are indispensable and trace inhibitors will cause harmful influence on microbes [16,17]. Anaerobic digestion is a process that waste is transformed into energy, which, however, is consumed in AD. Therefore, input energy should be lower than output energy so that this biological treatment makes sense. Methane is deemed to a renewable energy in the emerging

Fig. 1. Flow chart of anaerobic digestion (revised according to Wang et al. [10]). 121

Two-stage and supplement of organic waste Microwave irradiation

Wastewater sludge

Paper mill Municipal wastewater treatment plant N/A

Microwave irradiation and NaOH Microbial electrolysis cell with iron-graphite electrode

Electrohydrolysis pretreatment

Thermal pre-treatment

Thermal pre-treatment

Hydrothermal

Inoculation Feedstocks adjustment

Two-stage Activated carbon

Cysteine

Novel microbes Thermal

Rumen bacteria pulsed electric field Thermochemical

Lipase purple photosynthetic bacteria

Two-phase leach-bed process Fungal pre-treatment

Fungal pre-treatment

Inoculation

Alkali hydrogen peroxide

Temperature phase

Waste activated sludge

Lignocellulose waste pulp and paper mill sludge Primary and secondary sludge

Primary and secondary sludge

Fruit and vegetable waste

Canteen waste Canteen waste

Canteen waste Canteen waste

Canteen waste

Food waste Cow manure

Cow manure Pig slurry Pig slurry

Swine slaughterhouse waste Cattle manure leachate

Dairy manure leachate Chicken manure Rice straw

Yard waste

Yard waste

Cotton stock

Grass silage

Waste activated sludge

Microwave irradiation and CaO2

Waste activated sludge

122 Farm

Ohio Agricultural Research and Development Center University of South Florida campus Farm

Dairy cattle breeding base Biogas plant Farm

Slaughterhouse Local dairy cattle farm

Barn Pig farm Poultry farm

CSRM Breed plants

Dining hall

Students’ dining hall Canteen

School canteen Canteen

Farmers market

Municipal wastewater treatment plant Wastewater treatment plant and laboratory UASB reactor

Sewage treatment plant

Microwave irradiation and H2O2

Dairy effluent treatment plant Dairy wastewater treatment plant

Wastewater treatment plant

Source

Waste activated sludge

Waste activated sludge

Treatment

Feedstock

Table 1 Different methods for enhancement of methane production.

TS = 91.08 ± 0.00%, VS = 88.12 ± 0.01%, Cellulose = 50.42 ± 0.86%, Hemicellulose = 15.64 ± 0.03%, Lignin = 16.32 ± 0.40% TS = 27.0%, VS = 91%

TS = 14.58%, VS = 10.63%, Fibers = 21.5%TS, Proteins = 20.0%TS, Lipids = 14.4%TS TS = 9.15%, VS = 7.72%, Fibers = 35.2%TS, Proteins = 12.9%TS, Lipids = 15.2%TS TS = 15.01 ± 0.98%, VS = 14.18 ± 0.52%, Proteins = 3.58 ± 0.15% TS = 24.7%, VS = 95.2%TS, Carbohydrate = 34.3%, Protein = 22.4%, Lipid = 43.3% TS = 26.0%, VS = 24.1%, CODCr = 1.5 g/gVS TS = 28.60 ± 0.08%, VS = 27.42 ± 0.13%, Cellulose = 0.52 ± 0.03%, Hemicellulose = 1.46 ± 0.05%, Ligin less than 0.1% TS = 88.95 ± 4.74 g/L, VS = 82.90 ± 3.82 g/L, tCOD = 135.50 ± 7.50 g/L, tPolysaccharide = 81.96 ± 3.78 g/L, tProtein = 11.73 ± 1.51 g/L. VS = 71.7 ± 4.5%, COD = 19.6 g/L TS = 34.66%, VS = 19.52%, Fibers = 18.7%TS, Proteins = 0.6%TS, Lipids = 12.3%TS TS = 15.1 ± 0.08%, VS = 13.1 ± 0.08%, sCOD = 13250 ± 315 mg/L TS = 3.93%, VS = 73.62%, COD = 28000 mg/L TS = 88 ± 4 g/kg, VS = 55 ± 2 g/kg, tCOD = 93 ± 4 g/kg, sCOD = 13.9 ± 0.4 g/kg, TS = 91.70 ± 0.05%, VS = 91.52 ± 0.03% TS = 28.02 ± 6 g/L, TSS = 6.05 ± 1 g/L, VSS = 3.50 ± 0.2 g/L, BOD = 28 g/L, tCOD = 100 ± 20 g/L, sCOD = 40 ± 10 g/L TS = 6.60 ± 0.01%, VS = 4.98 ± 0.09%, Lignocellulose = 37.70 ± 1.32% TS = 25%, VS = 17% TS = 89.9 ± 0.2%, VS = 80.6 ± 0.2%, Cellulose = 37.8 ± 0.2%, Hemicellulose = 29.6 ± 0.7%, Lignin = 14.8 ± 0.4% TS = 94.3 ± 0.1%, VS = 98.9 ± 0.2%, Cellulose = 30.8 ± 0.9%, Hemicellulose = 15.9 ± 0.5% TS = 50.8 ± 3.4%, VS = 47.7 ± 4.4%

TS = 48.470 g/L, VS = 32.332 g/L

tCOD = 22000 mg/L, sCOD = 905 mg/L, TS = 11660 mg/L, VS = 9116 mg/L, SS = 4680 mg/L, sProteins = 780 mg/L, sCarbohydrates = 320 mg/L TS = 22000 ± 300 mg/L, tCOD = 24000 ± 400 mg/L, sCOD = 400 ± 10 mg/L, SS = 20000 + 200 mg/L, VS = 16150 ± 300 mg/L, sProteins = 41.3 ± 0.5 mg/L, sCarbohydrates = 5.2 ± 0.1 mg/L TS = 12950 ± 320 mg/L, VSS = 8620 ± 122 mg/L, tCOD = 9290 ± 130 mg/L, sCOD = 120 ± 10 mg/L, tProteins = 4980 ± 68mgCOD/L, tCarbohydrates = 1012 ± 49mgCOD/L TS = 29,615 ± 247 mg/L, VS = 17,120 ± 127 mg/L, tCOD = 30,640 ± 1,923 mg/L, sCOD = 340 ± 141 mg/L TSS = 117.9 ± 8.1 g/L, VSS = 77.2 ± 3.9 g/L, tCOD = 114.2 ± 9.2 g/L, sCOD = 4.1 ± 0.6 g/L, tProteins = 29.0 ± 1.8 g/L, tPolysaccharides = 13.5 ± 1.0 g/L TS = 38.97 g/L, VS = 28.87 g/L

VSS = 23.1 ± 0.9 g/L, sCOD = 4.1 ± 1.0 g/L

Characterization of substrate

80.2a 102.7a 22.4a 13.8a b

85.2b 16.1a a

70.1 63.0b 26.3a 41.0a 43.9a a

9.6a

250.0a

296.9a 170.2a 303.0a a

583.3a 326.0a a

502.9a 466.0a 472.0a a

35.5a

72.7a 254.3a

102.6a 192.4a

28.0

141.1b

44.6a

238.0

209.0a 230.0a 107.1b

124.5a 272.0a 263.0a

b

18.4a 36.4b

851.6a 428.6b

a

103.3b 58.0a 151.0b

14.6 13.3a 138.0a 740.6a 374.0a

102.0 238.0a

724.5 626.9a

130.0

54.5

64.0

285.0

a

[62]

[61]

[60]

[59]

[56] [57] [58]

[54] [55]

[51] [52] [53]

[50] [44]

[49]

[47] [48]

[45] [46]

[44]

[43]

[42]

[41]

[40]

[39]

[38]

[37]

[36]

[35]

reference

(continued on next page)

347.0a

350.0a a

Methane enhancement (%)

Methane yield (mL/g VS)

Y. Li, et al.

Applied Energy 240 (2019) 120–137

Applied Energy 240 (2019) 120–137

120.1b TS = 27.36 ± 0.53%, VS = 21.37 ± 0.41%,

[69] 14.7a

114.1b SS = 6.29 ± 0.18 g/L, VSS = 3.96 ± 0.12 g/L

[68] 78.3a

1500.0 428.4b TS = 56.7 g/L, VS = 44.4 g/L TS = 62 g/L, VS = 51 g/L

[66] [67]

48.6a 50.8b

[65]

60.0a 992.4b

There are many kinds of the substrate being applied in anaerobic digestion, such as sludge, food waste, farm waste, agriculture waste, and wastewater. Due to the low practical methane production, various methods have been adopted to enhance methanogenesis. To efficiently improve anaerobic digestion, it is of great importance to be aware of different substrates’ character and their mechanism during the AD. As shown in Table 1, different substrates have different content of carbohydrates and proteins which are the main micromolecules for acetogenic bacteria metabolism. In addition, different methane potential of carbohydrates and proteins results in different methane productions (0.373 versus 0.417 L/g). It is indicated that the proteins have a higher potential for methanogenesis than carbohydrates with the same quantity. The content of them in the substrate is a key factor for methane generation, furthermore, soluble state of them contributes to more biological reactions in the system. Although some kinds of substrates own high COD, their sCOD is low, which goes against the acidification. Therefore, steps should be taken to enhance methanogenesis according to their own characters and mechanism. As for different substrates, limitation of methanogenesis attributes to their different mechanism. Sludge, which full of organic matter, various bacteria, inorganic particles, colloid and so on, is a kind of solid sediment produced after wastewater treatment [25]. However, there are so many kinds of sludge from different place, but some of them may be hard to digest. For example, it is widely accepted that primary sludge is easy to digest while secondary sludge or waste activated sludge (WAS) are not [26]. Most of organics in the sludge are encased with

Reported. Calculated. N/A Not available.

Tryptone-based synthetic wastewater swine wastewater

b

Ferroferric oxide

Slaughterhouse wastewater Vinasse wastewater

a

Archaeal pre-treatment AnSBBR

Fischer-Tropsch wastewater

Pig farm

Magnetite

Molasses wastewater

Two-step heating

NaOH and mechanical pretreatment UASB Wheat straw

Regional sugar-processing factory Coal to Liquid Co, LTD Slaughterhouse Biohydrogen production process Wastewater treatment plant

Treatment

Courtyard

a

400.0 31.0b

b

33.2b 305.0a

TS = 89.53%, VS = 86.83%, Cellulose = 44.30%, Hemicellulose = 32.74%, Lignin = 6.30% TS = 89700 mg/L, TSS = 4269 mg/L, VS = 86200 mg/L, VSS = 4267 mg/L, tCOD = 128400 mg/L, sCOD = 119200 mg/L COD = 30,608.0 ± 843.5 mg/L

[64]

Methane enhancement (%) Methane yield (mL/g VS)

[63]

market which can be transformed into electric energy, heat energy and so on. Energy recovery potential of different substrates is diverse from each other. Thus, enhancement of efficiency in methane generation from different substrates is equivalent to more energy recovery in the anaerobic digestion. Furthermore, heat energy produced by methane can be utilized in the process of anaerobic digestion to adjust the temperature that is the factor that affects anaerobic microbial activity and anaerobic digestion efficiency which decrease the energy consumption for heat. Due to its lower investment and management costs, anaerobic digestion is better choice for energy generation. Because of the inadequacy of energy recovery from waste, strategies are adopted to raise the methane yield for implementing the circular use of resources and achievement of more energy production. In the literature, the progress of anaerobic digestion has been reviewed by some researchers. For example, Mao et al. summarized the factors affecting digestion efficiency, enhancement of biogas production by accelerants (greenery biomass, biological pure culture, and inorganic additives), the configuration of reactors, and type of anaerobic digestion process [18]. As hydrolysis is believed the rate-limited step in anaerobic digestion, most reviews focused on the advance in improving hydrolysis by physical, thermo-chemical, and biological pre-treatments [19–24]. It is well known that there are at least three stages involved in anaerobic digestion, i.e., hydrolysis, acidification, and methanogenesis. Until now, however, the advances in enhancing acidification and methanogenesis have not been reviewed. More important, enzymes are the key to anaerobic digestion, but the strategies for improving enzyme activity have not summarized. Also, although electron transfer has been reported to play an important role in anaerobic digestion, the research progress of the approaches for the acceleration of direct interspecies electron transfer in methane production has never been introduced. In addition, the recent advances in increasing reduction of carbon dioxide to methane, which has been widely observed in methanogenesis step, has not been summarized and reported in the literature. What’s more, the techno-economic assessment is presented in the end that is different from other reviews. All these are the object of this paper. In addition, the key points for future research in the area of anaerobic digestion are proposed. 2. Comparison of methane production from different substrates

Feedstock

Table 1 (continued)

Source

Characterization of substrate

reference

Y. Li, et al.

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extracellular polymeric substances (EPS) secreted by microbes and microbial cell membrane that restrains the rate of hydrolysis step. For this reason, application of anaerobic digestion has been limited which results in long hydraulic retention time (HRT) and low degradation efficiency [27]. Although total COD of sludge is normally high, soluble COD of it which can be utilized to proceed acidogenesis is low. If organic compounds can’t be best utilized in the AD which means substrates can’t be converted into the soluble state, practical methane generation will be lower than the theoretical value. The disintegration of EPS exposures carbohydrates and proteins in the sludge to acidogens and organic matters in the EPS will release into the system which contributes to more short-chain fatty acid generation and then more methanogenesis. Thermal hydrolysis application can achieve partial solubilization of sludge through raising dewaterability. As there are large numbers of organic compounds, such as protein and polysaccharide, in municipal organic wastes, it is attractive to recover valuable bioenergy from food waste [28]. It has been reported that 5% of organic content is used for cell growth during anaerobic digestion and 5% is transformed into heat even if the wastes are homogeneous and completely degradable [29]. Due to inaccessibility of some organic matter acting as binders for organic particles, recalcitrant organic matter, and the fraction of the substrate used for cell growth and maintenance, methane production would be low [30]. Simultaneously, low rate of anaerobic digestion contributes to a higher concentration of toxic substances produced in the process which needs more robust equipment to remain stable. Due to the high content of organic matters, food waste can’t yield too much biomethane as the theoretical value. Some of them are hard to be taken up by acidogens, so improving the transformation of organic matters makes them more accessible to microorganisms. Slaughterhouse waste, manure, and manure leachate are the parts of farm waste which comprise proteins and lipids. With complex chemical structure and high fat content, the lipid is difficult to degradable. Inaccessibility of lipid hinders its transformation and low utilization rate of it decreases the actual output. Some farm waste such as cattle manure waste is an abundant biodegradable waste with low C/N ratio which limits its effectiveness of the AD process. Measures should be taken to lower the harm of farm waste or adjust C/N ratio which is in favor of microorganism growth and more methane will be formed. The high alkalinity of farm waste goes against the growth of microorganism and conditioning of affecting factors enhances methane production. Commonly, wastewater in the slaughterhouse also owns an unbalanced C/N ratio. For those fat enriched wastewater, anaerobic digestion is found to be troublesome due to the potential of sludge floatation, the formation of fat scum layers at the surface of the reactor, which doesn’t digest and the inhibition/toxicity effects of the intermediate compounds (long-chain fatty acid) generated during the anaerobic digestion of the wastewater [31,32]. Adverse factors influence microbes’ metabolism so that the practical value of methane production is lower than that of the theoretical value. Diverse agriculture wastes, such as forestry and agriculture residues, are rich in lignocellulose and it is widely known that decomposition of lignocellulose will produce carbohydrate, and microorganism takes them up for methane yield [33]. However, their lignocellulosic structure hinders the process of hydrolysis. Therefore, the following steps are limited and practical methane production is low. Through some approaches, such as pretreatment and inoculation, carbohydrates that are the substrates for acidogens will be obtained after degradation of lignocellulose and more short-chain fatty acid production is in favor of methanogens to generate methane. Lignocellulose waste has a relatively high carbon to nitrogen ratio and inoculation that owns high nutrients content will lead to high enzymes activity and balanced C/N ratio, which improves methane yield from agriculture wastes[34].

3. Enhancement of hydrolysis Hydrolysis is the first step of anaerobic digestion, acceleration of it facilitates the following step proceeds. When a substrate is low degradable, hydrolysis will be inhibited, especially in treating the waste with high solid content [70]. Thus, steps should be taken to promote the hydrolysis rate and overall performance. For example, physical pretreatment, such as thermal treatment, mechanical treatment, and microwave treatment, and supplement of additives can realize it. Acidification is a step that micromolecules produced in the stage of hydrolysis can be taken up by acidogens and then turned into VFA. With more VFA production, more acetic acid will be formed, which are the direct substrates for methanogens metabolism, and more methane production will be realized in the end. 3.1. Electrical treatment Electrical treatment is widely used in enhancing anaerobic digestion. Pulsed electric field (PEF) has been reported to be a technique dealing with a range of biomass types and shown desired results [71–73]. Likewise, Safavi et al. [52] adopted this technology as a pretreatment of anaerobic digestion of pig slurry to enhance methane production. Hydrolysis stage has been promoted and realize 58% increment of methane. Feng et al. [40] integrated high solid anaerobic digestion with microbial electrolysis cell with the iron-graphite electrode. Fe2+ released in the process enhanced the activity of enzymes involved in hydrolysis which in turn contributed to increasing methane production by 22.4%. Choi et al. [74] also applied different supplemental voltages with microbial electrolysis cell resulting in the highest methane yield of 408.3 mL CH4/g COD glucose, which was 30.3% higher than that in the control. Ding et al. [75] combined microbial electrolysis cell with anaerobic membrane bioreactor and applied different voltage, which led to the increase of the absolute value of sludge particles and a decrease of sludge viscosity. Hydrolysis has been improved because more extracellular polymeric substance-polysaccharides were released into the system. Veluchamy et al. applied electric hydrolysis pretreatment for enhanced methane production from lignocellulose waste in pulp and paper mill sludge, which relied on electrophoresis, electrophoresis, electro-osmosis, and ohmic heating leading to the disintegration of particles and microbial cell lysis [76]. Methane yield was 301 ± 3 mL/g VS and increased by 13.8% after lignocellulosic waste pre-treated with electric hydrolysis. Hydrolysis step is also a focused step, therefore, promotion of hydrolysis to improve methane production will be the key solution for AD amended. PEF uses a rapidly pulsing, high-voltage electric field to disrupt cellular membranes and induce high permeabilization of biological cells, e.g. inactivate microorganisms [77]. As a dielectric, cell membrane has a membrane potential due to charges of opposite polarities distribution between interior and exterior of a cell membrane [78]. Electrical charges build up inside and outside of cell membrane when the cell is exposed to external electric field and then membrane potential increases. With more charges generated, the attraction between negative and positive charges from both sides of the membrane causes pressure to the membrane and attenuate its thickness [77]. When the elastic resistance of membrane reaches the limits, the natural structures of the cell is destroyed, thus organic substances in the cell will be released which stimulate the hydrolysis rate. Consequently, complex organic molecules in waste are converted into simpler forms that become easily accessible for acidogens to generate VFA. PEF pretreatment results in the permeability of the cell walls and the solubilization of organic solids. However, the mechanism of PEF pre-treatment is to produce small colloids from organic solids rather than solubilization, which contributes to more methane production [72].

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3.2. Biological method

and enzymes. Hinds et al. [60] enhanced methane yield in high-solids anaerobic digestion of yard wastes through inoculation with pulp and paper mill sludge (P&P sludge). With a complex microbial community acclimatizing to the high concentration of lignocellulosic wastes, pulp and paper mill sludge inoculated in the digestor accelerates the degradation of lignocellulosic [85,86]. In addition, pulp and paper mill sludge comprise high volatile solids content and granular nature, while sludge from wastewater anaerobic digestion has more dilute and flocculent nature, thus P&P sludge inoculated digestor may have faster process start-up. In a full-scale high-solids anaerobic digestion, yard waste in the digestor maintains the porosity for effective leachate recirculation because promotion in degradation of lignocellulosic biomass breaks its integrity [87]. Lipid-enriched swine slaughterhouse waste pretreated by hydrolysis of lipase enhanced methanogenesis [54]. It was reported that lipid was first hydrolyzed into long-chain fatty acid and glycerol, and further converted to hydrogen and acetate by acetogenic bacteria and then to methane by methanogenic archaea eventually in AD. With complicated structures, lipids will limit the rate of hydrolysis which in turn inhibit the efficiency of the AD process [88]. Hydrolysis is believed to be a ratelimiting step, so usage of lipase stimulates the hydrolysis of the substrate making it more accessible to microorganisms, and then products can be taken up by acetogens via metabolism to generate acetic acid, the predecessor of methane. Except for commercially procured lipase, enzymes directly separated from bacterial strains can also be applied to anaerobic digestion for enhancement of hydrolysis. Kanmani et al. [89] obtained partially purified lipase from staphylococcus pasteuri COM-4A in their laboratory and applied it in pre-treatment of coconut oil mill effluent, which contributed to the decrease of the overall COD load, with the maximum of 89%, via the effect of hydrolytic enzymes on the effluent organic substances. Dilution of the substrates with high organic load was also observed to promote hydrolysis efficiency.

Pre-treatment is reported to enhance the accessibility of substrate and biological treatment with fungal and archaea is almost applied to agriculture wastes, which can change its chemical composition and lignocellulosic structure of lignocellulose wastes, leading to breakage of linkage between polysaccharides and lignin, so that cellulose and hemicellulose are more accessible to microorganism and hydrolytic enzymes [79], as shown in Fig. 2. Thus, fungal pre-treatment with Pleurotus ostreatus and Trichoderma reesei was applied in the anaerobic digestion of rice straw [58]. Biological pretreatment compared to other methods is more environmentally friendly, consumes less energy and chemicals, and operates in a moderate condition, which reduces the generation of substances with inhibitory effects [76]. Due to the presence of fungi in the process, the nutrient transfer is a pivotal factor for fungi growth which depends on moisture content. Therefore, sufficient moisture ensures the growth of fungi and ligninolytic activity and excess moisture content increases the activity of lignin hydrolytic enzymes [80]. Degradation of lignocellulosic biomass in a rising trend with moisture is due to the absorption of water, which softens the substance, reduces the inner cohesive forces so that crystalline cellulose structure is swollen and more accessible to enzymes [81]. Dealt with fungi, P. streatus, lignin fiber was destroyed and secondary cell wall was exposed, which resulted in the increase of pore size and surface area, thus enzymes were easy to migrate through the cell wall. In addition, long incubation time due to the low removal rate of lignin was a barrier to apply fungal pre-treatment. With the treatment of fungi, the resistant character of lignocellulose was reduced by selectively degrading lignin which transformed more carbohydrate into methane [82]. What’s more, mechanical pre-treatment has been applied to decrease the size of particles interfering in the complex substrate hydrolysis and improve the activity and diversity of microorganisms, showing a good efficiency of methane yield due to the high utilization of substrates and more favorable microbes [83]. Inoculation or bioaugmentation with bacteria capable of hydrolyzing lignocellulosic substances is a cost-efficient technique which sometimes puts waste that full of the microorganism into the substrate, contributing to the increment of microbial abundance. The recalcitrance of lignin is a rate-limiting step of anaerobic digestion, and the association of cellulose, hemicellulose, and lignin acts as an obstacle to the microbial populations that perform hydrolytic conversion of cellulose [76]. Yue et al. [84] found that ruminant bacteria from cattle can be used as an inoculum because they are capable of producing extracellular substances that assist cellulolytic biofilms in adhesion and entrance to fibers, and then increase the abundance of relevant nutrients

3.3. Thermal hydrolysis Thermal hydrolysis is proved to be a technology which improves the hydrolysis rate and disintegration of substrates with the application of high temperature and pressure by addition of steam and subsequent decompression [90]. For those high suspended solid wastes full of refractory compounds, macromolecules in the substrate are hydrolyzed into micromolecules via thermal treatment. Choi et al. [91] adopted thermal hydrolysis to enhance methane production from sewage sludge, which was verified to improve sludge disintegration. Passos et al. [92] also found thermal hydrolysis can improve the solubilization in fractions of waste that remained in cell structure and were hardly utilized

Fig. 2. Schematic diagram of pre-treatment enhancing methane production from lignocellulosic wastes. 125

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by microorganisms. Simultaneously, it can decompose complex molecules and they would be easier to be degraded after that. Thermal hydrolysis not only collapses large particulate organics but also discharges the soluble matters from microbial cells. Dwyer et al. [93] found that thermal hydrolysis can facilitate hardly degradable substances such as melanoidins, soluble microbial products, and humic substances, transforming into low molecules which are easy to digest and disappearance of refractory organic matters in the sludge proves this conclusion. Choi et al. [94] suggested that anaerobic digestion of high suspended solid waste with the treatment of thermal hydrolysis created an efficient, robust and resilient system and promoted disintegration and solubilization of organic substances suspended in the reactor. Svensson et al. [95] applied thermal hydrolysis in the anaerobic digestion of sewage sludge and food waste and found it can increase degradation rates and methane production from a wide range of waste. Thermal hydrolysis leads to solubilization of organic compounds and releases degradable waste into liquid system. With the high temperature, this technique reduces some pathogens in the digestor and then facilitates fermentative bacteria for hydrolysis. The amount of soluble COD compared to the total COD can be used as an index of solubilization with thermal hydrolysis pretreatment. Svensson et al. [95] reported that highest solubilization of waste was 113% higher than the lowest solubilization. After pre-treated by this technique, more fractions in the waste become soluble and degradable, which benefits the microorganism metabolism and in favor of hydrolysis.

Application of dosing H2O2 is for the purpose of better usage of microwave disintegration so that microbes can be more accessible to their own substrate. Wang et al. [38] combined another additives, CaO2, with microwave irradiation. Just like the former, microwave irradiation is an efficient technique capable of disrupting sludge flocs and cells and releasing organic matters into aqueous phase [96,101–103] which enhanced the disintegration and solubilization of sludge. Microwave irradiation via thermal and athermal effects disintegrates compounds and breaks recalcitrant complex down into easily degradable ones [104]. Thermal effects mainly occur via denaturation of cell materials, while athermal effects include electroporation, dielectric cell membrane rupture, magnetic field coupling, and selective heating [96]. As a peroxide, CaO2 can also release hydroxyl radicals through decomposing to Ca(OH)2 and H2O2, and then microwave irradiation stimulated the decomposition of H2O2 to the formation of ·OH. Hydroxyl radical is a powerful oxidizing agent which can react with most of the organic substances and promote particulate COD of sludge disintegration. With the effect of hydroxyl radicals, the increment of the activity of protease and amylase contributes to more EPS dissolved via destroying cell wall and releases more hydrolase into the supernatant which will be more easily accessible to the substrate [105]. The major mechanism of the microwave with additives is that hydroxyl radicals generated by peroxide can promote disintegration of sludge and EPS, inducing the release of advantageous enzyme relevant with anaerobic digestion and more substrate taken up by methanogens are formed leading to increment of methane production which makes further efforts with microwave irradiation. From the perspective of methods above, it can be summed up that the acceleration of hydrolysis which is a rate-limiting step plays a crucial role in boosting methanogenesis. Impacts of cell’s destruction result in breakage of sludge turning it into easily degradable state and releases of the enzyme for AD which improves the efficiency of the entire process. A better surface area of sludge is elucidated by dissociation of EPS matrix and discharge of intracellular content into an aqueous phase that creates the opportunities for microbes to access to and make use of it. With regard to pig slurry, the pulsed electrical field can also cause cell destruction and release of organic substances in the cell to promote hydrolysis. Approximately 30% of municipal solid wastes are lignocellulosic green wastes, which are hard to digest, separation of municipal solid waste into biodegradable and non-biodegradable fractions can improve the efficiency of AD. The main aim of enhancing methane production in the agriculture waste digestion is to overcome the obstacle, low degradability of lignocellulosic biomass. Whether treated with different kind of fungi or inoculated with wastes full of microorganism, the key role is to decompose recalcitrant materials into carbohydrate so as to be taken up by hydrolytic bacterial for VFA production. Due to the low removal rate of biomass, a long incubation time of fungi hinders its application on a large scale [106]. However, fungal pretreatment can significantly reduce the required digestion time, potentially contributing to significant economic benefits. Degradation of lignocellulosic materials needs not only the disintegration of lignocellulosic, but also the transformation of sugars, VFA, and then produce methane in the end. Inoculation of P&P sludge that should have been treated and then discarded is conductive to lessen waste materials and enhance the microbial diversity and abundance in the digestor which results in more organic waste being hydrolyzed.

3.4. Radiant and oxidative destruction Microwave irradiation is known as an efficient technique for sludge solubilization of activated sludge to enhance methane formation and the presence of H2O2 can induce its performance [37]. It is noted that sludge hydrolysis is considered as a rate-limiting phase, microwave disintegration has been applied to overcome the obstacle. However, the application of microwave disintegration alone was not enough to significantly enhance hydrolysis of sludge and production of methane, because EPS present on the exterior of sludge flocs reduced the surface area that inhibited the microwave’s effect. Therefore, during the AD process, EPS would make further efforts to limit the access of organic substances to methanogenic microbes. Without sufficient substrates, the quantity of methane production is sure to decrease. As an oxidant, hydrogen peroxide released hydroxyl radicals which can promote the breakdown of sludge and dissociation of EPS via chemical oxidation at very low concentration [96]. H2O2 with a low cost works well in the acid condition, in addition, it can regulate activated sludge effectively in the same condition. Protease and amylase which played a key role in the AD were released from dissociated liquefiable EPS and contributed to sludge disintegration because the major component of microbes’ cell wall in the activated sludge are carbohydrates and proteins. The mechanism of biopolymer’s releasing into liquid medium is that rapid microwave irradiation alters the dipole arrangement of macromolecular compounds, breakage of hydrogen bonds [97]. Dosing too much H2O2 could result in the decrement of the extracellular enzyme, protease and amylase that are easily prone to OH radicals attack. Protease will transform into amino acids through maillard reaction and carbohydrates will turn into simple sugars via caramelization when putting higher specific energy by microwave irradiation. Balancing the amount of dosage should take into consideration because we need to make more sludge soluble, dissociate more EPS to release catalytic substances and keep it in an active state [98–100]. In this method, the biogas production increased by 85.6% and the production of methane was 250 mg/g VS at the end of 30 days. Hydrolysis of sludge is thought to be a rate-limiting phase, while microwave irradiation promotes its solubilization and then increase its hydrolysis rate which means that more VFA would be generated and used by methanogens for methanogenesis. Addition of H2O2 stimulates the EPS via breakage of hydrogen bondings and sludge dissociation through the destruction of microbes’ cell wall.

4. Enhancement of acidification Acidification is a process of VFA generation which is the substrates for methanogens to methane yield. Soluble monomers are converted into short-chain fatty acid, such as acetic acid and propionic acid, which are the end products of this process by acidogenic fermentation bacteria. However, acetic acid is the direct substrate for methanogens utilization. There are two pathways of VFA production. First, simple organic wastes, such as monosaccharides and proteins, are taken up by 126

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acetogens to produce short-chain acid (acetic acid, propionic acid, butyrate acid, etc.). Secondly, a little part of acetogens, such as homo acetogenic bacteria, can serve carbon dioxide and hydrogen as substrates and then turn them into acetic acid. Therefore, either pathway is promoted, the rate of acidification will be raised.

4.1.2. Biotransformation of carbon dioxide and hydrogen to acetic acid It is well known that VFA with more than three carbon chains cannot be accumulated because they are not direct substrate for methanogens, so they will be degraded to acetic acid in the fermentative stage. Acetic acid is usually produced from acidification of mono substrates, while they can be also formed through hydrogen and carbon dioxide in the acidification stage by homoacetogens. If biotransformation of hydrogen and carbon dioxide in the acidification stage can be increased, more acetic acid will be produced, which provide more direct substrate for acetotrophic methanogens to produce more methane in the methanogenesis stage. Simultaneously, the percentage of acetic acid in the acidification step ought to enhance. The increment of the abundance of Syntrophomonas just like the bacteria mentioned above could lead to the accumulation of acetic acid which stimulates Methanosaeta growth for more methanogenesis. Salomoni et al. [35] divided anaerobic digestion in two phases with two reactors: one is devoted to hydrolysis, acidification, and acetogenesis (fermentation), while the other is methanogenesis and fermentation phase is injected with CO2 generated in the methanogenesis phase. It was reported that injection of CO2 allowed its adsorption and a remarkable VFA increase which indicated that a part of CO2 was reduced to short-chain fatty acid according to the Wood–Ljungdahl pathway [116–118]. CO2 plays a special role in acidogenesis, as it can react with reductant, such as hydrogen, to generate acetic acid and stimulate the production of shortchain fatty acid. With the activity of some enzymes increased, biotransformation of carbon dioxide and hydrogen to acetic acid in the acidification stage were boosted, providing a more direct substrate for methanogens in the methanogenesis phase. Mohanakrishna et al. [119] utilized a microbial electrosynthesis (MES) to transform carbon dioxide to acetic acid by a biocathode under a mild potential condition and metabolic functions of microorganism and electrical potential significantly influenced these whole process. Homoacetogenic bacteria are a kind of acid-forming bacteria, which can converse two molecules of carbon dioxide to acetic acid with the hydrogen as a reducing agent. Meanwhile, acetogens ubiquitously exist in various kinds of environment, such as alkaline, high-salt and hot environment. Thus, they are flexible to be applied in a hybrid system with biological functions.

4.1. Production of acetic acid 4.1.1. Bioconversion of carbohydrates into acetic acid Acetic acid is a key intermediate product that can affect the methanogenesis, therefore, enhancement of acetic acid generation draws a lot of attention. Complex organic matter can be hydrolyzed into various kind of mono substrates, and some of them are easy to be degraded, but some of them are not. Therefore, bioconversion of hardly degradable substrate to acetic acid and hydrogen that are important intermediate products plays a key role in enhancing methane generation. Monosaccharides are the main hydrolysis products in the sugar-enriched waste and there are two types of configuration (i.e., dextrorotatory and levorotatory). It was reported that not only D-monosaccharide but also L-monosaccharide is reported to be generated in the hydrolysis steps [49]. Although monosaccharides with D-configuration are easily digested into VFA, monosaccharides with L-configuration are hard to digest because they are inhibitors of bacterial growth [107]. Liu et al. [49] found that adding cysteine into anaerobic digestion of food waste facilitated the conversion of adverse configuration. It was verified that some enzymes, such as acetate kinase, CoA and L-glucose dehydrogenase, were increased by cysteine [49] and L-glucose dehydrogenase can catalyze L-glucose metabolized to pyruvic acid that is an important metabolic intermediate for acidification of L-glucose, which could enhance acetic acid production eventually [108]. It is widely known that the contact between enzymes and substrates is a prerequisite for achieving highly efficient biological reactions. What’s more, improving the acidification of refractory substrates to acetic acid is conductive to methanogens’ metabolism for methane yield. It was concluded that sCOD and acetic acid production in the fermentative step is the main driver of the overall methane production [35]. Acidification stage is concerned with the abundance of the relevant microbial community. Dong et al. [109] found that porphyromonadaceae family which generated acetic acid from carbohydrates or proteins were dominant in the system after thermal pre-treatment, and production of VFA was 348.63 mg COD/gVSS which was 6.8 times higher than the control. Zhang et al. [110] also found that the percentage of order Clostridiales in the microorganism which can degrade carbohydrate, sugar, amino acid, etc. and acetic acid and methane production were both raised. Desulfovibrio and Levilinea have the ability to transform organic matter or carbohydrates to acetic acid, CO2 and H2 through degradation of sulfate and propionic/butyric acid respectively [111]. A novel method was put forward by Luo et al. [112], which enhanced the acidification step by addition of polycyclic aromatic hydrocarbon. The activity of key enzymes relevant to acetic acid production and the quantities of their corresponding encoding genes were promoted and their microbial community structures shifted a lot. In the metabolic pathway, VFA is formed via transformation of acetyl-CoA to acetyl phosphate by PTA and then converted into acetic acid by AK. With the presence of more active enzymes, such as AK, more acetic acid will be generated. In addition, more genes associated with the enzyme of acetic acid formation regulate the synthesis and expression of AK and ATP enzymes which resulted in more acetic acid production. Other VFA, such as propionic and butyric acid, can be degraded to acetic acid with the generation of hydrogen which can be taken up together with CO2, yielding from the metabolism of acetotrophic methanogens, by hydrogenotrophic methanogens to produce methane [113–115]. In addition, it is reported that Magnetite also facilitates consumption of H2 by functional microbes, and propionic acid and/or butyrate acid oxidizing bacteria coupled with H2-utilizing bacteria produce more acetic acid for acetogens consumption and utilization [68].

4.2. Production of propionic acid Proteins and carbohydrates degradation plays an important role in VFA production. VFA can be converted into acetic acid by hydrogenconsuming bacteria and then turned into methane. Propionic acid is the most general short-chain fatty acid that ranks only second to acetic acid and easy to be transformed into acetic acid. Therefore, many researchers investigated the enhancement of propionic acid production in the anaerobic digestion of sludge, such as primary sludge and wasteactivated sludge, which contain a large portion of carbohydrates and proteins. In general, primary sludge, served as solid organic waste, contains a large amount of carbohydrates, while waste activated sludge (WAS) composed mainly of the bacterial cell contains a large portion of proteins. Due to the high temperature, fermentation under mesophilic and thermophilic is beneficial to facilitate the solubility of organic compounds and improve the biological and chemical reaction rates [43,120]. Zhang et al. [121] investigated the effect of pH variation on short-chain fatty acid production from waste activated sludge under mesophilic and thermophilic conditions and found maximum propionic acid was appeared under pH 9.0 and pH8.0 respectively which indicated that high temperature and alkaline pH condition facilitated propionic acid production. Under the uncontrolled pH, the dominant VFA was propionic acid. Conversion of hydrolysis products implies that more short-chain fatty acids are generated indicating a higher substrate utilization of acidogens. Yuan et al. [122] also found the same conclusion that propionic acid production from excess sludge was significantly improved and maintained stable under alkaline condition. Although increasing pH will result in more VFA production, it took 127

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longer time to generate the same amount of VFA which may be attributed to the toxic effects of high alkalinity to acidogens while low alkalinity will raise microbiological activity [123]. It is widely known that domestic sludge consists mainly of proteins, carbohydrates, lipids and fermentation of these substrates in the acidification steps is associated with VFA production. Alkaline condition improves the solubility of carbohydrates and proteins, especially proteins due to their high content in the sludge, and then enhances the conversion of them for more VFA productions. Feng et al. [124] found that the addition of carbohydrates and adjust pH can enhance VFA production, especially propionic acid, and pH variation can not only improve VFA production but also the percentage of individual VFA. What’s more, the key enzymes associated with propionic acid appeared the highest activities at pH 8.0 which indicated that high propionic acid content in the VFA. In view of a large amount of proteins remaining in the fermentation liquid after VFA generation which indicated the low C/N ratio of waste activated sludge, enhancement of proteins bioconversion is conductive to VFA yield. The carbon to nitrogen mass ratio of WAS is approximately 7.1/1, therefore, addition carbohydrates substrate increases the C/N ratio to a higher level from 20/1 to 30/1 which is more beneficial to the microorganism and VFA production. According to the effect of pH variation after supplementing of carbohydrates, OAATC, which is a key enzyme relevant to propionic acid synthesis, exhibits higher activity at pH 8.0. OAATC catalyzes pyruvate and methylmalonyl CoA to oxaloacetate and propionyl CoA respectively, which is associated with supply carbon flux from the central carbon metabolism to propionic acid. CoA transferase is also a key enzyme responsible for catalysis of the transformation of propionyl CoA to propionic acid [124]. Luo et al. [125] also conducted the investigation of carbohydrates addition in continuous-flow reactors, however, they found the effect of sludge retention time (SRT) and temperature on anaerobic digestion of waste activated sludge for propionic acid production. Increasing the SRT and temperature in a proper range was in favor of VFA production, especially propionic acid production and the activity of acid-forming enzymes were reported to be enhanced. It was reported that acidogens were more active at the SRT shorter than eight days. In the experimental results of Luo et al. [125], propionic acid was the dominant product when SRT was shorter than 8d and its concentration was 143.3 mg COD/g VSS, which was the highest production in their experiment. However, increasing the temperature to 55 °C will lead to a dramatical drop of VFA, which may attribute to VFA consumption by methanogens and high temperature that is more appropriate for methanogenic bacteria. At 37 °C, the activities of acid-forming enzymes reached the highest, especially enzymes responsible for propionic acid production. Jiang et al. [126] found that waste activated sludge digestion with a surfactant in it would result in more VFA production and the maximum VFA production appeared at sodium dodecylbenzene sulfonate (SDBS) dosage of 0.02 g/g and fermentation time of 6d. While high concentration of surfactant, such as 0.2 g/g, will cause inhibition to the activity of acidogens due to its toxic effects. Proteins and polysaccharides are the main constituents of WAS and degradation of them during the anaerobic digestion will generate VFA. The solubilization of sludge particulate organic-carbon is beneficial to VFA production. The presence of SDBS enhances the solubilization of proteins, polysaccharides, and EPS. EPS contains microbiologically produced polymers (polysaccharides and proteins), therefore, solubilization of EPS causes the break-up of sludge and releases these polymers into the aqueous phase. Acidification occurs accompanied by degradation of these polymers and more VFA is generated. With the dosage of SDBS, degradation of glucose and L-alanine was 1.22 and 1.44-fold respectively than the control and microbial activity was significantly raised. Li et al. [127] also found the effect of pH value on VFA production, while they conducted the investigation of anaerobic digestion of primary sludge. Primary sludge is a kind of sludge that generated from the primary settling tank in wastewater treatment plants, while WAS is

produced in the secondary sedimentation tank. Primary sludge contains a large amount of easily degradable fractions, thus, fermentation of primary sludge which is served as a substrate is feasible to generate more VFA. It was reported above that anaerobic digestion of WAS under alkaline condition facilitated the VFA production, especially propionic acid [124], and it can also enhance VFA production from primary sludge [127]. According to the sCOD in the fermented liquid, it raised as the pH value increased, which means more substrates became soluble under weak base condition. Soluble COD is derived from not only sludge fermentation (proteins and polysaccharides) but also VFA. The number of soluble proteins are higher than that of polysaccharides, which indicates proteins are the main content in the primary sludge and plays a key role in the VFA generation [122]. Simultaneously, degradation of proteins will form NH4+ which increases the pH value in the system and enhances VFA production. 5. Enhancement of methanogenesis 5.1. Enhancement of acetic acid conversion to methane Acetic acid is the direct substrate for methanogens to methanogenesis, thus, enhancement of acetic acid conversion can increase methanogenesis. Adjustment of appropriate conditions for anaerobic digestion is beneficial to maximum methane generation. Microbial electrolysis cell with iron-graphite electrode creates a preferable condition for microbes [40]. Served as an electron donor and a reductant, iron could decrease the oxidative-reductive potential (ORP) which is beneficial for the growth of microorganisms especially for those anaerobic bacteria and facultative anaerobic bacteria. The dosage of proper magnetite can also establish a more reductive anaerobic micro-environment which raises the abundance of methanogens, which in turn enhances methanogenesis and produce more alkalinity relating to degradation of VFA and increase effluent pH value. GAC/NZVI mediator in the EGSB reactor for tetracycline wastewater treatment was found to enhance methane production [16] and zero valent iron (ZVI) in it could lower the oxidation-reductive potential (ORP) of the system. Served as a representative of the oxidation-reduction reactions in the fermenters, ORP value is a useful parameter to monitor the fermentation process. For anaerobic microorganism, low concentration of oxygen and ORP play a key role in their growth, because there is no cytochrome with high potential, cytochrome oxidase which acts as driver to promote those biological chemical reactions proceeded in the high potential and -SH of enzymes necessary for their growth should be reduced completely, which can be work actively with enzymatic function. When the OPR around the microorganism decreases to satisfy their needs, active metabolism in the cell proceeds and reduzates are generated, creating a more suitable environment for the microorganism. Steps, such as the addition of magnetite and zero-valent iron which served as an electron donor and reductant, can also be adopted to reduce the ORP value and establish a more reductive anaerobic micro-environment which is conductive to the growth of microorganisms especially for those anaerobic bacteria and facultative anaerobic bacteria. It also alters the community structure and the abundance of dominant bacteria in the reactors which improve the performance of the system and methanogenesis. Supplement of additives, such as mineral nutrients, metal oxide nanoparticles, bioaugmentation, and enzyme, stimulates methane production as well as improves process stability. Featured with desirable characters, biochar with porous structure can absorb ammonia to lessen its inhibition and immobilize methanogens [128–130]. Although it is widely known that ammonia can lead to inhibition of methanogens, the mechanism of this process is controversial. A few studies of AD in the pure culture indicated that there are two patterns. First, ammonia inhibited the activity of methane synthetase; Second, owing to its hydrophobicity, free ammonia getting into the microbial cell by means of passive diffusion caused imbalance of proton and deficiency of 128

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potassium which proved to evaluate the pH of the digester’s environment to facilitate CO2 sequestration with increased buffering capacity [131]. Biochar supplementation resulted in the decrease of TAN through sorption and precipitation and the increase of alkalinity in continuous anaerobic of sludge [132]. The cation exchange capacity of the functional groups via hydrogen bondings decided the sorption capacity [133,134]. Corn stover biochar and pine biochar with high calcium, magnesium and iron can not only increase the alkalinity of the digester but also provide essential elements for some methanogens to grow [131]. In the presence of Fe, regulating the redox conditions, hydrolysis and acidogenesis reaction rates can be accelerated to stimulate the degradation of volatile fatty acid (VFA) which means more substrates were fermented and more predecessor for methanogenesis were converted into methane [135]. A study found [136] that biochar addition significantly increased the concentration of Co and Ni that are the two essential cofactor key enzymes involved in methanogenesis and ionic state of them in the solution which can be taken up by microorganisms improves substrate utilization, methane production [137,138] and promoted microbial diversity. Ni, Co, and Fe are important cofactors of enzymes involved in the acetoclastic pathway [139] and are also reported to promote the methanogenesis because methanogenic archaea need to utilize hydrogenases containing Fe and Ni as cofactors [140]. The relative content of a group of slowly-growing bacteria, order/class anaerolineae, served as a degrader of recalcitrant compounds for the release of soluble organic carbon which can be taken up by another microorganism for growth [141] were in a rising trend. With the particular porous structures, biochar induced the formation of biofilm on the surface and inside the pore channel and immobilized microorganism which is beneficial to their growth, activity and resistance [129]. It could be included that dosing biochar selectively promoted the enhancement of the concentration of some acetate-utilizing bacteria over others. Shen et al. [142] also amended anaerobic digestion of sludge with corn stover biochar and put forward that biochar with large surface area and highly porous nature can absorb CO2 and high monovalent and divalent cation concentration in biochar accelerated carbonation converting CO2 into carbonate/bicarbonate and increasing the pH which was beneficial to the anaerobic digestion, especially for the growth and activity of methanogens for methanogenesis [143,144]. Nutrients are necessary for microbial growth. Except for nitrogen sources or phosphate sources, the deficiency of inorganic nutrients inhibits the metabolism of methanogens, which is more serious in the anaerobic biological treatment than in the aerobic treatment. Brulé et al. [145] found that a decrease of methane production and high content of VFA in the system were due to the deficiency of trace nutrients. The concentration of trace nutrients increase in the appropriate range is corresponding with the growth rate of methanogens. Although trace metals make effects on many aspects of bacterial metabolism, archaea bacteria involved in the final steps of AD, methane production, have a higher requirement of some trace element, such as Fe, Cu, Ni, Co etc. than fermentative bacteria which converts complex organic matter into VFA. Trace nutrients, such as iron, stimulated the growth of hydrogenotrophic methanogens by consuming hydrogen which was an adverse intermediate for AD in thermodynamics to generate methane [146], and high calcium and magnesium can not only increase the alkalinity of the digester but also provide essential elements for some methanogens to grow [131]. In addition, supplement of iron not only provides nutrients for methanogens growth but also promotes the availability of other metals by binding amino. The character of the substrate decides its alkalinity. Thus, adjustment of pH makes it more appropriate for methanogens to produce methane in the methanogenesis stage. It was reported that alkalinity pre-treatment enhanced the methanogenesis from anaerobic digestion of waste activated sludge [38] which indicates that pH value affects the anaerobic digestion. Due to the organic matter conversion into volatile fatty acid, pH value in the system will decrease which inhibits the activity of methanogens and lowers the transformation of VFA into

methane. Dai et al. [147] investigated the effect of pH on anaerobic codigestion of waste activated sludge and perennial ryegrass and found that the activity of acetotrophic methanogens, such as Methanosarcinaceae and Methanosaetaceae, was raised and observed methane enhancement via consumption of acetic acid, formic acid and hydrogen after pH increase. Methanobacterium which utilized carbon dioxide and hydrogen to generate methane was inhibited in a rarely low pH. It was also reported by Zhang et al. [110] that sludge which pretreated with pH 10 showed a significantly higher percentage of active Archaea than control. Although the alkaline condition is in favor of the active of methanogens, the active microorganisms will be killed or inhibited in the extreme alkali environment, because they are too toxic for their growth. Shen et al. also found that absorption of CO2 which decreased the pH in the reactor and acceleration of carbonation converting CO2 into carbonate/bicarbonate which led to the increment of pH in the system was beneficial to the anaerobic digestion, especially for the growth and activity of methanogens for methanogenesis. What’s more, VFA generation results in a decrease of pH and increase the degradation of VFA by methanogens will lead to more alkalinity and effluent pH value. Some substrate with high alkalinity can sustain pH in the desired range and stable operation of the reactor. Regulation of experimental conditions by shortening HRT with increasing OLR imposed pressure on the shift of microbial community which could wash out some of slow-growing methanogens [148]. Simultaneously, raising the temperature of AD could reach a faster reaction rate, produce more biogas as well as reduce pathogen content and occurrence of foaming [149]. During the methanogenesis, complex substances are decomposed through the interaction of bacteria and archaea and variable operation parameters cause instability of methane yield, thus, minimizing changes in microbe dynamics under the situation of the high load could maintain highly efficient operation of the system [150]. During treatment of high concentration of organic wastewater, high organic loading rate affects the AD system and degrades its performance, because organic loading rate increases with an increment of substrate concentration and decrement of hydraulic retention time. It also significantly disturbs the microbial community via a rapid change of intermediate abundance, and excess accumulation of intermediate has a negative influence on methanogens, which decrease the methane production. Well-developed sludge granulation is a prerequisite factor for the high performance of UASB reactor [151], while a high load of wastewater will wash out these granules [152,153]. It also caused VFA production and accumulation, acidification of the system and inhibition of the process. Therefore, proper organic loading rate should be applied to achieve high organic removal rate and stable methane production. Balancing the C/N ratio in feedstocks is significant for stable operation of anaerobic digestion. When the C/N ratio is high in the feedstocks, nitrogen will be consumed rapidly by methanogens to meet their protein requirements, which results in low methane production. Nitrogen will exist and accumulate in the form of ammonia with the low C/N ratio in the feedstocks, which inhibits the methanogens metabolism due to its toxicity. Sensai et al. [154] adopted anaerobic codigestion of distillers grains and swine manure to adjust C/N ratio and illustrated that maximum methanogenesis was achieved with the C/N ratio of 30/1. There is a wide range for sustainable anaerobic process, lower C/N ratio of feedstocks in this range improves the activity of methanogen in a long time, which contributes to a high methane production rate [155]. Wang et al. [155] found that NH4+ accumulation in the low C/N ratio provided an appropriate condition for methanogen to generate methane in the methanogenesis stage, because VFA produced by acidogens and NH4+ constituted a buffering system for methanogen metabolism, and activity of methanogen was reported to be strongly enhanced by the feedstocks with high nitrogen content, which resulted in more VFA converting into methane. Microorganism in anaerobic digestion contains bacteria and archaea, and one is responsible for fermentation, the other is for methanogenesis at the final stage through 129

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intermediate degradation [156]. Due to the limitation of the substrate, methanogenic archaea are strictly dependent on syntrophic partner bacteria [150]. Hence, the addition of nutrients to provide an appropriate condition for the syntrophic association between bacteria and archaea is beneficial to AD and methanogenesis. Carbohydrates are necessary for methanogens growth and compensating the deficiency of carbon source can promote microbial metabolism. Carbon dioxide is an end product of acidification and methanogenesis stage, and conversion of CO2 into carbohydrate can not only provide energy for methanogens but also contribute to stable performance. Neshat et al. [55] found that purple photosynthetic bacteria were one of the special photoautotrophic organisms that were able to utilize CO2 in the presence of light to produce carbohydrate through anoxygenic photosynthetic, which provided energy for the organisms in the dark cycles [157]. Supplement of this kind of bacteria is conductive to balancing C/N ratio. Carbohydrate generated in this unique pathway compensates the shortage in feedstock with low C/N ratio and provides carbon source in the carbon deficiency. If C/N ratio is low, which means carbon source is insufficient for the microorganism to metabolism, the concentration of nitrogen source is relatively high which might lead to the high content of ammonia nitrogen and then inhibits the anaerobic digestion process. The increase of C/N ratio means CO2 fixation which enhances the content of carbon and the activity of methanogens. Hydrogen sulfide is a product in the anaerobic digestion, which can easily diffuse through the cell membrane into the cytoplasm and

damage methanogens, especially acetotrophic methanogens [131]. Sulfate is reduced to hydrogen sulfide by sulfate-reducing bacteria, and competition will be occurred in the anaerobic fermentation between sulfate-reducing bacteria and methanogen due to the utilization of substrates, such as hydrogen and acetic acid. Therefore, elimination of hydrogen sulfate is necessary for more methane yield. Dai et al. [158] pretreated sludge at pH 10 for eight days and adjusted the system at neutral pH for methane generation, which can simultaneously reduce hydrogen sulfate and enhance methane generation. Alkaline pretreatment of substrate limits the sulfate-reducing bacteria metabolism and hydrogen sulfate production. Thus, the relative abundance of Bacteroidetes and Firmicutes was raised, which resulted in increasing the concentration of acetic acid and hydrogen. With the high level of these two substrates, the activity of methanogens will be raised. Except for inhibitor generated in the anaerobic digestion, inhibitor in the substrate goes against the growth of methanogens. For example, by reason of strong bacteriostatic, antibiotics and its intermediates in tetracycline wastewater disturb activity of the anaerobic microbial community, which decrease the stability of anaerobic biological treatment and then inhibit methane production [159]. Zhang et al. [16] applied GAC into the anaerobic digestion of tetracycline wastewater to decrease the inhibition of tetracycline on methanogens and enhance methanogenesis. Therefore, high-efficiency degradation of toxic substances could destroy its antibacterial activity and elevate the microorganism activity. After more toxic compounds have been degraded, the abundance of some

Fig. 3. The mechanism of electron transfer (a: Direct interspecies electron transfer (DIET); b: DIET induced by conductive material). 130

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microbes increases, such as Methanosaeta and Syntrophus. Methanosaeta is a kind of acetotrophic methanogens and strict anaerobe. Syntrophus can utilize butyric acid to generate acetic acid and hydrogen, contributing to the accumulation of acetic acid, which can be taken up by Methanosaeta and is beneficial for their growth.

process. In this anaerobic reactor with MEC, methane was theoretically produced from two pathways. First, VFA and hydrogen were consumed by acidotrophic methanogens and hydrogenotrophic methanogens respectively via anaerobic digestion of sludge. Secondly, CO2 accepted electron released from Fe0 and organics to produce methane via the biocathode reactions [165]. With regard to one of the pathways for methane production, increment of applied voltage in a proper range promoted electron transfer in the MEC which in turn led to speed up the cathodic reaction rate that methane production became more quickly. As for the pathway, consuming H2 which is an adverse product inhibiting the degradation of organic acid by hydrogenotrophic methanogens is beneficial to the anaerobic digestion and promotes their growth which stimulates the methane production. Redox reaction in the MEC reactor provided a suitable environment for methanogens. Synergistic effects of high-solid anaerobic digestion and MEC enhanced the reaction between CO2 and H2 with methanogens as catalyze. With more intermediates being consumed and remaining stable, entire condition is suitable for archaea to methanogenesis, thus hydrogenotrophic methanogens reduce more CO2 to CH4. Interspecies electron transfer was enhanced to induce redox reaction that more end products were generated. Upon the study of Wang et al. [65], magnetite (Fe3O4) was added in the anaerobic sequential batch reactor as a typical conductive material with the attempt to enhance removal of contaminants and production of methane during Fischer-Tropsch wastewater treatment. Rich in oxygenated by-products (monohydric alcohols, monocarboxylic organic acid, and ketones), hydrocarbons and aldehydes, the residual wastewater still had a large amount of alcohols and organic acid that contributed to high COD content and low pH value after recycling by rectifying column [166]. Although AD has been believed to be a technically feasible and cost-effective strategy for high-strength organic waste, slowly syntrophic metabolism of propionic acid and butyric acid, which is commonly described as interspecies H2 transfer, will limit this process [166–169]. Not required the catalysis of multiple enzymes to produce hydrogen to function as an electron shuttle, direct interspecies electron transfer is a more effective mechanism than interspecies hydrogen transfer. It is also reported that supplement of electrically conductive materials, such as magnetite, biochar, and carbon cloth, enhance organics (e.g. ethanal, acetic acid, propionic acid and butyric acid) degradation and methane production, and promotion of DIET is the main reason for these results [170–174]. Enriched functional microbes induced by proper dosage of magnetite could facilitate reduction of CO2 to CH4 and consume hydrogen produced by acidification to stimulate contaminants removal.

5.2. Enhancement of carbon dioxide conversion to methane With the enhancement of acetic acid production, more methane will be generated by acetotrophic methanogens if we balance the abundance between acidogens and methanogens. If the consumption rate of VFA is lower than that of production, VFA will be accumulated and the toxicity of intermediate will inhibit the activity of methanogens. However, there is another pathway of methane formation. CO2 can be reduced to CH4 via utilization by hydrogenotrophic methanogens. Therefore, if the rate of this pathway is raised, more methane is bound to yield. Viggi et al. [160] discovered that the positive effect of biochar is directly related to the electron-donating capacity of the materials, but was independent of its bulk electrical conductivity and specific surface area which played a major role in the biochar-mediated interspecies electron transfer in methanogenic consortia. High conductivity of biochar particles leads to the positive stimulatory effect on methanogenesis which accelerated the process of interspecies electron transfer between syntrophic bacteria resulting in more metabolism to enhance methane production. Enriched functional microbes induced by proper dosage of magnetite could facilitate reduction of CO2 to CH4 [65]. Zhang et al. [16] applied granular activated carbon and nano-scale zero valent iron into anaerobic digestion of tetracycline wastewater and achieved more methane production. Granular activated carbon (GAC) is reported to strengthen ethanol transfer to methane via DIET between special species [161] and EPS secreted by microorganisms boosts interspecies mass and electron transfer [16]. The related mechanism is shown in Fig. 3. H2 produced from oxidation of iron in the system serves as an electron donor for hydrogenotrophic methanogens to methanogenesis. Served as an electron channel, GAC and NZVI adjusted electrical syntrophy between methanogens and hydrolytic acidification bacteria [162]. Simultaneously, function microbes like dissimilatory iron reduction bacteria (DIRB) that could effectively transfer electron to methanogens would be enriched with the proper magnetite addition, and then methane production increased with the reduction of Fe(III) to Fe(II) [163]. DIET between methanogens and iron-reducing bacteria proceeded with adding GAC/NZVI in the reactor [161]. CO2 accepted electron from Geobacter to be reduced to methane, in addition, NZVI and H2 could also act as an electron donor to participate in the reactions above [164]. Applied voltage can accelerate electron transfer in the system, and therefore the application of microbial electrolysis cell (MEC) boosts electron transfer in anaerobic digestion. Combination of high-solid anaerobic digestion and microbial electrolysis cell with iron-graphite electrode has been observed to enhance the production of methane from waste activated sludge’s degradation [40]. Electrode accepted electron from anode which was generated by exoelectrogen and then transferred it to cathode. The reaction at the cathode with the reduction of CO2 was catalyzed by electrochemically active microorganisms (H2utilizing methanogens). As a trace nutrient, iron stimulates the growth of hydrogenotrophic methanogens by consuming hydrogen which is an adverse intermediate for AD in thermodynamics to generate methane [146]. However, pH in the digester would increase when applying voltage was too high (0.6 V), because a large amount of hydrogen ion to carry on a cathodic reduction for the production of methane and hydrogen. Excess alkalinity decreased the activity of methanogens and H2 served as a substrate of hydrogenotrophic methanogens availed to their growth, but the adverse effects of pH was dominant [40]. If applied MEC in the AD to enhance methane production, pH in the digester would increase because a large amount of hydrogen ion to carry on a cathodic reduction for the production of methane and hydrogen. Maintaining the pH value in an optimum range is in favor of AD

5.3. Techno-economic assessment Although there are large amounts of strategies in lab-scale showing high potential in the anaerobic digestion since they achieve the enhancement of methane production, the economic assessment is limited in the literature. Most methods lead to high methane yield, but they are not economically feasible because the cost they consumed is higher than that of the increased methane. Thus, comparison of the input and output energy in different methods helps us to estimate their economic benefits. With the assessment of economic investment, we can check the real feasibility of these technologies when approaches are implemented in the anaerobic digester from wastewater plant in full-scale. In the following, a simple and useful tool for process feasibility assessment in terms of economic efficiency is shown in order to evaluate the potential and sustainability of different methods. For this, we introduce a ratio to compare the cost of strategies input relative to the cost produced by extra methane (equation (6)) and treatment is economically feasible when ratiocost is less than 1. According to the U.S. Energy information administration (EIA), the commercial price of natural gas is 7.88 US dollars per thousand cubic feet which is equivalent to 0.000278 US dollars per liter. It is well known that 98% of the ingredient in natural 131

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gas is methane, therefore, we can suppose that the cost of methane is approximately equal to that of natural gas.

Ratiocost =

Cost of strategies input C = Price of extra methane P

consumption that heats sludge from the room temperature to the specified temperature. Secondly, the quantity of heat that maintain the substrate at the specified temperature. Thirdly, the cost of coal that is utilized to burn in order to produce the equivalent heat is input cost. According to the research of Cheng et al. [176], the specific heat capacity and the thermal conductivity of sludge is relevant to its TS concentration. The relation of specific heat capacity and TS is shown in Eq. (7). In addition, when the TS concentration of sludge is lower than 15%, the thermal conductivity of sludge is 0.6 W/(m K). Total heat consumption is calculated by Eq. (8). Sludge moisture content is 90% and its density is 1.07 g/ml. In the research of Choi et al.[91], a batch test was performed using serum bottle with a working volume of 200 mL in a 500 mL serum bottle. The depth of 500 mL serum bottle is 180 mm (Alibaba Enterpriser, https://www.1688.com), therefore, the height of heat conduction is 72 mm. The highest methane production is 273.2 mL/g COD under the reaction temperature of 150 °C at the reaction time of 60 min while that of control is 194.5 mL/g COD. We assume that the room temperature is 20 °C, thus, total heat consumption is 50 kJ. It is well known that the fuel value of coal is 23000 kJ/kg coal and the average sales price of coal in the U.S. is 0.037 U.S. dollars/ kg coal. In order to generate 50 kJ heat for this process, 0.00217 kg coal is needed that is worth 0.00008 U.S. dollars. With the substrate COD of 169 g/L, extra creative value by methane is 0.00074 U.S. dollars which come to that ratiocost is 0.108.

(6)

Due to the low degradability of some recalcitrant organic matters in the substrate, pretreatments are applied in order to improve their solubilization and speed up the rate of hydrolysis. As for electrical treatment, the cost of strategies input is electricity consumption. The commercial price of electricity in the U.S. this year which is shown in the EIA is 11.01 cents per kilo watthour that is amount to 0.1101 U.S. dollars per kilo watthour. Safavi et al. [52] applied the electrical intensity at 50 kWh/m3 for 4L pig slurry in anerobic digestion to realize the 1.18 m3 extra CH4 production after treated by a pulsed electric field. Therefore, the cost of strategies input is electrical intensity multiplies treated substrate volume and the price of electricity, which is equal to 0.02202 U.S. dollars and the price of extra methane is 0.32804 U.S. dollars. The ratiocost is 0.067 which means this pre-treatment is economically feasible. 31.4L/kg VSS extra methane production after electrical treatment with the energy consumption of 4.9 kJ that is equivalent to 0.0013kWh was achieved by Feng et al. [40]. The VSS of sludge mixture is 62.7 g/Lsubstrate and the working volume of the reactor is 2L, therefore, the P is 0.0011U.S. dollars and the C is 0.00014U.S. dollars. The ratiocost is 0.127. Radiant pretreatment feeds in energy and then output energy in the form of methane which simplifies the process of comparison. Zhao et al. [175] studied optimization of microwave pretreatment of lignocellulosic waste for enhancing methane production. A maximum methane yield of 221 mL/g substrate compared to 170 mL/g substrate in control was obtained under the optimum pretreatment where substrate concentration is 20.1 g/L and pretreatment time is 14.6 min. The microwave power was maintained at 500 W for treatment and the process was carried out in serum bottles with a working volume of 150 mL. Therefore, input energy is 0.122 kW·h and extra output methane volume is 153.765 mL. What’s more, the input cost is 0.0134 U.S. dollars and output cost is 0.00004 U.S. dollars and the ratiocost is 313.5 which means microwave pretreatment in this research is not economically feasible. The result is in accordance with that of Eswari et al. [37] which conducted the economic assessment of their research that utilization microwave or combined with H2O2 pretreated substrate in AD. However, there is still some microwave treatment in the full-scale plant, which may attribute to the decrease of sludge disposal cost and solids disposal cost due to the reduced quantity of sludge solids under the microwave treatment. Taking all cost reduced into account leads to a feasible strategy. Compared to the economic assessment of electrical treatment, that of thermal hydrolysis is more complicated. Firstly, the energy

C (kJ/(kg· K )) = 1.0 + 0.005TS(%)

(7)

Q = cm T + ·d· T ·s

(8)

In terms of biological method, inoculation and bioaugmentation are common strategies. For example, lipase was brought in anaerobic digestion of waste in swine slaughterhouse [54] and this strategy input is the cost of lipase. The price of lipase isolated from pig pancreas is 193.3 U.S. dollars per 500 g where the cost is indexed with the exchange rate from dollar to Chinese Yuan (CNY) of 1:6.96 (Sigma-Aldrich, Shanghai, China). When the lipase concentration is at 0.2% w/v, 132.5 mL CH4/g VS extra methane yield is achieved. The organic loading is 20 g VS/L of the substrate and the working volume is 250 mL. We can calculate that the cost of input is 0.00019 U.S. dollars and the price of output is 0.00018 U.S. dollars. The ratiocost is 1.06 which indicates the input is similar to the output but it is a little higher than the value that methane produces. That is to say, this strategy is an economic balance, therefore, we don’t need to spend our time doing useless work. It may contribute to the high price of lipase isolated from pig pancreas which is almost nine times higher than the market price of food grade lipase (21.5 U.S. dollars per 500 g, Alibaba Enterpriser, https://www.1688.com). However, lipase from pig pancreas is more efficient than food grade lipase due to its high purity and dependency. Thus, the balance between price and efficiency is the matter we should weigh, then we can find a

Table 2 Cost consumption by different treatment and price produced by extra methane. Treatment Electrical Electrical Electrical Microwave Microwave with H2O2 Microwave with CaO2 Microwave with NaoH Thermal Biological Biological Activated carbon Cysteine magnetite ferroferric oxide NaOH

Energy consumption (kWh) 0.2 0.0013 1.12 0.122 68.8 0.016 0.167 0.014 – – – – – – –

Extra methane production (mL) 6

1.18 × 10 3940 5.9 × 104 153.765 2.5 × 105 61,288 10,836 2660 662.5 194.1 1.26 × 107 8953.2 7460 78.3 682

132

Price of additives ($/g)

Input cost ($)

Output cost ($)

Ratiocost

Reference

– – – – 0.012 0.043 0.0036 – 0.043 – 0.015 0.243 0.025 0.025 0.0036

0.022 0.00014 0.1232 0.01340 10.968 16.68 0.0314 0.00008 0.00019 0 155.4 0.055 0.0099 0.5 0.00089

0.3280 0.00110 0.0164 0.00004 0.0695 0.017 0.0030 0.00074 0.00018 0.00005 3.51 0.037 0.0021 0.00002 0.00019

0.067 0.127 7.512 313.5 157.8 979.0 10.47 0.108 1.06 0 44.27 1.486 4.714 22,970 4.684

[52] [40] [41] [175] [37] [38] [39] [91] [54] [60] [48] [49] [65] [68] [3]

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moderate lipase to consume relative low cost and create relative high value. As for those inoculation with waste such as pulp and paper mill sludge to enhance anaerobic digestion, input cost is zero, because we don’t need to spend our money on the waste and treat waste simultaneously [60]. The extra value we create is the cost of the increased methane production. In the previous comparison, some pretreatments which almost act on the stage of hydrolysis are demonstrated. Other strategies are mostly the addition of different substances, such as a chemical reagent, biological additives, greenery biomass and so on, in the process of anaerobic digestion, and what we need to calculate the price of these substances to compared with the extra value produced by methane. What’s more, some researches adjust pH to realize more methane generation through the addition of alkali reagent, which can be calculated by the cost of substance input. More economic comparisons of different strategies are concluded in Table 2. It is obvious that most of the methods are not economically feasible which is similar to the result of Cano et al. [177] because all these methods are just applied in the lab scale and assessment are calculated according to the process of research in the laboratory. In the full-scale operation, taking sludge treatment as an example, the volume of sludge decreased a lot in the anaerobic digestion as well as the solid in the sludge which spends a large amount of money treating to reach harmlessness. Therefore, the sum of reduced cost in waste treatment and increased value produced by methane is the output cost. Some researches conduct an economic assessment in their article which is different from mine. They compare the cost of total energy input and the price of total methane. With this approach, we can’t judge whether the price spent in enhancing methane generation is higher than the methods’ extra expense that gives us the decision to take steps. What’s more, some researches take heat value of methane and cost of these energy produced by methane as evaluation criteria. As we know, the combustion of methane needs extra cost. In addition, the heat value of methane is 39.82 MJ/m3 and the value of it is 1.2166$/ m3, but the commercial price of methane is 0.278$/m3 which is of great difference. Among the treatment compared in this review, most of the electrical treatment is economically feasible as well as a kind of thermal pretreatment which all make actions on the stage of hydrolysis. There are also some biological treatments by supplement of fungi, bacteria or archaea, however, it is complex to evaluate its input cost, because, in the lab-scale research, we usually purchase a tube of microorganism in the form of a dry powder which can be made into microorganism in glycerin as much as you want. Therefore, we can hardly calculate the price of bacteria utilized in the research. Biological treatments usually make actions on the stage of hydrolysis and acidification. For the stage of acidification and methanogenesis, the strategies for methane enhancement almost adjust pH or supplement of some additives which are a convenience to operate that decrease the manpower cost. Also, the enhancement of carbon dioxide conversion into acetic acids or methane is always through altering the configuration of reactors that is not compared in this review. The most economically feasible treatment is inoculation of different wastes, such as sludge, manure, which are needed to treated to decrease their harmless. Through the approach, we not only decrease more solid waste but also increase methane yield and our net cost is the price of extra methane generation.

decomposed into sugar, proteins, lipids, etc. and then are hydrolyzed into micromolecules, such as monosaccharides, amino acid and so on, which are easier to be taken up by fermentative bacteria to form volatile fatty acid. Due to the special characters of sludge, dissociation of sludge results in cell destruction accompanied by extracellular polymeric substances decomposed, releasing hydrolytic enzymes into the system, which promotes the process of hydrolysis. Acidification is a process that transforms micromolecules into VFA. There are various substrates which can be converted into VFA, while some of them have a low productivity. Conversion of the adverse configuration of monosaccharides facilitates the VFA generation and decrease the inhibition of microorganisms. In the metabolic pathway, VFA is formed via transformation of acetyl-CoA and butyryl-CoA to acetyl phosphate and butyryl phosphate by PTA and PTB, and then convert into acetic and butyric acid by AK and BK, respectively. Therefore, the increase of some relevant enzymes, such as acetyl-CoA, butyryl-CoA, PTA, PTB, AK, and BK are beneficial to the generation of VFA. In addition, acetic acid can be formed with consuming hydrogen and carbon dioxide by homoacetogens via the Wood−Ljungdahl pathway. Thus, enhancement of acetic acid which are the direct substrate for methanogens production via this pathway promotes methanogenesis. For example, CO2 generation in the methanogenesis stage can be injected into the fermentative stage, inducing more acetic acid yield in this stage. After enhancement of the first three stages, methanogenesis stage is a key process for more methane production. Methanogenesis stage is the process of electron transfer. Thus, the increment of electron transfer contributes to more methane production. In the previous study, served as primary shuttle compound, H2 can transfer an electron from organic acid oxidizers to hydrogenotrophic methanogens [168]. While recent studies revealed that direct interspecies electron transfer (DIET) is more effective mechanism because it doesn’t need to produce hydrogen to function as an electron shuttle with multiple enzymatic steps and save energy for microorganism to metabolize, and outer membrane electrically pili and c-type cytochromes are utilized as a carrier for electron transfer in DIET. Thus, acceleration of DIET is bound to promote methane production. Although it’s presumably universal for each individual microorganism, it’s even of greater significance for the syntrophic organism, such as methanogenic syntrophic communities. Conductive material or application of MEC will accelerate DIET and promote methane production via consumption of acetic acid by acetotrophic methanogens and reaction between hydrogen and carbon dioxide with catalysis of hydrogenotrophic methanogens. The activity of methanogens is a key factor for methanogenesis. As for strictly anaerobic microbes, a variation of the environment is adverse for their metabolism and growth. PH values seriously affect methane production and methane content in the biogas. In the start-up process, pH will decrease accompanied by VFA production, which inhibits the growth of methanogens. Neutral pH is beneficial for methanogens growth and alkaline condition promotes hydrolysis stage. Therefore, separation of the fermentative stage and methanogenesis stage can adjust the pH of these two stages respectively to make it more appropriate for different microorganisms. The balance of C/N ratio provides enough carbon sources and nitrogen sources for methanogens metabolism. Deficiency of carbon sources inhibits the methanogens growth and ammonia generation due to reductant nitrogen will hinder the anaerobic digestion via decrease of the activity of methanogens. Deficiency of carbon sources will lead to quickly consumption of nitrogen to meet their protein requirement. Trace element, such as Fe, Ni, and Co are essential for methanogens, because they are significant constitutes of coenzyme like CoA which participate in the process of methanogens. Simultaneously, they provide nutrients for methanogens growth and zero-valent iron can lower the environment oxidative-reductive potential creating a better environment for the microorganism. Decrease of inhibitors generation accompanied with anaerobic digestion and inherent inhibitors in the substrate are in favor of microorganism. Carbon dioxide generated in the AD decreases the pH value

6. Conclusion and perspective Various factors influence methane production in view of the entire process and each stage. However, regardless of any method you employ, the mechanism of enhancing methane production is according to the following portions. For the whole anaerobic digestion, hydrolysis, acidification, acetogenesis, and methanogenesis are four steps of it, and enhancement of any steps will lead to more methane generation. Hydrolysis is a rate-limiting process for anaerobic digestion and various methods are put forward to improve it, such as physical, chemical, biological or combination of any of them. Complex organic matters are 133

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in the system, therefore, adsorption of it or consumption of it for acetic acid and methane production facilitate the methanogenesis. Some complex organic matters are toxicity for microorganism, degradation of them contributes to more VFA generation and decrease the toxic influence on the microorganism. Methods for methane enhancement in the recent studies are almost in a lab-scale system, but particular application aims at full-scale anaerobic digestion which is more complicate to the operation and regulates. Development of reactor with special hydraulic flow pattern in full scale which meets the ecological requirement of the different anaerobic microbial population is a future research direction. Anaerobic bacteria have low metabolic rate compared with aerobic bacteria, thus the efficiency of the anaerobic biological treatment is relatively low. Raising the anaerobic bacterial abundance promotes its capacity and efficiency of treatment. Inoculation is a preferable choice, and wastes are degraded with addition into digestor. Besides, creating a more suitable condition for the different microorganism is in favor of their physiological stabilization. Although there is some synergetic effect between acidogens and methanogens, their ecological state and metabolic rate is of great difference. Therefore, instability of the ecological system will occur when acidogens and methanogens present in the same reactor. Separation of two kinds of bacteria in two reactors, which perform their own functions, promotes the degradation rate and stabilization of anaerobic biological treatment. It was also found in our research group that toxic compound, polycyclic aromatic hydrocarbon, can enhance the methane production and degradation of PAH in the codigestion of food waste and activated sludge which not only reduce toxic substances but also increase the energy production. It is a considerable direction for future investigation for methane enhancement. However, its mechanism is hard to explain, which need more deep investigations. Not all the strategies for methane enhancement is economically feasible. In general, the cost of energy they produced in the process doesn’t satisfy the cost they consume. Electrical pretreatment consumes less energy in the lab-scale research which means less input cost in the anaerobic digestion. However, thermal and radiant treatment cost too much energy to raise the temperature of the substrate and destroy its structure for increasing hydrolysis rate. Supplement of additives is in favor of the acidification and methanogenesis stage, but chemical reagent is expensive. In order to raise conversion of carbon dioxide to acetic acids and methane, integration of the process that recovers heat energy from methane engine can satisfy the energy consumption so that strategies are more economically feasible because methane of the same volume creates more cost in the form of energy than in the form of merchandise. Also, inoculation of waste can create net profit and decrease the waste solid treatment cost. Although methods in lad-scale are not economically feasible, integration and application in the full-scale will produce more value, it needs to take into consideration that strategies should be more energy self-sufficient and economically feasible.

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Acknowledgment

[27]

This work was supported by the National Science Foundation of China (51425802 and 51778454).

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