Anaerobic Mixed Consortia Research Articles

Optimization of Long-Chain Fatty Acid Synthesis from CO 2 Using Response Surface Methodology

A microbial electrosynthesis cell (MES) is an electrochemical technique in which electrolithoautotrophic electroactive microbes fix carbon dioxide (CO2) to longer-chain volatile fatty acids (VFAs). The synthesis of VFAs from CO2 requires optimization to improve the MES performance for industrial feasibility. This work studied the effect of different parameters, such as pH, headspace gas pressure, ethanol concentration, electrolyte, and trace element concentrations, on VFA synthesis from CO2 that used mixed anaerobic consortia in serum bottles. The operational parameters were varied according to a central composite design (CCD) and response surface methodology (RSM). A global optimum for the response variables was determined. The optimum values of different operating factors to maximize VFA production that was obtained from the optimizations were 1.12 × 105 Pa pressure, pH 7.149, ethanol = 2,318.7 mg/L, three times the standard Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ) 300 electrolyte concentration and five times the standard DSMZ 300 trace elements concentration. The experimental study was validated in triplicate, which considered the optimal values. The rate of total VFA production that was obtained from the operational parameters experimental study was 43.7 ± 5.9 mg/L/day under this optimized condition, which agreed with the predicted value of 30.39 mg/L/day. For the media optimization, validation of the experimental study was conducted at the optimal values, and their experimental response was 79.75 ± 19.26 mg/L/day, and the predicted response was 75.89 mg/L/day for total VFA production rates.

Assessing the syngas biomethanation in anaerobic sludge digestion under different syngas loading rates and homogenisation

Gasification offers digestate conversion into syngas, which can be metabolised to CH4 by a mixed anaerobic consortium. However, low gas-liquid mass transfer and CO presence can limit further scale-up. The present study assessed syngas biomethanation (55% H2, 45% CO) in a thermophilic lab-scale continuous anaerobic digester processing sewage sludge. Different syngas loading rates and mechanical stirring compared to pneumatic mixing were investigated. Results showed that anaerobic culture acclimation for CO utilisation is unnecessary. Moreover, the mechanical stirring replacement for the pneumatic system enhanced the gas-liquid mass transfer rate. Hence, the biological conversion of CO and H2 was improved by 30%. Nevertheless, increased pH to 8.6 resulted in acetic acid accumulation (>7.5 g/L). These findings improve syngas biomethanation efficiency and suggest including a proper pH control before further scale-up.

“Biomethanation of syngas by enriched mixed anaerobic consortium in pressurized agitated column”

In a circular economy approach, heterogeneous wastes can be upgraded to energy in the form of syngas via pyrogasification, and then to methane via biomethanation. Working at high pressure is a promising approach to intensify the process and to reduce gas–liquid transfer limitations. However, raising the pressure could lead to reaching the CO inhibition threshold of the microorganisms involved in syngas-biomethanation. To investigate the impact on pressure on the process, a 10L continuous stirred tank reactor working at 4 bars and 55 °C was implemented. Syngas (40% CO, 40% H2, 20% CO2) biomethanation was performed successfully and methane productivity as high as 6.8 mmolCH4/Lreactor/h with almost full conversion of CO (97%) and H2 (98%) was achieved. CO inhibition was investigated and carboxydotrophs appeared less resistant to high CO exposition than methanogens.

Biofuel generation from high-fat effluent in a biofilm reactor with backwash

ABSTRACT The performance of an up-flow 0.5 m high and 0.076 m diameter fixed-film bioreactor treating lipid-rich synthetic wastewater has been investigated under the present study. The initial porosity of the filter bed using immobilised microorganisms were on the support matrices has been measured to be 0.67. Excess biomass accumulation on the support leads to an increase in biofilm thickness with time. The filter bed has been regenerated by backwashing after the stipulated time to regain its efficiency in terms of COD removal through the revival of the contact surface. A mathematical model has been developed to predict the loss of contact area and decrease in the porosity of bed with operation time. The backwashing time has been correlated to the inlet concentration of substrate. Methanogens like Methanococcus sp. and Methanosarcina sp. are detected in the mixed anaerobic consortia and favourable growth condition for methanogens in the reactor leads to a maximum cumulative methane production at the exit of the reactor estimated to be 785 ml CH4 (STP)/(g VSS added). The originality of the present research lies in investigating erucic acid-rich wastewater in a bioreactor with backwash and proposing an integration of this bioconversion with syngas-based 2G-Biorefinery.

ANAMMOX-denitrification biomass in microbial fuel cell to enhancethe electricity generation and nitrogen removal efficiency.

The inoculum biomass was collected from a pilot-scale (3 m3 process tank) nitritation-anaerobic ammonium oxidation (ANAMMOX) (deammonification moving bed biofilm (DeaMBBR)) reactor demonstrating the highest total nitrogen removal rate (TNRR) of 0.33kgNm-3day-1. This biomass was used for inoculating the anodic chamber of a microbial fuel cell (MFC) to investigate the capacity of DeaMBBR biomass to act as an exo-electrogenic consortia. Performance of MFCs inoculated with ANAMMOX-specific consortia collected from DeaMBBR (MFC-ANA) and another MFC-CON inoculated with a septic tank mixed anaerobic consortium as a control was investigated for electrochemical performance and wastewater treatment efficiency. These MFCs were operated for the total duration of 419days during which regular feed was given and performance was monitored for first 30 cycles and last 30 cycles, with each cycle of 3day duration. The MFC-ANA continuously generated bio-energy with higher volumetric power density (9.5Wm-3 and 6.0Wm-3) in comparison to MFC-CON (4.9 and 2.9Wm-3) during the first 30 and last 30 cycles of operational period, respectively. MFC-ANA also achieved 84 ± 2% and 80 ± 2% of COD removal efficiency and 89 ± 4% and 73 ± 2% of total nitrogen removal efficiency during first 30 and last 30 cycles of operational period, respectively. The improvement ofnitrogen removal and power production in case of MFC-ANA over MFC-CON could be attributed to the ANAMMOX-denitrifiers populations and Trichococcus (14.92%) as denitrifying exo-electrogenic microbes (4.46%), respectively.

Improvement of waste-fed bioelectrochemical system performance by selected electro-active microbes: Process evaluation and a kinetic study

In this work, bioaugmentation strategy was tested to enhance electricity production efficiency from municipal waste liquor feedstock in microbial fuel cells (MFC). During the experiments, MFCs inoculated with a mixed anaerobic consortium were enriched by several pure, electro-active bacterial cultures (such as Propionibacterium freudenreichii, Cupriavidus basilensis and Lactococcus lactis) and behaviours were assessed kinetically. It turned out that energy yield could be enhanced mainly at high substrate loadings. Furthermore, energy production and COD removal rate showed an optimum and could be characterized by a saturation range within the applied COD loadings, which could be elucidated applying the Monod-model for describing intracellular losses. Polarization measurements showed the positive effect of bioaugmentation also on extracellular losses. The data indicated a successful augmentation process for enhancing MFC efficiency, which was utmost in case of augmentation strain of Propionibacterium freudenreichii.

Long-term operation of microbial electrosynthesis cell reducing CO2 to multi-carbon chemicals with a mixed culture avoiding methanogenesis

In microbial electrosynthesis (MES), CO2 can be reduced preferably to multi-carbon chemicals by a biocathode-based process which uses electrochemically active bacteria as catalysts. A mixed anaerobic consortium from biological origin typically produces methane from CO2 reduction which circumvents production of multi-carbon compounds. This study aimed to develop a stable and robust CO2 reducing biocathode from a mixed culture inoculum avoiding the methane generation. An effective approach was demonstrated based on (i) an enrichment procedure involving inoculum pre-treatment and several culture transfers in H2:CO2 media, (ii) a transfer from heterotrophic to autotrophic growth and (iii) a sequential batch operation. Biomass growth and gradual acclimation to CO2 electro-reduction accomplished a maximum acetate production rate of 400mgLcatholyte−1d−1 at −1V (vs. Ag/AgCl). Methane was never detected in more than 300days of operation. Accumulation of acetate up to 7–10gL−1 was repeatedly attained by supplying (80:20) CO2:N2 mixture at −0.9 to −1V (vs. Ag/AgCl). In addition, ethanol and butyrate were also produced from CO2 reduction. Thus, a robust CO2 reducing biocathode can be developed from a mixed culture avoiding methane generation by adopting the specific culture enrichment and operation procedures without the direct addition of chemical inhibitor.

Simultaneous Removal of Phenol and Dissolved Solids from Wastewater Using Multichambered Microbial Desalination Cell.

Microbial desalination cell (MDC) has great potential toward direct electricity generation from wastewater and concurrent desalination through potential difference developed due to microbial activity. Degradation of phenol by isolate Pseudomonas aeruginosa in anodic chamber and simultaneous desalination of water in middle desalination chamber of multichamber MDC is demonstrated in this study. Performance of the MDCs with different anodic inoculum conditions, namely pure culture of P. aeruginosa (MDC-1), 50% v/v mixture of P. aeruginosa and anaerobic mixed consortia (MDC-2) and anaerobic mixed consortia (MDC-3), was evaluated to compare the phenol degradation in anodic chamber, bioelectricity generation, and simultaneous total dissolved solids (TDS) removal from saline water in desalination chamber. Synergistic effect between P. aeruginosa and mixed anaerobic consortia as inoculum was evident in MDC-2 demonstrating phenol degradation of 90%, TDS removal of 75% in 72h of reaction time along with higher power generation of 27.5mW/m(2) as compared to MDC-1 (95%, 64%, 12.8mW/m(2), respectively) and MDC-3 (58%, 52%, 4.8mW/m(2), respectively). The results illustrate that the multichamber MDC-2 is effective for simultaneous removal of phenol and dissolved solids contained in industrial wastewaters.

Specific inhibitors for identifying pathways for methane production from carbon monoxide by a nonadapted anaerobic mixed culture

Specific inhibitors such as 2-bromoethanesulfonate (BES) and vancomycin were employed in activity batch tests to decipher metabolic pathways that are preferentially used by a mixed anaerobic consortium (sludge from an anaerobic digester) to transform carbon monoxide (CO) into methane (CH4). We first evaluated the inhibitory effect of both BES and vancomycin on the microbial community, as well as the efficiency and stability of vancomycin at 35 °C, over time. The activity tests with CO2-H2, CO, glucose, acetate, formate, propionate, butyrate, methanol, and ethanol showed that vancomycin does not inhibit some Gram-negative bacteria, and 50 mmol/L BES effectively blocks CH4 production in the sludge. However, when sludge was incubated with propionate, butyrate, methanol, or ethanol as the sole energy and carbon source, methanogenesis was only partially inhibited by BES. Separate tests showed that 0.07 mmol/L vancomycin is enough to maintain its inhibitory efficiency and stability in the population for at least 32 days at 35 °C. Using the inhibitors above, it was demonstrated that CO conversion to CH4 is an indirect, 2-step process, in which the CO is converted first to acetate and subsequently to CH4.

Ozone pretreatment of wheat straw for enhanced biohydrogen production

Ozonation was tested as a pretreatment method for enhanced biohydrogen production from wheat straw. Ozone pretreatment effectively degraded wheat straw lignin, and the delignification increased with increase in the applied ozone dose. Results of reducing sugar measurement showed that under our experimental conditions ozone pretreatment significantly increased reducing sugar yields. A simultaneous enzyme hydrolysis and dark fermentation experiment was then conducted using a mixed anaerobic consortium, and the results demonstrated that ozone pretreatment significantly increased biohydrogen production. Compared to the untreated one, hydrogen production in the samples ozonated for 15, 30, 45 and 90 min increased 107%, 134%, 158% and 138%, respectively. Slight inhibitory effect on the dark fermentation was observed with the sample ozonated for 90 min, and the inhibitory effect was due to prolonged ozonation. These results proved that enhancement of biohydrogen production from lignocellulosic biomass using ozone as a pretreatment method is technically feasible.

Effect of ozone pretreatment on hydrogen production from barley straw

Application of ozone technology to lignocellulosic biohydrogen production was explored with a barley straw. Ozone pretreatment effectively degraded the straw lignin and increased reducing sugar yield. A simultaneous enzyme hydrolysis and dark fermentation experiment was conducted using a mixed anaerobic consortium together with saccharification enzymes. Both untreated and ozonated samples produced hydrogen. Compared to the untreated group, hydrogen produced by the groups ozonated for 15, 30, 45 and 90 min increased 99%, 133%, 166% and 94%, respectively. Some inhibitory effect on hydrogen production was observed with the samples ozonated for 90 min, and the inhibition was on the fermentative microorganisms, not the saccharification enzymes. These results demonstrate that production of biohydrogen from barley straw, a lignocellulosic biomass, can be significantly enhanced by ozone pretreatment.

Fermentative hydrogen production from beet sugar factory wastewater treatment in a continuous stirred tank reactor using anaerobic mixed consortia

A low pH, ethanol-type fermentation process was evaluated for wastewater treatment and bio-hydrogen production from acidic beet sugar factory wastewater in a continuous stirred tank reactor (CSTR) with an effective volume of 9.6 L by anaerobic mixed cultures in this present study. After inoculating with aerobic activated sludge and operating at organic loading rate (OLR) of 12 kgCOD·m−3·d−1, HRT of 8h, and temperature of 35°C for 28 days, the CSTR achieved stable ethanol-type fermentation. When OLR was further increased to 18 kgCOD·m−3·d−1 on the 53rd day, ethanol-type fermentation dominant microflora was enhanced. The liquid fermentation products, including volatile fatty acids (VFAs) and ethanol, stabilized at 1493 mg·L−1 in the bioreactor. Effluent pH, oxidation-reduction potential (ORP), and alkalinity ranged at 4.1–4.5, −250-(−290) mV, and 230–260 mgCaCO3·L−1. The specific hydrogen production rate of anaerobic activated sludge was 0.1 L·gMLVSS−1·d−1 and the COD removal efficiency was 45%. The experimental results showed that the CSTR system had good operation stability and microbial activity, which led to high substrate conversion rate and hydrogen production ability.

Metabolic shift and electron discharge pattern of anaerobic consortia as a function of pretreatment method applied during fermentative hydrogen production

We have made an attempt to evaluate the variation in the electron discharge (ED) pattern of anaerobic consortia as a function of pretreatment viz., chemical, heat-shock, acid and oxygen-shock in comparison with untreated mixed consortia during fermentative hydrogen (H 2) production. Experiments were performed with dairy wastewater as substrate using anaerobic mixed consortia as biocatalyst (pretreated individually and in combination). Cyclic voltammetry (CV) elucidated significant variation in the ED pattern of mixed consortia along with H 2 production and substrate degradation (SD) as a function of pretreatment method applied. Higher ED was observed with all pretreated consortia which can be attributed to the stable proton (H +) shuttling due to the suppression of methanogenic activity. Oxygen-shock method and untreated consortia showed lower H 2 production and higher SD among the variations studied, while, combined pretreated consortia resulted higher H 2 production and lower SD. Lower ED observed with untreated consortia suggests the H + reduction during methanogenesis rather than the inter-conversion of metabolites, which is presumed to be necessary for H 2 production. ED observed with combined pretreated consortia corroborated well with the observed H 2 production. Redox pairs were visualized on the voltammograms with almost all the experimental variations studied except untreated consortia. The potentials ( E 0) of redox pairs observed were corresponding to intracellular electron carriers viz., NAD +/NADH ( E 0 −0.32 V) and FAD +/FADH 2 ( E 0 −0.24 V).

Biohydrogen production process optimization using anaerobic mixed consortia: A prelude study for use of agro-industrial material hydrolysate as substrate

Efficient biohydrogen production from lignocellulosic hydrolysate assumes considerable practical and academic importance. The impact of glucose to xylose ratio, medium pH, inoculum size and age on biohydrogen production indicated that glucose to xylose ratio is the critical parameter for effective H 2 production compared to either pure glucose or xylose as carbon source. Inoculum size and its age contributed more than 70% to overall H 2 production and revealed significance at individual as well as interactive level. Maximum interaction of 39% and 32% was noticed with inoculum size and its age vs. glucose to xylose ratio (2:3), respectively. The H 2 production yield enhanced from 140 to 357 ml/g substrate upon statistical optimization revealing >240% improvement.

Fermentative biohydrogen production by mixed anaerobic consortia: Impact of glucose to xylose ratio

Glucose and xylose are the dominant monomeric carbohydrates present in agricultural materials which can be used as potential building blocks for various biotechnological products including biofuels production. Hence, the imperative role of glucose to xylose ratio on fermentative biohydrogen production by mixed anaerobic consortia was investigated. Microbial catabolic H 2 and VFA production studies revealed that xylose is a preferred carbon source compared to glucose when used individually. A maximum of 1550 and 1650 ml of cumulative H 2 production was observed with supplementation of glucose and xylose at a concentration of 5.5 and 5.0 g L −1, respectively. A triphasic pattern of H 2 production was observed only with studied xylose concentration range. pH impact data revealed effective H 2 production at pH 6.0 and 6.5 with xylose and glucose as carbon sources, respectively. Co-substrate related biohydrogen fermentation studies indicated that glucose to xylose ratio influence H 2 and as well as VFA production. An optimum cumulative H 2 production of 1900 ml for 5 g L −1 substrate was noticed with fermentation medium supplemented with glucose to xylose ratio of 2:3 at pH 6. Overall, biohydrogen producing microbial consortia developed from buffalo dung could be more effective for H 2 production from lignocellulosic hydrolysates however; maintenance of glucose to xylose ratio, inoculum concentration and medium pH would be essential requirements.

Harnessing of biohydrogen by acidogenic fermentation of Citrus limetta peelings: Effect of extraction procedure and pretreatment of biocatalyst

The extracts of Citrus limetta (sweet lime) peelings were evaluated as a fermentable substrate for hydrogen (H2) production by dark-fermentation (acidogenic) using both anaerobic mixed consortia and selectively enriched acidogenic mixed consortia. Extraction was carried by pretreating sweet lime peelings at 121 °C (1 bar pressure) at variable pH (6 and 7) and digestion time (20 and 40 min). Maximum organic matter extraction was observed at pH 7.0 (40 min). Fermentation was performed at different organic loading conditions [OL1, 1.17 kg COD/m3; OL2, 2.35 kg COD/m3; OL3, 4.69 kg COD/m3] under acidophilic microenvironment. H2 production was found to depend on the concentration of the substrate and composition. Increase in organic load showed consistent improvement in H2 production. Operation at OL3 employing selectively enriched inoculum documented higher cumulative H2 production (10.07 mmol) and H2 production rate (0.345 mmol/h) (pH 7; 40 min). Substrate degradation was also found to increase with increase in organic loading. Maximum substrate degradation (SD) was registered at pH 6 (40 min) with anaerobic culture (2.80 kg CODR/m3; ξCOD 31.82%) and at pH 7 (40 min) with selectively enriched acidogenic culture (3.20 kg CODR/m3; ξCOD 36.36%). Concentration of volatile fatty acids (VFAs) also improved with increase in organic load. Maximum VFA concentration (1098 mg/l) was observed with OL3 (pH 7; 40 min) by using selectively enriched culture.

Non-catalyzed microbial fuel cell (MFC) with open air cathode for bioelectricity generation during acidogenic wastewater treatment

Single chambered mediatorless microbial fuel cell (MFC Nafion-117 membrane) fabricated with non-catalyzed electrodes was operated with open-air cathode to evaluate bioelectricity generation from domestic wastewater under acidogenic conditions (pH, 6) using anaerobic mixed consortia as anodic biocatalyst. Experimental data illustrated the feasibility of bioelectricity generation from domestic wastewater treatment. A steady increase in MFC performance was observed from the first cycle (0.248 V; 27.3 mW/m 2; 1.06 W/kg COD R) during the startup phase prior to stabilization on fourth cycle (0.449 V; 144.6 mW/m 2; 4.64 W/kg COD R). Sharp increase in power generation was observed after the fourth hour (125.4 mW/m 2; 289.61 mA/m 2) which continued up to the sixth hour (155.92 mW/m 2; 325.51 mA/m 2) and gradually decreased thereafter. Voltammogram evidenced clear redox peaks (E 0′, − 0.334 V) related to redox mediator NAD +/NADH (E 0′, − 0.32 V) suggesting a strong reducing phase. Higher energy (1.33 J) was observed at the fourth hour in concurrence with the effective electron discharge and higher substrate degradation.

Bio-electrochemical evaluation of fermentative hydrogen production process with the function of feeding pH

Fermentative hydrogen (H 2) production process in concurrence with feeding pH [aciodophilic (pH 6.0) and neutral (pH 7.0)] and reactor operation mode (continuous and fed-batch) was evaluated in a biofilm configured reactor [upflow mode; retention time, 24 h; operating temperature, 28 ± 2 °C; organic loading rate, 3.4 kg COD/m 3 day] using anaerobic mixed consortia. Acidophilic pH showed relatively effective performance with respect to H 2 production compared to neutral operation. Neutral pH illustrated effective substrate removal efficiency over the corresponding acidophilic operation. Fed-batch mode of operation with acidophilic pH showed highest H 2 production among the studied experimental variations. The pattern of soluble metabolites distribution showed the persistence of acid-forming metabolic flow associated with acidogenesis which may be considered as optimum microenvironment for effective H 2 production. Bio-electrochemical behavior of mixed anaerobic consortia (whole cell) during H 2 production process was evaluated employing cyclic voltammetry (CV) in electrochemical cell [platinum as working electrode; Ag/AgCl as reference electrode; graphite rod as counter electrode; wastewater as electrolyte] to gain insight into the possible mechanism based on intracellular electron transfer involved in the fermentative metabolic process. Voltammogram profiles visualized well defined redox pairs in forward and reverse scans at both pH conditions and the signals corresponded to intracellular electron carrier, NADH/NAD + ( E 0′, −0.32 V). Relatively higher energy output was observed in acidophilic operation which might be attributed to the possibility of efficient proton (H +) transfer between metabolic intermediates.

Biological hydrogen production from probiotic wastewater as substrate by selectively enriched anaerobic mixed microflora

Biohydrogen production from probiotic wastewater using mixed anaerobic consortia is reported in this paper. Batch tests are carried out in a 5.0 L batch reactor under constant mesophillic temperature (37 °C). The maximum hydrogen yield 1.8 mol-hydrogen/mol-carbohydrate is obtained at an optimum pH of 5.5 and substrate concentration 5 g/L. The maximum hydrogen production rate is 168 ml/h. The hydrogen content in the biogas is more than 65% and no significant methane is observed throughout the study. In addition to hydrogen, acetate, propionate, butyrate and ethanol are found to be the main by-products in the metabolism of hydrogen fermentation.

Influence of anodic biofilm growth on bioelectricity production in single chambered mediatorless microbial fuel cell using mixed anaerobic consortia

The effect of anodic biofilm growth and extent of its coverage on the anodic surface of a single chambered mediatorless microbial fuel cell (MFC) was evaluated for bioelectricity generation using designed synthetic wastewater (DSW) and chemical wastewater (CW) as substrates and anaerobic mixed consortia as biocatalyst. Three MFCs (plain graphite electrodes, air cathode, Nafion membrane) were operated separately with variable biofilm coverage [control; anode surface coverage (ASC), 0%], partially developed biofilm [PDB; ASC ∼44%; 90 days] and fully developed biofilm [FDB; ASC ∼96%; 180 days] under acidophilic conditions (pH 6) at room temperature. The study depicted the effectiveness of anodic biofilm formation in enhancing the extracellular electron transfer in the absence of mediators. Higher specific power production [29 mW/kg COD R (CW and DSW)], specific energy yield [100.46 J/kg VSS (CW)], specific power yield [0.245 W/kg VSS (DSW); 0.282 W/kg VSS (CW)] and substrate removal efficiency of 66.07% (substrate degradation rate, 0.903 kg COD/m 3-day) along with effective functioning fuel cell at relatively higher resistance [4.5 kΩ (DSW); 14.9 kΩ (CW)] correspond to sustainable power [0.008 mW (DSW); 0.021 mW (CW)] and effective electron discharge (at higher resistance) and recovery (Coulomb efficiency; 27.03%) were observed especially with FDB operation. Cyclic voltammetry analysis documented six-fold increment in energy output from control (1.812 mJ) to PDB (10.666 mJ) operations and about eight-fold increment in energy from PDB to FDB (86.856 mJ). Biofilm configured MFC was shown to have the potential to selectively support the growth of electrogenic bacteria with robust characteristics, capable of generating higher power yields along with substrate degradation especially operated with characteristically complex wastewaters as substrates.