The Impact of Preservation Techniques on Methane-Arrested Anaerobic Digestion of Nutrient-Rich Feedstocks.

The carboxylate platform is a promising biomass-to-energy pathway that uses methane-arrested anaerobic digestion (MAAD) to convert biomass to carboxylic acids, which can be chemically converted to industrial chemicals and liquid fuels. Lignocellulose is an energy-rich carbon source, but lacks nutrients necessary for microbial growth. Chicken manure (rural waste) and sewage sludge (urban waste) are rich in nitrogen and useful macronutrients; therefore, co-digesting these wastes with lignocellulose improves MAAD performance. However, waste nutrients must be digested immediately, or preserved. This study investigated the effects of various preservation techniques - frozen (fresh), air-dried, and baked - on chicken manure and sewage sludge. Batch experiments were performed with office paper (carbon source) and chicken manure or sewage sludge (nutrient source) with different methods of preservation. Fresh substrates produced higher acid yields and biomass conversion (the amount of biomass consumed during digestion) than dried substrates. Baked chicken manure showed reduced conversion and total acid production, which suggests that oven-drying reduces digestibility. From the batch data, the Continuum Particle Distribution Model (CPDM) predicted results of a four-stage countercurrent digestion. The data are displayed on maps showing the impact of liquid residence time (LRT) and volatile solids loading rate (VSLR) on conversion and product concentration. Co-digesting office paper and wet chicken manure at a non-acid volatile solid (NAVS) concentration of 300 g/Lliq, the model predicted a high total acid concentration of 52.8 g/L and conversion of 0.89 g NAVSdigested/NAVSfed at a volatile solid loading rate of 4 g/(Lliq·day) and liquid retention time of 35 days.

Valorizing prickly pear cladodes via methane-arrested anaerobic digestion for carboxylic acid production.

To address climate change, liquid biofuels are an essential alternative to fossil fuels, especially for transportation. The carboxylate platform uses methane-arrested anaerobic digestion (MAAD) to ferment biomass to carboxylic acids, which can be chemically converted to liquid fuels via the carboxylate platform. Most biomass sources require expensive pretreatments to remove lignin; however, prickly pear (Opuntia ficus-indica) cladodes have low lignin content and therefore do not require pretreatment. Furthermore, this sugar-rich feedstock is readily digested to high concentrations of carboxylic acids. At various substrate concentrations, batch MAAD of prickly pear cladodes yielded primarily acetic, butyric, and caproic acids. From these batch data, continuum particle distribution modeling (CPDM) simulated four-stage countercurrent digestion. At a non-acid volatile solid (NAVS) concentration of 100 g/Lliq , CPDM predicts a high total acid concentration of 93 g/L and conversion of 0.93 g NAVSdigested /NAVSfed at a volatile solid loading rate of 6 g/(Lliq ·d) and liquid retention time of 35 days. Without chemical pretreatment, co-digestion, or in situ product removal, prickly pear produced high yields, biomass conversion, product concentration, and selectivity compared to previously studied lignocellulosic feedstocks.

Conversion of acetone and mixed ketones to hydrocarbons using HZSM-5 catalyst in the carboxylate platform.

In this study, two different feeds were treated to produce hydrocarbons: (1) reagent-grade acetone, and (2) mixed ketones obtained from lignocellulosic biomass via the carboxylate platform. Acetone and mixed ketones underwent catalytic self-condensation over HZSM-5. For acetone, HZSM-5(80) was used, and the experiments were conducted in two sets: (1) vary temperature (305-415°C) at P = 101 kPa (abs) and weight hourly space velocity (WHSV) = 1.3 h-1; (2) vary WHSV (1.3-7.9 h-1) at T = 350 and 415°C, and P = 101 kPa (abs). For acetone over HZSM-5(280), the experiments were conducted in two sets: (1) vary WHSV (1.3-6.5 h-1) at T = 415°C, and P = 101 kPa (abs); and (2) vary WHSV (1.3-11.8 h-1) at P = 790 kPa (abs) and T = 415°C. For mixed ketones, HZSM-5(280) was used at WHSV = 1.9 h-1, T = 430-590°C, and P = 101 kPa (abs). For acetone at higher temperatures, the conversion was 100% and the liquid products were aromatics centered on C8. At low temperatures, conversion was less and the carbon liquid distribution was centered on C9 (mainly mesitylene). For mixed ketones, catalyst deactivation was higher causing product concentrations to change over time, and the highest conversion reached was 40%.

Assessment of corn stover pretreated with shock and alkali using methane-arrested anaerobic digestion.

Corn stover, an underutilized agricultural residue, is a promising lignocellulosic feedstock for producing biofuels. To fully utilize it, pretreatment is needed. Typically, pretreatments are rapidly assessed using extracellular enzymes that release sugars from cellulose and hemicellulose. In contrast, this study uses methane-arrested anaerobic digestion (MAAD) to assess pretreatments. Although time consuming, MAAD is a more accurate assessment technique when lignocellulose is employed in the carboxylate platform, a promising approach that utilizes nearly all biomass components. Using recommended pretreatment conditions identified from a previous study, three corn stover pretreatments were compared using MAAD: (1) shock-only, (2) NaOH-only, and (3) shock + NaOH. Air-dried sewage sludge was used as nutrient source. At 100 g/L initial substrate concentration, compared to untreated corn stover, shock-only decreased conversion (amount of biomass digested) by 14%, NaOH-only increased conversion by 82%, and shock + NaOH increased conversion by 104%. Using batch MAAD data, the continuum particle distribution model simulated four-stage countercurrent fermentation. At an industrial non-acid volatile solids (NAVS) concentration of 300 g/Lliq , for both NaOH-only and shock + NaOH, the model predicts total carboxylic acid concentration of about 58 g/L and conversion of about 0.85 g NAVSdigested /g NAVSfed at liquid retention time of 35 days and volatile solid loading rate of 4 g/(Lliq ⋅day). At this long solid residence time, shock is not necessary; however, with short solid residence times, shock acts synergistically to aid NaOH pretreatment. Shock treatment offers a way to reduce pretreatment costs without sacrificing pretreatment efficacy.

Assessment of shock pretreatment and alkali pretreatment on corn stover using enzymatic hydrolysis.

This study investigates digestibility enhancements of lignocellulose from shock pretreatment, alkaline pretreatment, and combination. Shock pretreatment subjects aqueous slurries of lignocellulose to shock waves, which disrupts its structure rendering it more susceptible to hydrolysis. Alkaline pretreatment submerges the biomass in aqueous alkali (NaOH, Ca(OH)2 ), which removes lignin and acetyl groups. As indicators of digestibility, cellulase (CTec3) and hemicellulase (HTec3) were used to saccharify the pretreated corn stover and the resulting filtrate which contains about 10% of the sugars. Shock is most effective when it precedes alkaline pretreatment, presumably because it opens the biomass structure and enhances diffusion of pretreatment chemicals. Lignocellulose digestibility from calcium hydroxide treatment improves significantly with oxygen addition. In contrast, sodium hydroxide is a more potent alkali, and thereby eliminates the need for oxygen to enhance pretreatment. At low hydroxide loadings (<4 g OH- /100 g dry biomass), both NaOH and Ca(OH)2 provide similar increases in digestibility; however, at high hydroxide loadings, NaOH is superior. For animal feed, Ca(OH)2 treatment is recommended, because residual calcium ions are valuable nutrients. In contrast, for methane-arrested anaerobic digestion, NaOH treatment is preferred because NaHCO3 is a stronger buffer. At 50°C, shock pretreatment improves sugar yields at all NaOH loadings. The effect of shock is most pronounced when the no-shock control employed the same soaking-and-drying procedure as the shock treatment. The recommended conditions are shock treatment (5.52 bar [abs] initial H2 /O2 pressure) followed by 50°C alkaline treatment with NaOH loading of 4 g OH- /100 g dry biomass for 1 h.

Microbial communities for valorizing biomass using the carboxylate platform to produce volatile fatty acids: A review.

The carboxylate platform employs a diverse microbial consortium of anaerobes in which the methanogens are inhibited. Nearly all biomass components are digested to a mixture of C1-C8 monocarboxylic acids and their corresponding salts. The methane-arrested anaerobic digestion proceeds readily without needing to sterilize biomass or equipment. It accepts a wide range of feedstocks (e.g., agricultural residues, municipal solid waste, sewage sludge, animal manure, food waste, algae, and energy crops), and produces high product yields. This review highlights several important aspects of the platform, including its thermodynamic underpinnings, influences of inoculum source and operating conditions on product formation, and downstream chemical processes that convert the carboxylates to hydrocarbon fuels and oxygenated chemicals. This review further establishes the carboxylate platform as a viable and economical route to industrial biomass utilization.

Arrested methanogenesis digestion of high-strength cheese whey and brewery wastewater with carboxylic acid production.

A new anaerobic digestion process based on arrested methanogenesis (AM) was developed to treat high-strength cheese whey and brewery wastewater with simultaneous carboxylic acid production. This study specifically determined the links between wastewater characteristics, microbial community structure, and the operation of AM digesters at the bench scale. The highest total carboxylic acids concentration (78 g/L) was achieved after 15 days under batch condition at 40 °C and near-neutral pH. Lactate conversion to chain-elongated volatile fatty acid was observed. Under fed-batch conditions, the highest total acid productivity was 16 g/(Lliq·d) with substrate conversion of 0.66 g CODdigested/g CODfed at hydraulic residence time (HRT) of 4 days. Fed-batch digestion with biomass recycling resulted in a 2-fold increase in VFAs concentration (30 g/L) and a higher diversity in the microbial consortia. Experimental results show that highly efficient, robust, and productive community structure was established for sustainable carboxylate production from widely varying high-strength wastewaters.

Kinetic modeling of countercurrent saccharification.

BACKGROUND: Countercurrent saccharification is a promising way to minimize enzyme loading while obtaining high conversions and product concentrations. However, in countercurrent saccharification experiments, 3-4 months are usually required to acquire a single steady-state data point. To save labor and time, simulation of this process is necessary to test various reaction conditions and determine the optimal operating point. Previously, a suitable kinetic model for countercurrent saccharification has never been reported. The Continuum Particle Distribution Modeling (CPDM) satisfactorily predicts countercurrent fermentation using mixed microbial cultures that digest various feedstocks. Here, CPDM is applied to countercurrent enzymatic saccharification of lignocellulose. RESULTS: CPDM was used to simulate multi-stage countercurrent saccharifications of a lignocellulose model compound (α-cellulose). The modified HCH-1 model, which accurately predicts long-term batch saccharification, was used as the governing equation in the CPDM model. When validated against experimental countercurrent saccharification data, it predicts experimental glucose concentrations and conversions with the average errors of 3.5% and 4.7%, respectively. CPDM predicts conversion and product concentration with varying enzyme-addition location, total stage number, enzyme loading, liquid residence time (LRT), and solids loading rate (SLR). In addition, countercurrent saccharification was compared to batch saccharification at the same conversion, product concentration, and reactor volume. Results show that countercurrent saccharification is particularly beneficial when the product concentration is low. CONCLUSIONS: The CPDM model was used to simulate multi-stage countercurrent saccharification of α-cellulose. The model predictions agreed well with the experimental glucose concentrations and conversions. CPDM prediction results showed that the enzyme-addition location, enzyme loading, LRT, and SLR significantly affected the glucose concentration and conversion. Compared to batch saccharification at the same conversion, product concentration, and reactor volume, countercurrent saccharification is particularly beneficial when the product concentration is low.

Development of modified HCH-1 kinetic model for long-term enzymatic cellulose hydrolysis and comparison with literature models.

BACKGROUND: Enzymatic hydrolysis is a major step for cellulosic ethanol production. A thorough understanding of enzymatic hydrolysis is necessary to help design optimal conditions and economical systems. The original HCH-1 (Holtzapple-Caram-Humphrey-1) model is a generalized mechanistic model for enzymatic cellulose hydrolysis, but was previously applied only to the initial rates. In this study, the original HCH-1 model was modified to describe integrated enzymatic cellulose hydrolysis. The relationships between parameters in the HCH-1 model and substrate conversion were investigated. Literature models for long-term (> 48 h) enzymatic hydrolysis were summarized and compared to the modified HCH-1 model. RESULTS: A modified HCH-1 model was developed for long-term (> 48 h) enzymatic cellulose hydrolysis. This modified HCH-1 model includes the following additional considerations: (1) relationships between coefficients and substrate conversion, and (2) enzyme stability. Parameter estimation was performed with 10-day experimental data using α-cellulose as substrate. The developed model satisfactorily describes integrated cellulose hydrolysis data taken with various reaction conditions (initial substrate concentration, initial product concentration, enzyme loading, time). Mechanistic (and semi-mechanistic) literature models for long-term enzymatic hydrolysis were compared with the modified HCH-1 model and evaluated by the corrected version of the Akaike information criterion. Comparison results show that the modified HCH-1 model provides the best fit for enzymatic cellulose hydrolysis. CONCLUSIONS: The HCH-1 model was modified to extend its application to integrated enzymatic hydrolysis; it performed well when predicting 10-day cellulose hydrolysis at various experimental conditions. Comparison with the literature models showed that the modified HCH-1 model provided the best fit.

Energy Portfolio Assessment Tool (EPAT): Sustainable energy planning using the WEF nexus approach - Texas case.

The paper introduces a holistic framework that identifies the links between energy and other systems (water, land, environment, finance, etc.), and measures the impact of energy portfolios, to offer a solid foundation for the best sustainable decision making in energy planning. The paper presents a scenario-based holistic nexus tool, Energy Portfolio Assessment Tool (EPAT) that provides a platform for energy stakeholders and policymakers to create and evaluate the sustainability of various scenarios based on the water-energy-food (WEF) nexus approach. The tool is applied to a case study in Texas, USA. Scenarios considered are set by the U.S. Energy Information Administration (EIA): EIA Reference Case - 2015, EIA Clean Power Plan (CPP) & Reference Case - 2030, and EIA No-CPP & Reference Case - 2030. In the presence of the CPP, total water withdrawal is expected to decrease significantly, while total water consumption is projected to experience a slight decrease due to the increase in water consumption in electricity generation caused by the new electricity mix. The CPP is successful in decreasing emissions, but is accompanied by tradeoffs, such as increased water consumption and land use by electricity generation. The absence of the CPP will lead to an extreme surge in total water withdrawn and consumed, and in emissions. Therefore, conservation policies should move from the silo to the nexus mentality to avoid unintended consequences that result in improving one part of the nexus while worsening the other parts.

Insights into Xylan Degradation and Haloalkaline Adaptation through Whole-Genome Analysis of Alkalitalea saponilacus, an Anaerobic Haloalkaliphilic Bacterium Capable of Secreting Novel Halostable Xylanase.

The obligately anaerobic haloalkaliphilic bacterium Alkalitalea saponilacus can use xylan as the sole carbon source and produce propionate as the main fermentation product. Using mixed carbon sources of 0.4% (w/v) sucrose and 0.1% (w/v) birch xylan, xylanase production from A. saponilacus was 3.2-fold greater than that of individual carbon sources of 0.5% (w/v) sucrose or 0.5% (w/v) birch xylan. The xylanse is halostable and exhibits optimal activity over a broad salt concentration (2⁻6% NaCl). Its activity increased approximately 1.16-fold by adding 0.2% (v/v) Tween 20. To understand the potential genetic mechanisms of xylan degradation and molecular adaptation to saline-alkali extremes, the complete genome sequence of A. saponilacus was performed with the pacBio single-molecule real-time (SMRT) and Illumina Misseq platforms. The genome contained one chromosome with a total size of 4,775,573 bps, and a G+C genomic content of 39.27%. Ten genes relating to the pathway for complete xylan degradation were systematically identified. Furthermore, various genes were predicted to be involved in isosmotic cytoplasm via the "compatible-solutes strategy" and cytoplasmic pH homeostasis though the "influx of hydrogen ions". The halostable xylanase from A. saponilacus and its genomic sequence information provide some insight for potential applications in industry under double extreme conditions.

Draft Genome Sequence of Natranaerobius trueperi DSM 18760T, an Anaerobic, Halophilic, Alkaliphilic, Thermotolerant Bacterium Isolated from a Soda Lake.

The anaerobic, halophilic, alkaliphilic, thermotolerant bacterium Natranaerobius trueperi was isolated from a soda lake in Wadi An Natrun, Egypt. It grows optimally at 3.7 M Na+, pH 9.5, and 43°C. The draft genome consists of 2.63 Mb and is composed of 2,681 predicted genes. Genomic analysis showed that various genes are potentially involved in the adaptation mechanisms for osmotic stress, pH homeostasis, and high temperatures.

Draft Genome Sequence of Natronolimnobius baerhuensis CGMCC 1.3597T, an Aerobic Haloalkaliphilic Archaeon Isolated from a Soda Lake.

The haloalkaliphilic archaeon Natronolimnobius baerhuensis was isolated from a soda lake in Inner Mongolia (China), growing optimally at about 20% NaCl and pH 9.0. The draft genome consists of approximately 3.91 Mb and contains 3,810 predicted genes. Some genes that regulate intracellular osmotic stress and pH homeostasis were identified, providing insight into specific adaptations to this double-extreme environment.

Shock treatment of corn stover.

Corn stover digestibility was enhanced via shock treatment. A slurry of lime-treated corn stover was placed in a partially filled closed vessel. From the ullage space, either a shotgun shell was fired into the slurry, or a gas mixture was detonated. Various conditions were tested (i.e., pressures, depth, solids concentrations, gas mixtures). A high pressurization rate (108,000 MPa/s shotgun shells; 4,160,000 MPa/s hydrogen/oxygen detonation) was the only parameter that improved enzymatic digestibility. Stoichiometric propane/air deflagration had a low pressurization rate (37.2 MPa/s) and did not enhance enzymatic digestibility. Without shock, enzymatic conversion of lime-treated corn stover was 0.80 g glucan digested/g glucan fed with an enzyme loading of 46.7 mg protein/g glucan. With shock, the enzyme loading was reduced by ∼2× while maintaining the same conversion. Detonations are extraordinarily fast; rapidly cycling three small vessels (0.575 m3 each) every 7.5 s enables commercially relevant shock treatment (2,000 tone/day). © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:815-823, 2017.

Creating Economic Incentives for Waste Disposal in Developing Countries Using the MixAlco Process.

In rapidly growing developing countries, waste disposal is a major challenge. Current waste disposal methods (e.g., landfills and sewage treatment) incur costs and often are not employed; thus, wastes accumulate in the environment. To address this challenge, it is advantageous to create economic incentives to collect and process wastes. One approach is the MixAlco process, which uses methane-inhibited anaerobic fermentation to convert waste biomass into carboxylate salts, which are chemically converted to industrial chemicals and fuels. In this paper, humanure (raw human feces and urine) is explored as a possible nutrient source for fermentation. This work focuses on fermenting municipal solid waste (energy source) and humanure (nutrient source) in batch fermentations. Using the Continuum Particle Distribution Model (CPDM), the performance of continuous countercurrent fermentation was predicted at different volatile solid loading rates (VSLR) and liquid residence times (LRT). For a four-stage countercurrent fermentation system at VSLR = 4 g/(L∙day), LRT = 30 days, and solids concentration = 100 g/L liquid, the model predicts carboxylic acid concentration of 68 g/L and conversion of 78.5 %.

Consolidated bioprocessing of microalgal biomass to carboxylates by a mixed culture of cow rumen bacteria using anaerobic sequencing batch reactor (ASBR).

This study employed mixed-culture consolidated bioprocessing (CBP) to digest microalgal biomass in an anaerobic sequencing batch reactor (ASBR). The primary objectives are to evaluate the impact of hydraulic residence time (HRT) on the productivity of carboxylic acids and to characterize the bacterial community. HRT affects the production rate and patterns of carboxylic acids. For the 5-L laboratory-scale fermentation, a 12-day HRT was selected because it offered the highest productivity of carboxylic acids and it synthesized longer chains. The variability of the bacterial community increased with longer HRT (R2=0.85). In the 5-L laboratory-scale fermentor, the most common phyla were Firmicutes (58.3%), Bacteroidetes (27.4%), and Proteobacteria (11.9%). The dominant bacterial classes were Clostridia (29.8%), Bacteroidia (27.4%), Tissierella (26.2%), and Betaproteobacteria (8.9%).

Optimum alcohol concentration for chain elongation in mixed-culture fermentation of cellulosic substrate.

Medium-chain fatty acids (MCFA, e.g., caproic, heptanoic, caprylic acid) are more valuable than short-chain fatty acids (SCFA, e.g., acetic, propionic, butyric, valeric acid). SCFAs are major products in methane-inhibited mixed-culture anaerobic fermentation. By feeding ethanol to the fermentor, MCFA formation is enhanced through chain elongation. Microorganisms such as Clostridium kluyveri elongate short-chain acids by combining them with alcohol. Very low ethanol concentration reduces chain elongation rates, whereas very high ethanol concentrations inhibit microorganisms. To maximize MCFA production, different ethanol concentrations were investigated in the mixed-culture fermentation of office paper and chicken manure. At 10 g/L ethanol concentration, 10 g/L MCFA was formed. High ethanol concentrations (above 40 g/L) inhibit microorganisms resulting in no chain elongation. For chain elongation, propanol was found to be more inhibitory than ethanol. The data suggest that MCFA production will increase by continuously extracting MCFA and maintaining 5-10 g/L ethanol concentration by periodic addition. Biotechnol. Bioeng. 2016;113: 2597-2604. © 2016 Wiley Periodicals, Inc.

Assessment of Shock Pretreatment of Corn Stover Using the Carboxylate Platform.

Corn stover was pretreated with lime and shock, a mechanical process that uses a shockwave to alter the biomass structure. Two pretreatments (lime-only and lime + shock) were evaluated using enzymatic hydrolysis, batch mixed-culture fermentations, and continuous countercurrent mixed-culture fermentation. In a 120-h enzymatic hydrolysis, shock pretreatment increased the glucan digestibility of submerged lime pretreatment (SLP) corn stover by 3.5 % and oxidative lime pretreatment (OLP) corn stover by 2.5 %. The continuum particle distribution model (CPDM) was used to simulate a four-stage continuous countercurrent mixed-culture fermentation using empirical rate models obtained from simple batch experiments. The CPDM model determined that lime + shock pretreatment increased the total carboxylic acids yield by 28.5 % over lime-only pretreatment in a countercurrent fermentation with a volatile solids loading rate (VSLR) of 12 g/(L/day) and liquid retention time (LRT) of 30 days. In a semi-continuous countercurrent fermentation performed in the laboratory for 112 days with a VSLR of 1.875 g/(L day) and LRT of 16 days, lime + shock pretreatment increased the total carboxylic acid yield by 14.8 %. The experimental results matched closely with CPDM model predictions (4.05 % error).

Evaluating the performance of carboxylate platform fermentations across diverse inocula originating as sediments from extreme environments.

To test the hypothesis that microbial communities from saline and thermal sediment environments are pre-adapted to exhibit superior fermentation performances, 501 saline and thermal samples were collected from a wide geographic range. Each sediment sample was screened as inoculum in a 30-day batch fermentation. Using multivariate statistics, the capacity of each community was assessed to determine its ability to degrade a cellulosic substrate and produce carboxylic acids in the context of the inoculum sediment chemistry. Conductance of soils was positively associated with production of particular acids, but negatively associated with conversion efficiency. In situ sediment temperature and conversion efficiency were consistently positively related. Because inoculum characteristics influence carboxylate platform productivity, optimization of the inoculum is an important and realistic goal.

Comparison of three screening methods to select mixed-microbial inoculum for mixed-acid fermentations.

Using a mixed culture of microorganisms, the carboxylate platform converts biomass into hydrocarbons and chemicals. To develop a method that identifies the highest performing inoculum for carboxylate fermentations, five bacterial communities were screened and ranked by three fermentation performance tests: (1) 30-day batch screen, (2) 28-day continuum particle distribution model (CPDM), and (3) 5-month continuous countercurrent fermentation trains. To screen numerous inocula sources, these tests were used sequentially in an aseptic environment. For the batch-fermentation screen, Inoculum 1 achieved the highest conversion. For the CPDM evaluation, the operating map for Inoculum 1 had the highest performance. For the continuous countercurrent fermentation, the train resulting from Inoculum 1 was among the best performers. This study suggests that the three screens are a useful and predictive method for choosing optimal inocula sources. The bacterial community with optimal performance in these three screens could be considered for use in commercial-scale fermentations.