Conversion of municipal solid waste to carboxylic acids using a mixed culture of mesophilic microorganisms

Waste biomass was anaerobically converted to carboxylate salts by using a mixed culture of acid-forming microorganisms. Municipal solid waste (MSW) was the energy source (carbohydrates) and sewage sludge (SS) was the nutrient source (minerals, metals, and vitamins). Four fermentors were arranged in series and solids and liquids were transferred countercurrently in opposite directions, which allows both high conversions and high product concentrations. Fresh biomass was added to Fermentor 1 (highest carboxylic acid concentration) and fresh media was added to Fermentor 4 (most digested biomass). All fermentations were performed at 40 degrees C. Calcium carbonate was added to the fermentors to neutralize the acids to their corresponding carboxylate salts. Iodoform was used to inhibit methane production and urea was added as a nitrogen source. Product concentrations were up to 25 g/L, with productivities up to 1.4 g total acid/(L liquid d). Mass balances with closure between 93% and 105% were obtained for all systems. Continuum particle distribution modeling (CPDM) was applied to correlate batch fermentation data to countercurrent fermentation data and predict product concentration over a wide range of solids loading rates and residence times. CPDM for lime-treated MSW/SS fermentation system predicted the experimental total acid concentration and conversion within 4% and 16% respectively.     

Anaerobic mixed‐culture fermentation of aqueous ammonia‐treated sugarcane bagasse in consolidated bioprocessing

The MixAlco process is an example of consolidated bioprocessing (CBP) in which anaerobic mixed‐culture fermentation biochemically converts any biodegradable feedstock into carboxylate salts. Downstream processing thermochemically transforms the resulting salts into mixed alcohol fuels or gasoline. To enhance digestibility, sugarcane bagasse was treated under mild conditions (55°C, 24 h, and 30% aqueous ammonia solution with a loading of 10 mL/g dry biomass). Using NH₄HCO₃ buffer, the feedstock (80% ammonia‐treated sugarcane bagasse/20% chicken manure) was anaerobically fermented by a mixed culture of marine microorganisms at 55°C. Four‐stage countercurrent fermentations were performed at various volatile solids loading rates (VSLRs) and liquid residence times (LRTs). The highest acid productivity (1.14 g/(L day)) occurred at a total acid concentration of 29.8 g/L. The highest conversion (65%) occurred at a total acid concentration of 27.6 g/L. The continuum particle distribution model (CPDM) predicted the experimental total acid concentrations and conversions within 4.98% and 10.41%, respectively. When using NH₄HCO₃ buffer, ammonia pretreatment is an attractive option. The CPDM “map” shows that both high volatile solid conversions (78.8%) and high acid concentrations (32.6 g/L) are possible with 300 g/(L liquid) substrate concentration, 30 days LRT, 2 g/(L day) solid loading rate and NH₄HCO₃ buffer. Biotechnol. Bioeng. 2010;106: 216–227. © 2010 Wiley Periodicals, Inc.     

Fermentation of corn stover to carboxylic acids

This article describes countercurrent fermentation to anaerobically convert corn stover and pig manure to mixed carboxylic acids using a mixed culture of mesophilic microorganisms. Corn stover was pretreated with lime to increase digestibility. The Continuum Particle Distribution Model (CPDM) was used to simulate continuous fermentors based on data collected from batch experiments. This model saves considerable time in determining optimum operating conditions. For 80% corn stover/20% pig manure, the highest total carboxylic acid productivity was 1.81 g/(L of liquid. d) at a concentration of 21.4 g total acid/L. The highest total acid selectivity, yield, and conversion were 0.714 g total acid/g volatile solids (VS) digested, 0.550 g total acid/g VS fed, and 0.770 g VS digested/g VS fed, respectively, at a concentration of 16.0 g total acid/L. CPDM predicted the acid concentration and conversion within 13.4 and 11.6%, respectively.     

Thermodynamic prediction of hydrogen production from mixed-acid fermentations

The MixAlco? process biologically converts biomass to carboxylate salts that may be chemically converted to a wide variety of chemicals and fuels. The process utilizes lignocellulosic biomass as feedstock (e.g., municipal solid waste, sewage sludge, and agricultural residues), creating an economic basis for sustainable biofuels. This study provides a thermodynamic analysis of hydrogen yield from mixed-acid fermentations from two feedstocks: paper and bagasse. During batch fermentations, hydrogen production, acid production, and sugar digestion were analyzed to determine the energy selectivity of each system. To predict hydrogen production during continuous operation, this energy selectivity was then applied to countercurrent fermentations of the same systems. The analysis successfully predicted hydrogen production from the paper fermentation to within 11% and the bagasse fermentation to within 21% of the actual production. The analysis was able to faithfully represent hydrogen production and represents a step forward in understanding and predicting hydrogen production from mixed-acid fermentations.     

Anaerobic thermophilic fermentation for carboxylic acid production from in-storage air-lime-treated sugarcane bagasse

Wet storage and in situ lime pretreatment (50 °C, 1-atm air, 56 days, excess lime loading of 0.3 g Ca(OH)₂/g dry biomass) of sugarcane bagasse (4,000 g dry weight) was performed in a bench-scale pile pretreatment system. Under thermophilic conditions (55 °C, NH₄HCO₃ buffer, methane inhibitors), air-lime-treated bagasse (80 wt.%) and chicken manure (20 wt.%) were anaerobically co-digested in 1-L rotary fermentors by a mixed culture of marine microorganisms (Galveston, TX). During four-stage countercurrent fermentation, the resulting carboxylic acids consisted of primarily acetate (average 87.7 wt.%) and butyrate (average 9.0 wt.%). The experimental fermentation trains had the highest yield (0.47 g total acids/g volatile solids (VS) fed) and highest selectivity (0.79 g total acids/g VS digested) at a total acid concentration of 28.3 g/L, which is equivalent to an ethanol yield of 105.2 gal/(tonne VS fed). Both high total acid concentrations (>44.7 g/L) and high substrate conversions (>77.5%) are predicted for countercurrent fermentations of bagasse at commercial scale, allowing for an efficient conversion of air-lime-treated biomass to liquid transportation fuels and chemicals via the carboxylate platform.     

Ammonium carboxylate production from sugarcane trash using long-term air-lime pretreatment followed by mixed-culture fermentation

Sugarcane trash (ST) was converted to ammonium carboxylates using a novel bioprocessing strategy known as long-term air-lime pretreatment/mixed-culture fermentation. At mild conditions (50 °C, 5 weeks, 1-atm air, and excess lime loading of 0.4 g Ca(OH)₂/(g dry biomass)), air-lime pretreatment of ST had moderate delignification (64.4%) with little loss in polysaccharides. Without employing detoxification, sterility, expensive nutrients, or costly enzymes, the feedstock (80% treated ST/20% chicken manure) was fermented to primarily ammonium acetate (>75%) and butyrate by a mixed culture of marine microorganisms at 55 °C. In the best four-stage countercurrent fermentation, the product yield was 0.36 g total acids/(g VS fed) and the substrate conversion was 64%. Model predictions indicate both high acid concentrations (>47.5 g/L) and high substrate conversions (>70%) are possible at industrial scale.     

Fixed-bed fermentation of rice straw and chicken manure using a mixed culture of marine mesophilic microorganisms

A mixture of rice straw (80%) and chicken manure (20%) was pretreated and fermented to carboxylic acids by using a mixed culture of marine mesophilic microorganisms. Two sets of four fermentors, built from PVC pipes, were used for both biomass pretreatment and fermentation. Four 1L fermentors (F1-F4) were arranged in series, where liquid fermentation products were transferred from one fermentor to the other, to form a train. A liquid volume of 10mL and 15mL were transferred every four days for Trains A and B, respectively. The maximum total acid concentration for F1 in Train A was 34.2g/L and the maximum acid concentration in F2-F4 was approximately 44g/L. The maximum total acid concentration in F1 in Train B was 30.5g/L and the maximum acid concentration in F2-F4 was approximately 48g/L. The conversion in each of the fermentors in Train A varied from 0.821 to 0.879g VS digested/g VS fed and the yield was in the range 0.489-0.609g total acids/g VS fed. The conversion and yield in Train B were 0.741-0.914g VS digested/g VS fed and 0.563-0.669g total acids/g VS fed, respectively. The continuum particle distribution model (CPDM) predicted acid concentrations and retention times in the fixed-bed fermentation system with R(2) of 0.67-0.84 in Trains A and B.     

Measurement,control, and modeling of submerged acetic acid fermentation

For control and optimization of large scale bioprocesses, mathematical models are needed to describe transient growth and/or product formation. Such models can only be developed from reliable experimental data. A computerized experimental system was applied to submerged acetic acid fermentation with industrial Acetobacter strains in order to obtain quantitatively reproducible long-term data. Automated repeated-batch fermentations were carried out over a period of one year. It was found that consideration of substrate, product, and biomass concentrations alone was not sufficient to describe transient culture conditions. At least one more internal parameter must be taken into account. A delay-time model was developed which takes into consideration the variable concentration of an internal component of the cells, the ribonucleic acid. This model was used to simulate the acetic acid fermentation. The simulation results agreed well with the experimental data. Thus, the validity of the model assumptions could be confirmed. The model was capable of simulating the lag-phase of growth as well as lysis of microorganisms due to product inhibition.     

Ring-opening bulk polymerization of epsilon-caprolactone and trimethylene carbonate catalyzed by lipase Novozym 435.

The affects of lipase concentration on ring-opening bulk polymerizations of epsilon-caprolactone and trimethylene carbonate were studied by using Novozym 435 (immobilized form of lipase B from Candida antarctica) as biocatalyst. The polymerization of epsilon-caprolactone was carried out in bulk at 70 degrees C. Three lipase concentrations of 9.77, 1.80 and 0.50 mg/mmol epsilon-CL were used in the experiment. The results showed that increasing the lipase concentration used in the polymerization system resulted in an increased rate of monomer consumption. For an enzyme concentration of 9.8 mg lipase per mmol monomer, an 80% monomer conversion was achieved in a 4-h time period, while for the lower enzyme concentration of 1.8 mg lipase per mmol monomer, 48 h were needed to reach monomer conversion. Linear relationships between Mn and monomer conversions were observed in all three enzyme concentrations, suggesting that the product molecular weight may be controlled by the stoichiometry of the reactants for these systems. At the same monomer conversion level, however, Mn decreased with increasing enzyme concentration. After correcting for the amount of monomer consumed in initiation, the plot of ln[([M]o - [M]i)/([Mt] - [M]i)] versus reaction time was found to be linear, suggesting that the monomer consumption followed a first-order rate law and no chain termination occurred. For the TMC systems, the polymerization was carried out in bulk at 55 degrees C. Similar to the epsilon-CL systems, increasing the Novozym 435 concentration from 8.3 to 23.6 mg/mmol TMC increased the rate of monomer conversion. Unlike the epsilon-CL systems, however, nonlinear relationships were obtained between Mn and monomer conversion, indicating that possible chain transfer and/or slow initiation had taken place in these systems. Consistent with the above result, nonlinear behavior was observed for the plot of ln[[M]o/[M]t] versus reaction time.     

Simple, rapid method for converting a peptide from one salt form to another

A method for converting peptide trifluoroacetate salts to the corresponding acetate salts has been developed. The procedure involves reversed phase HPLC with a volatile buffer system. The method is exemplified by the conversion of Growth Hormone-Release Factor, GRF(1-44)-NH2 trifluoroacetate to the acetate which was achieved in greater than 95% recovery. Extensive analytical studies of the product confirmed the absence of trifluoroacetate and that the acetate salt was obtained without degradation. This procedure is expected to be generally applicable to other peptides and to other salt form conversions.     

An immobilized cell reactor with simultaneous product separation. II. Experimental reactor performance

The simultaneous separation of volatile fermentation products from product-inhibited fermentations can greatly increase the productivity of a bioreactor by reducing the product concentration in the bioreactor, as well as concentrating the product in an output stream free of cells, substrate, or other feed impurities. The Immobilized Cell Reactor-Separator (ICRS) consists of two column reactors: a cocurrent gas-liquid "enricher" followed by a countercurrent "stripper" The columns are four-phase tubular reactors consisting of (1) an inert gas phase, (2) the liquid fermentation broth, (3) the solid column internal packing, and (4) the immobilized biological catalyst or cells. The application of the ICRS to the ethanol-from-whey-lactose fermentation system has been investigated. Operation in the liquid continuous or bubble flow regime allows a high liquid holdup in the reactor and consequent long and controllable liquid residence time but results in a high gas phase pressure drop over the length of the reactor and low gas flow rates. Operation in the gas continuous regime gives high gas flow rates and low pressure drop but also results in short liquid residence time and incomplete column wetting at low liquid loading rates using conventional gas-liquid column packings. Using cells absorbed to conventional ceramic column packing (0.25-in. Intalox saddles), it was found that a good reaction could be obtained in the liquid continuous mode, but little separation, while in the gas continuous mode there was little reaction but good separation. Using cells sorbed to an absorbant matrix allowed operation in the gas continuous regime with a liquid holdup of up to 30% of the total reactor volume. Good reaction rates and product separation were obtained using this matrix. High reaction rates were obtained due to high density cell loading in the reactor. A dry cell density of up to 92 g/L reactor was obtained in the enricher. The enricher ethanol productivity ranged from 50 to 160 g/L h while the stripper productivity varied from 0 to 32 g/L h at different feed rates and concentrations. A separation efficiency of as high as 98% was obtained from the system.     

Diffusion and reaction within porous packing media: a phenomenological model

A phenomenological model has been developed to describe biomass distribution and substrate depletion in porous diatomaceous earth (DE) pellets colonized by Pseudomonas aeruginosa. The essential features of the model are diffusion, attachment and detachment to/from pore walls of the biomass, diffusion of substrate within the pellet, and external mass transfer of both substrate and biomass in the bulk fluid of a packed bed containing the pellets. A bench-scale reactor filled with DE pellets was inoculated with P. aeruginosa and operated in plug flow without recycle using a feed containing glucose as the limiting nutrient. Steady-state effluent glucose concentrations were measured at various residence times, and biomass distribution within the pellet was measured at the lowest residence time. In the model, microorganism/substrate kinetics and mass transfer characteristics were predicted from the literature. Only the attachment and detachment parameters were treated as unknowns, and were determined by fitting biomass distribution data within the pellets to the mathematical model. The rate-limiting step in substrate conversion was determined to be internal mass transfer resistance; external mass transfer resistance and microbial kinetic limitations were found to be nearly negligible. Only the outer 5% of the pellets contributed to substrate conversion. (c) 1993 Wiley & Sons, Inc.     

Continuous gluconic acid production by isolated yeast-like mould strains of <Emphasis Type="Italic">Aureobasidium pullulans</Emphasis>

By extensive microbial screening, about 50 strains with the ability to secrete gluconic acid were isolated from wild flowers. The strains belong to the yeast-like mould Aureobasidium pullulans (de Bary) Arnaud. In shake flask experiments, gluconic acid concentrations between 23 and 140 g/l were produced within 2 days using a mineral medium. In batch experiments, various important fermentation parameters influencing gluconic acid production by A. pullulans isolate 70 (DSM 7085) were identified. Continuous production of gluconic acid with free-growing cells of the isolated yeast-like microorganisms was studied. About 260 g/l gluconic acid at total glucose conversion could be achieved using continuous stirred tank reactors in defined media with residence times (RT) of about 26 h. The highest space-time-yield of 19.3 g l(-1) x h(-1)) with a gluconic acid concentration of 207.5 g/l was achieved with a RT of 10.8 h. The possibility of gluconic acid production with biomass retention by immobilised cells on porous sinter glass is discussed. The new continuous gluconate fermentation process provides significant advantages over traditional discontinuous operation employing Aspergillus niger. The aim of this work was the development of a continuous fermentation process for the production of gluconic acid. Process control becomes easier, offering constant product quality and quantity.     

Bioconversion of wheat straw to ethanol: Chemical modification,enzymatic hydrolysis,and fermentation

Native wheat straw (WS) was pretreated with various concentrations of H₂SO₄ and NaOH followed by secondary treatments with ethylene diamine (EDA) and NH₄OH prior to enzymatic saccharification. Conversion of the cellulosic component to sugar varied with the chemical modification steps. Treatment solely with alkali yield 51–75% conversion, depending on temperature. Acid treatment at elevated tempeatures showed a substantial decrease in the hemicellulose component, whereas EDA-treated WS (acid pretreated) showed a 69–75% decrease in the lignin component. Acid-pretreated EDA-treated straw yielded a 98% conversion rate, followed by 83% for alkali–NH₄OH treated straws. In other experiments, WS was pretreated with varying concentration of H₂SO₄ or NaOh followed by NH₄OH treatment prior to enzymatic hydrolysis. Pretreatment of straw with 2% NaOH for 4 h coupled to enzymatic hydrolysis yield a 76% conversion of the cellulosic component. Acid–base combination pretreatment yielded only 43% conversions. A reactor column was subsequently used to measure modification–saccharification–fermentation for wheat straw conversion on a larger scale. Thirty percent conversions of wheat straw cellulosics to sugar were observed with subsequent fermentation to alcohol. The crude cellulase preparation yielded considerable quantities of xylose in addition to the glucose. Saccharified materials were fermented directly with actively proliferating proliferating yeast cells without concentration of the sugars.     

Environmental analysis of producing biochar and energy recovery from pulp and paper mill biosludge

Sweden is one of the largest exporters of pulp and paper products in the world. It follows that huge quantities of sludge rich in carbonaceous organic material and containing heavy metals are generated. This paper carried out a comparative environmental analysis of three different technologies, which can be adopted to produce biochar and recover energy from the biosludge, using landfilling as the reference case. These three thermochemical biosludge management systems—using incineration, pyrolysis, and hydrothermal carbonization (HTC)—were modeled using life cycle assessment (LCA). Heat generated in the incineration process (System A) was considered to be for captive consumption within the kraft pulp mills. It was assumed that the biochars—pyrochar and hydrochar—produced from pyrolysis (System B) and HTC (System C), respectively, were added to the forest soils. The LCA results show that all the alternative systems considerably improve the environmental performance of biosludge management, relative to landfilling. For all systems, there are net reductions in greenhouse gas emissions (–0.89, –1.43, and –1.13 tonnes CO₂‐equivalent per tonne dry matter biosludge in Systems A, B, and C, respectively). System B resulted in the lowest potential eutrophication and terrestrial ecotoxicity impacts, whereas System C had the least acidification potential. The results of this analysis show that, from an environmental point of view, biochar soil amendment as an alternative method for handling pulp and paper mill biosludge is preferable to energy recovery. However, an optimal biochar system needs to factor in the social and economic contexts as well.     

The effects of potassium and chloride ions on the ethanolic fermentation of sucrose by Zymomonas mobilis 2716

Summary The inclusion of specific salts in Zymomonas mobilis batch sucrose fermentations can limit by-product formation. Sorbitol and fructo-oligosaccharide formation can be reduced and ethanol production enhanced by manipulating mineral salt concentrations. Chloride salts reduced the production of biomass and sorbitol in favour of fructo-oligosaccharide formation at concentrations lower than 10 g NaCl/l or MgCl₂. Higher concentrations led to the accumulation of glucose and fructose. Low concentrations of KH₂PO₄ (<20 g/l) enhanced biomass formation, and the concomitant reduction in sorbitol and fructo-oligosaccharides favoured enhanced ethanol formation. At concentrations above 20 g/l, its effects were similar to those obtained with the chloride salts. Invertase addition at the start of fermentation increased sorbitol formation, whereas addition after the completion of sucrose hydrolysis resulted in the conversion of fructo-oligosaccharides formed into fructose or ethanol. Fermentation with 250 g/l of sugar-cane syrup ( = 130 g sucrose/l) in the presence of 8 g KH₂PO₄/l, with 0.05 g invertase/l added on the completion of sucrose hydrolysis, resulted in a conversion efficiency of 94% with complete carbon accountability, and only 7 g sorbitol/l. Offprint requests to: H. W. Doelle     

Ethanol production from xylose by Thermoanaerobacter ethanolicus in batch and continuous culture

The fermentation of xylose by Thermoanaerobacter ethanolicus ATCC 31938 was studied in pH-controlled batch and continuous cultures. In batch culture, a dependency of growth rate, product yield, and product distribution upon xylose concentration was observed. With 27 mM xylose media, an ethanol yield of 1.3 mol ethanol/mol xylose (78% of maximum theoretical yield) was typically obtained. With the same media, xylose-limited growth in continuous culture could be achieved with a volumetric productivity of 0.50 g ethanol/liter h and a yield of 0.42 g ethanol/g xylose (1.37 mol ethanol/mol xylose). With extended operation of the chemostat, variation in xylose uptake and a decline in ethanol yield was seen. Instability with respect to fermentation performance was attributed to a selection for mutant populations with different metabolic characteristics. Ethanol production in these T. ethanolicus systems was compared with xylose-to-ethanol conversions of other organisms. Relative to the other systems, T. ethanolicus offers the advantages of a high ethanol yield at low xylose concentrations in batch culture and of a rapid growth rate. Its disadvantages include a lower ethanol yield at higher xylose concentrations in batch culture and an instability of fermentation characteristics in continuous culture.     

多级逆流工艺促进城市污泥厌氧发酵生产挥发性脂肪酸

采用一种新型的厌氧发酵工艺——多级逆流发酵工艺对城市污泥进行厌氧发酵, 实现高效产挥发性脂肪酸的目的。结果表明, 实验条件下应用多级逆流发酵工艺, 挥发性脂肪酸浓度与产率分别达到(10.5±0.5) g/L和0.20 gVFAs/ gVS, 与普通厌氧发酵工艺相比, 分别提高了31%和54%。此外, 在多级逆流工艺中, 底物有机质去除率可达50%, 较普通厌氧发酵提高了37%。进一步分析多级逆流工艺产酸的机制, 发现产酸效率的提高在于降低了发酵产物对厌氧产酸细菌的抑制效应, 并且工艺的VFAs产率以及有机质去除率分别取决于第一级和第三级厌氧发酵过程。因此, 城市污泥采用多级逆流工艺厌氧发酵不仅能够有效促进挥发性脂肪酸的生成, 而且能够较大程度上提高污泥中有机质的去除率。     

Continuous SSCF of AFEX™ pretreated corn stover for enhanced ethanol productivity using commercial enzymes and Saccharomyces cerevisiae 424A (LNH‐ST)

High productivity processes are critical for commercial production of cellulosic ethanol. One high productivity process—continuous hydrolysis and fermentation—has been applied in corn ethanol industry. However, little research related to this process has been conducted on cellulosic ethanol production. Here, we report and compare the kinetics of both batch SHF (separate hydrolysis and co‐fermentation) and SSCF (simultaneous saccharification and co‐fermentation) of AFEX? (Ammonia Fiber Expansion) pretreated corn stover (AFEX?‐CS). Subsequently, we designed a SSCF process to evaluate continuous hydrolysis and fermentation performance on AFEX?‐CS in a series of continuous stirred tank reactors (CSTRs). Based on similar sugar to ethanol conversions (around 80% glucose‐to‐ethanol conversion and 47% xylose‐to‐ethanol conversion), the overall process ethanol productivity for continuous SSCF was 2.3‐ and 1.8‐fold higher than batch SHF and SSCF, respectively. Slow xylose fermentation and high concentrations of xylose oligomers were the major factors limiting further enhancement of productivity. Biotechnol. Bioeng. 2013; 110: 1302–1311. © 2012 Wiley Periodicals, Inc.