Engineering the Metabolic Landscape of Microorganisms for Lignocellulosic Conversion.

Bacteria and yeast are being intensively used to produce biofuels and high-added-value products by using plant biomass derivatives as substrates. The number of microorganisms available for industrial processes is increasing thanks to biotechnological improvements to enhance their productivity and yield through microbial metabolic engineering and laboratory evolution. This is allowing the traditional industrial processes for biofuel production, which included multiple steps, to be improved through the consolidation of single-step processes, reducing the time of the global process, and increasing the yield and operational conditions in terms of the desired products. Engineered microorganisms are now capable of using feedstocks that they were unable to process before their modification, opening broader possibilities for establishing new markets in places where biomass is available. This review discusses metabolic engineering approaches that have been used to improve the microbial processing of biomass to convert the plant feedstock into fuels. Metabolically engineered microorganisms (MEMs) such as bacteria, yeasts, and microalgae are described, highlighting their performance and the biotechnological tools that were used to modify them. Finally, some examples of patents related to the MEMs are mentioned in order to contextualize their current industrial use.

Enhancing Phosphorus and Nitrogen Uptake in Maize Crops with Food Industry Biosolids and Azotobacter nigricans.

The problem of phosphorus and nitrogen deficiency in agricultural soils has been solved by adding chemical fertilizers. However, their excessive use and their accumulation have only contributed to environmental contamination. Given the high content of nutrients in biosolids collected from a food industry waste treatment plant, their use as fertilizers was investigated in Zea mays plants grown in sandy loam soil collected from a semi-desert area. These biosolids contained insoluble phosphorus sources; therefore, given the ability of Azotobacter nigricans to solubilize phosphates, this strain was incorporated into the study. In vitro, the suitable conditions for the growth of Z. mays plants were determined by using biosolids as a fertilizer and A. nigricans as a plant-growth-promoting microorganism; in vitro, the ability of A. nigricans to solubilize phosphates, fix nitrogen, and produce indole acetic acid, a phytohormone that promotes root formation, was also evaluated. At the greenhouse stage, the Z. mays plants fertilized with biosolids at concentrations of 15 and 20% (v/w) and inoculated with A. nigricans favored the development of bending strength plants, which was observed on the increased stem diameter (>13.5% compared with the negative control and >7.4% compared with the positive control), as well as a better absorption of phosphorus and nitrogen, the concentration of which increased up to 62.8% when compared with that in the control treatments. The interactions between plants and A. nigricans were observed via scanning electron microscopy. The application of biosolids and A. nigricans in Z. mays plants grown in greenhouses presented better development than when Z. mays plants were treated with a chemical fertilizer. The enhanced plant growth was attributed to the increase in root surface area.

Bioconversion of H2 and CO2 from dark fermentation to methane: Effect of operating conditions on methane concentration.

The main goal of this study was to assess the methane production in a biotrickling filter (BTF) using a synthetic gas mixture (H2/CO2: 60/40), evaluating the effect of the empty bed gas residence time (EBRT), pH, and temperature. The BTF was inoculated with acclimated granular anaerobic sludge. Three EBRT were tested: 11.6, 5.8, and 2.9 h. The decrease in EBRT (from 11.6 to 5.8 h) increased 1.3-fold the methane content (69 ± 3%) with H2 and CO2 removals of 100% and 24 ± 6%, respectively. The following reduction to 2.9 h showed no effect on CH4 content. The increment of the pH had no significant effect; however, the highest CH4 percentage (74%) was observed at a pH of 8.5. The system showed flexibility to adapt to changes in temperature without drastically diminishing CH4 concentration. In these stages, the principal hydrogenotrophic archaea detected was Methanobacterium flexile. Soluble microbial products such as butanol, caproate, and iso-valerate were detected in all the operating stages. This study demonstrates the potential of methane generation from a dark fermentation gaseous effluent.

The influence of total solids content and initial pH on batch biohydrogen production by solid substrate fermentation of agroindustrial wastes.

Hydrogen is a valuable clean energy source, and its production by biological processes is attractive and environmentally sound and friendly. In México 5 million tons/yr of agroindustrial wastes are generated; these residues are rich in fermentable organic matter that can be used for hydrogen production. On the other hand, batch, intermittently vented, solid substrate fermentation of organic waste has attracted interest in the last 10 years. Thus the objective of our work was to determine the effect of initial total solids content and initial pH on H2 production in batch fermentation of a substrate that consisted of a mixture of sugarcane bagasse, pineapple peelings, and waste activated sludge. The experiment was a response surface based on 2(2) factorial with central and axial points with initial TS (15-35%) and initial pH (6.5-7.5) as factors. Fermentation was carried out at 35 °C, with intermittent venting of minireactors and periodic flushing with inert N2 gas. Up to 5 cycles of H2 production were observed; the best treatment in our work showed cumulative H2 productions (ca. 3 mmol H2/gds) with 18% and 6.65 initial TS and pH, respectively. There was a significant effect of TS on production of hydrogen, the latter decreased with initial TS increase from 18% onwards. Cumulative H2 productions achieved in this work were higher than those reported for organic fraction of municipal solid waste (OFMSW) and mixtures of OFMSW and fruit peels waste from fruit juice industry, using the same process. Specific energetic potential due to H2 in our work was attractive and fell in the high side of the range of reported results in the open literature. Batch dark fermentation of agrowastes as practiced in our work could be useful for future biorefineries that generate biohydrogen as a first step and could influence the management of this type of agricultural wastes in México and other countries and regions as well.

Re-fermentation of washed spent solids from batch hydrogenogenic fermentation for additional production of biohydrogen from the organic fraction of municipal solid waste.

In the first batch solid substrate anaerobic hydrogenogenic fermentation with intermittent venting (SSAHF-IV) of the organic fraction of municipal solid waste (OFMSW), a cumulative production of 16.6 mmol H(2)/reactor was obtained. Releases of hydrogen partial pressure first by intermittent venting and afterward by flushing headspace of reactors with inert gas N(2) allowed for further hydrogen production in a second to fourth incubation cycle, with no new inoculum nor substrate nor inhibitor added. After the fourth cycle, no more H(2) could be harvested. Interestingly, accumulated hydrogen in 4 cycles was 100% higher than that produced in the first cycle alone. At the end of incubation, partial pressure of H(2) was near zero whereas high concentrations of organic acids and solvents remained in the spent solids. So, since approximate mass balances indicated that there was still a moderate amount of biodegradable matter in the spent solids we hypothesized that the organic metabolites imposed some kind of inhibition on further fermentation of digestates. Spent solids were washed to eliminate organic metabolites and they were used in a second SSAHF-IV. Two more cycles of H(2) production were obtained, with a cumulative production of ca. 2.4 mmol H(2)/mini-reactor. As a conclusion, washing of spent solids of a previous SSAHF-IV allowed for an increase of hydrogen production by 15% in a second run of SSAHF-IV, leading to the validation of our hypothesis.

Improvement of biohydrogen production from solid wastes by intermittent venting and gas flushing of batch reactors headspace.

Headspace of batch minireactors was intermittently vented and gas flushed with N2 in order to enhance H2 production (PH) by anaerobic consortia degrading organic solid wastes. Type of inocula (meso and thermophilic), induction treatment (heat-shock pretreatment, HSP, and acetylene, Ac), and incubation temperature (37 and 55 degrees C) were studied by means of a factorial design. On average, it was found that mesophilic incubation had the most significant positive effect on PH followed by treatment with Ac, although the units with the best performance (high values of PH, initial hydrogen production rate, and short lag time) were those HSP-induced units incubated at 37 degrees C (type of inocula was not significant). In this way, after 720 h of incubation PH was inhibited in those units by H2 partial pressure (pH2) of 0.54 atm. Venting and gas flushing with N2 was efficient to eliminate that inhibition achieving additional hydrogen generation in subsequent incubation cycles although smaller than the first one. Thus, four cycles of PH were obtained from the same substrate with neither addition of inocula nor application of induction treatment obtaining an increment of 100% in the generated H2. In those subsequent cycles there was a positive correlation between PH and organic acids/solvent ratio; maximum values were found in the first cycle. Solventogenesis could be clearly distinguished in third and fourth production cycles, probably due to a metabolic shift originated by high organic acid concentrations.