Influence of pH and temperature on the performance and microbial community during the production of medium-chain carboxylic acids using winery effluents as substrate.

Winery effluents containing high ethanol concentrations and diverse organic matter are ideal substrates for producing medium-chain carboxylic acids via fermentation and chain elongation. However, the process needs to be better understood. This study presents novel insights into the bioconversion mechanisms of medium-chain carboxylic acids by correlating fermentation and chain elongation kinetic profiles with the study of microbial communities at different pH (5 to 7) conditions and temperatures (30 to 40 °C). It was found that high productivities of MCCA were obtained using a native culture and winery effluents as a natural substrate. Minor pH variations significantly affected the metabolic pathway of the microorganisms for MCCA production. The maximal productivities of hexanoic (715 mg/L/d) and octanoic (350 mg/L/d) acids were found at pH 6 and 35 °C. Results evidence that the presence of Clostridium, Bacteroides, and Negativicutes promotes the high productions of MCCA. The formation of heptanoic acid was favor when Mogibacterium and Burkholderia were present.

Feedstock design for quality biomaterials.

Feedstock design is crucial for lignocellulosic biomass use. Current strategies for feedstock design cannot be readily applied to improve the quality of biomass-based materials, limiting the sustainability and economics of lignocellulosic biorefineries. Recent studies have advanced the understanding of biomass structure-property relationships and discovered several characteristics, such as molecular weight, uniformity, linkage profile, and functional groups, that are critical for manufacturing diverse quality biomaterials. These discoveries call for fundamentally different strategies for feedstock development. Such strategies need to rediscover the roles of monolignol biosynthesis enzymes and leverage lignin polymerization enzymes to achieve precise control of lignin molecular structure. These innovations could transform biomass into feedstock for high-quality biomaterials, addressing essential environmental challenges and empowering the bioeconomy.

New insights into microbial interactions and putative competitive mechanisms during the hydrogen production from tequila vinasses.

This study aimed to characterize the prokaryotic community and putative microbial interactions involved in hydrogen (H2) production during the dark fermentation (DF) process, applying principal components analysis (PCA) to correlate changes in operational, physicochemical, and biological variables. For this purpose, a continuous stirred-tank reactor-type digester fed with tequila vinasses was operated at 24, 18, and 12 h of hydraulic retention times (HRTs) to apply organic loading rates of 20, 36, and 54 g-COD L-1 d-1, corresponding to stages I, II, and III, respectively. Results indicated high population dynamics for Archaea during the DF process toward a decrease in total sequences from 6299 to 99. Concerning the Bacteria community, lactic acid bacteria (LAB) were dominant reaching a relative abundance of 57.67%, while dominant H2-producing bacteria (HPB) decreased from 25.76% to 21.06% during stage III. Putative competitive exclusion mechanisms such as competition for substrates, bacteriocins production, and micronutrient depletion carried out by Archaea and non-H2-producing bacteria (non-HPB), especially LAB, could negatively impact the dominance of HPB such as Ethanoligenens harbinense and Clostridium tyrobutyricum. As a consequence, low maximal volumetric H2 production rate (672 mL-H2 L-1 d-1) and yield (3.88 mol-H2 assimilated sugars-1) were obtained. The global scenario obtained by PCA correlations suggested that C. tyrobutyricum positively impacted H2 molar yield through butyrate fermentation using the butyryl-CoA:acetate CoA transferase pathway, while the most abundant HPB E. harbinense decreased its relative abundance at the shortest HRT toward the dominance of non-HPB. This study provides new insights into the microbial interactions and helps to better understand the DF performance for H2 production using tequila vinasses as substrate. KEY POINTS: • E. harbinense and C. tyrobutyricum were responsible for H2 production. • Clostridiales used acetate and butyrate fermentations for H2 production. • LAB won the competition for sugars against Clostridiales during DF. • Putative bacteriocins production and micronutrients depletion could favor LAB.

Prokaryotic population dynamics and interactions in an AnSBBR using tequila vinasses as substrate in co-digestion with acid hydrolysates of Agave tequilana var. azul bagasse for hydrogen production.

AIMS: The purpose of this study was to characterize the prokaryotic community and putative microbial interactions between H2 -producing bacteria (HPB) and non-HPB using two anaerobic sequencing batch biofilm reactors (AnSBBRs) fed with tequila vinasses in co-digestion with acid hydrolysates of Agave tequilana var. azul bagasse (ATAB). METHODS AND RESULTS: Two AnSBBRs were operated for H2 production to correlate changes in physicochemical and biological variables by principal component analysis (PCA). Results indicated that H2 yield was supported by Ethanoligenens harbinense and Clostridium tyrobutyricum through the PFOR pathway. However, only E. harbinense was able to compete for sugars against non-HPB. Competitive exclusion associated with competition for sugars, depletion of essential trace elements, bacteriocin production and resistance to inhibitory compounds could be carried out by non-HPB, increasing their relative abundances during the dark fermentation (DF) process. CONCLUSIONS: The global scenario obtained by PCA correlated the decrease in H2 production with the lactate:acetate molar ratio in the influent. At the beginning of co-digestion, this ratio had the minimum value considered for a net gain of ATP. This fact could cause the reduction of the relative abundance of C. tyrobutyricum. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study that demonstrated the feasibility of H2 production by Clostridiales from acid hydrolysates of ATAB in co-digestion with tequila vinasses.

Standardized protocol for determination of biohydrogen potential.

Biohydrogen production potential (BHP) depends on several factors like inoculum source, substrate, pH, among many others. Batch assays are the most common strategy to evaluate such parameters, where the comparison is a challenging task due to the different procedures used. The present method introduces the first internationally validated protocol, evaluated by 8 independent laboratories from 5 different countries, to assess the biohydrogen potential. As quality criteria, a coefficient of variation of the cumulative hydrogen production (H max) was defined to be <15 %. Two options to run BHP batch tests were proposed; a manual protocol with periodic measurements of biogas production, needing conventional laboratory materials and analytical equipment for biogas characterization; and an automatic protocol, which is run in a device developed for online measurements of low biogas production. The detailed procedures for both protocol options are presented, as well as data validating them. The validation showed acceptable repeatability and reproducibility, measured as intra- and inter-laboratory coefficient of variation, which can be reduced up to 9 %.

Bioderived ionic liquid-based pretreatment enhances methane production from Agave tequilana bagasse.

In recent years, bioderived ionic liquids have gained attention as a new promising approach for lignocellulosic biomass pretreatment. In this work, Agave tequilana bagasse (ATB), an attractive bioenergy feedstock in Mexico, was pretreated with a bioderived ionic liquid (cholinium lysinate) for the first time. Optimization of the pretreatment conditions, in-depth biomass characterization and methane generation via anaerobic digestion are the main contributions of this work. The results indicated optimized pretreatment conditions of 124 °C, 205 min and 20% solids loading by applying a central composite design. The optimized pretreated ATB was able to produce an elevated sugar yield of 51.4 g total sugars per g ATB due to their high delignification (45.4%) and changes in their chemical linkages although an increase in cellulose crystallinity was found (0.51 untreated vs. 0.62 pretreated). Finally, the mass balance showed that 38.2 kg glucose and 13.1 kg xylose were converted into 12.5 kg of methane per 100 kg of untreated ATB, representing 86% of the theoretical methane yield and evidencing the potential of this biorefinery scheme.

Enhancing biohydrogen production from Agave tequilana bagasse: Detoxified vs. Undetoxified acid hydrolysates.

The aim of this work was to compare the biohydrogen production potential of undetoxified and detoxified acid hydrolysates from A. tequilana bagasse. Detoxification was carried out with activated carbon at different concentrations and pH values. Results indicated that pH was not a significant variable, while the lowest evaluated concentration of activated carbon (1% p/v) significantly promoted the highest removal of acetic acid (89%) with minimal losses of fermentable sugars. Regarding dark fermentation experiments, central composite designs were used to optimize COD and pH variables for both substrates, undetoxified and detoxified hydrolysates (activated carbon 1% p/v and pH 0.6). At optimal conditions, the detoxified hydrolysate produced 33% more biohydrogen than the undetoxified one. Hydrogen molar yields were 1.71 and 1.23 mol H2/molsugar, respectively. This improvement was correlated to changes in metabolic byproducts, since the detoxified hydrolysate produced only acetic and butyric acids, while lactic acid was detected in the undetoxified hydrolysate.

Mild reaction conditions induce high sugar yields during the pretreatment of Agave tequilana bagasse with 1-ethyl-3-methylimidazolium acetate.

Sequential 2k factorial and central composite designs were used to optimize Agave tequilana bagasse (ATB) pretreatment by using 1-ethyl-3-methylimidazolium acetate ([Emim][OAc]). Reaction time, temperature and solids loading were the studied factors while sugar yield was the response variable. Results indicated that optimal conditions (119 °C, 142 min) using high solids loading (30%) were achieved at lower temperatures and reaction times than those previously reported in the literature. It was also revealed that solid recovery after pretreatment with [Emim][OAc] is a key factor. The increase in enzymatic digestibility of pretreated ATB was correlated to a decrease in crystallinity and lower lignin content as observed using microscopy techniques and weaken chemical bonds by Fourier transform infrared spectroscopy. Yields of glucose and xylose in the hydrolysate were 41.3, and 13.0 kg per 100 kg of untreated ATB, which are equivalent to glucan and xylan conversions of 75.9% and 82.9%, respectively.

Agave tequilana bagasse for methane production in batch and sequencing batch reactors: Acid catalyst effect, batch optimization and stability of the semi-continuous process.

Agave tequilana bagasse is the main solid waste of the tequila manufacturing and represents an environmental issue as well as a potential feedstock for biofuel production due to its lignocellulosic composition and abundance. In this contribution, this feedstock was subjected to pretreatments with HCl and H2SO4 for sugar recovery and methane was produced from the hydrolysates in batch and sequencing batch reactors (AnSBR). Sugar recovery was optimized by using central composite designs at different levels of temperature, acid concentration and hydrolysis time. Results showed that at optimal conditions, the HCl pretreatment induced higher sugar recoveries than the H2SO4 one, 0.39 vs. 0.26 g total sugars/g bagasse. Furthermore, the H2SO4 hydrolysate contained higher concentrations of potential inhibitory compounds (furans and acetic acid). Subsequent anaerobic batch assays demonstrated that the HCl hydrolysate is a more suitable substrate for methane production; a four-fold increase was found. A second optimization by using HCl as acid catalyst and methane production as the response variable demonstrated that softer hydrolysis conditions are required to optimize methane production as compared to sugar recovery (1.8% HCl, 119 °C and 103min vs. 1.9% HCl, 130 °C and 133min). This softer conditions were used to feed an AnSBR for 110 days and evaluate its stability at three different cycle times (5, 3 and 2 days). Results showed stable reactor performances at cycle times of 5 and 3 days, obtaining the highest methane yield and production at 3 days, 0.28 NL CH4/g-COD and 1.04 NL CH4/d respectively. Operation at shorter cycle times is not advised due to microbial imbalance.

CO2 Removal from Biogas by Cyanobacterium Leptolyngbya sp. CChF1 Isolated from the Lake Chapala, Mexico: Optimization of the Temperature and Light Intensity.

In the present study, the capacity of the cyanobacterium Leptolyngbya sp. CChF1 to remove CO2 from real and synthetic biogas was evaluated. The identification of the cyanobacterium, isolated from the lake Chapala, was carried out by means of morphological and molecular analyses, while its potential for CO2 removal from biogas streams was evaluated by kinetic experiments and optimized by a central composite design coupled to a response surface methodology. Results demonstrated that Leptolyngbya sp. CChF1 is able to remove CO2 and grow indistinctly in real or synthetic biogas streams, showing tolerance to high concentrations of CO2 and CH4, 25 and 75%, respectively. The characterization of the biomass composition at the end of the kinetic assays revealed that the main accumulated by-products under both biogas streams were lipids, followed by proteins and carbohydrates. Regarding the optimization experiments, light intensity and temperature were the studied variables, while synthetic biogas was the carbon source. Results showed that light intensity was significant for CO2 capture efficiency (p = 0.0290), while temperature was significant for biomass production (p = 0.0024). The predicted CO2 capture efficiency under optimal conditions (27.1 °C and 920 lx) was 93.48%. Overall, the results of the present study suggest that Leptolyngbya sp. CChF1 is a suitable candidate for biogas upgrading.

Methane production from acid hydrolysates of Agave tequilana bagasse: evaluation of hydrolysis conditions and methane yield.

Evaluation of diluted acid hydrolysis for sugar extraction from cooked and uncooked Agave tequilana bagasse and feasibility of using the hydrolysates as substrate for methane production, with and without nutrient addition, in anaerobic sequencing batch reactors (AnSBR) were studied. Results showed that the hydrolysis over the cooked bagasse was more effective for sugar extraction at the studied conditions. Total sugars concentration in the cooked and uncooked bagasse hydrolysates were 27.9 g/L and 18.7 g/L, respectively. However, 5-hydroxymethylfurfural was detected in the cooked bagasse hydrolysate, and therefore, the uncooked bagasse hydrolysate was selected as substrate for methane production. Interestingly, results showed that the AnSBR operated without nutrient addition obtained a constant methane production (0.26 L CH4/g COD), whereas the AnSBR operated with nutrient addition presented a gradual methane suppression. Molecular analyses suggested that methane suppression in the experiment with nutrient addition was due to a negative effect over the archaeal/bacterial ratio.