Chain Elongation Process Research Articles

Outstanding enhancement of caproate production with microwave pyrolyzed highly reductive biochar addition

The integration of biochar into microbial Chain Elongation (CE) proves to be an effective tool of producing high-value bio-based products. This study innovatively applied biochar fabricated under microwave irradiation with carbon fiber cloth assistance into CE system. Results highlighted that microwave biochar achieved maximal CE efficiency yielding 8 g COD/L, with 3-fold increase to the blank group devoid of any biochar. Microwave biochar also obtained the highest substrate utilization rate of 94 %, while conventional biochar group recorded 90 % and the blank group was of 74 %. Mechanistic insights revealed that the reductive surface properties facilitated CE performance, which is relevant to fostering dominant genera of Parabacteroides, Bacteroides, and Macellibacteroides. By metagenomics, microwave biochar up-regulated functional genes and enzymes involved in CE process including ethanol oxidation, the reverse β-oxidation pathway, and the fatty acid biosynthesis pathway. This study effectively facilitated caproate production by utilizing a new microwave biochar preparation strategy.

Comprehensive comparative study on n-caproate production by Clostridium kluyveri: batch vs. continuous operation modes

Recently, there has been notable interest in researching and industrially producing medium-chain carboxylic acids (MCCAs) like n-caproate and n-caprylate via chain elongation process. This study presents a comprehensive assessment of the behavior and MCCA production profiles of Clostridium kluyveri in batch and continuous modes, at different ethanol:acetate molar ratios (1.5:1, 3.5:1 and 5.5:1). The highest n-caproate concentration, 12.9 ± 0.67 g/L (92.9 ± 1.39 % MCCA selectivity), was achieved in batch mode at a 3.5:1 ratio. Interestingly, higher ratios favored batch mode selectivity over continuous mode when this was equal or higher to 3.5:1. Steady state operation yielded the highest n-caproate (9.5 ± 0.13 g/L) and n-caprylate (0.35 ± 0.020 g/L) concentrations at the 3.5:1 ratio. Increased ethanol:acetate ratios led to a higher excessive ethanol oxidation (EEO) in both operational modes, potentially limiting n-caproate production and selectivity, especially at the 5.5:1 ratio. Overall, this study reports the efficient MCCA production of both batch and continuous modes by C. kluyveri.

Nano zero valent iron stimulated acetaldehyde oxidation, electron transfer, and RBO pathway for enhanced medium-chain carboxylic acids production

Nano zero-valent iron (NZVI) has been shown to effectively enhance the chain elongation (CE) process, addressing the issue of limited yield of medium-chain carboxylic acids (MCCA) from organic wastewater. However, the specific impact of NZVI on the metabolism of CE bacteria (CEB) is not well understood. In this study, it was aimed to investigate the mechanism by which an optimal concentration of NZVI influences CE metabolism, particularly in relation to ethanol oxidation, electron transfer, and MCCA synthesis. This was achieved through single-factor influence experiments and metagenomic analysis. The results showed that the addition of 1 g/gVSS NZVI achieved the highest MCCA yield (n-caproic acid + n-octanoic acid) at 2.02 g COD/L, which was 4.9 times higher than the control. This improvement in MCCA production induced by NZVI was attributed to several factors. Firstly, NZVI facilitated the oxidation of acetaldehyde, leading to its reduced accumulation in the system (from 18.4 % to 5.8 %), due to the optimized chemical environment created by NZVI corrosion, including near-neutral pH and a more reductive oxidation–reduction potential (ORP). Additionally, the inherent conductivity property of NZVI and the additional Fe ions released during corrosion improved the electron transfer efficiency between CEB. Lastly, both the composition of microbial communities and the abundance of unique enzyme genes confirmed the selective stimulation of NZVI on the reverse β-oxidation (RBO) pathway. These findings provide valuable insights into the role of NZVI in CEB metabolism and its potential application for enhancing MCCA production in CE bioreactors.

Perspectives on carboxylates generation from Ecuadorian agro-wastes

Carboxylates generation from banana (peel and pulp), coffee, and cacao fermentation agro-waste, upon uncontrolled and controlled pHs of 6.6 (heat-driven methanogens inactivation) and 5.2 (pH inactivation), was studied. Regarding volatile fatty acids (VFAs), acetic was the highest for cocoa (96.2 g kg−1TVS) at pH 4.5. However, butyric was relevant for banana pulp (90.7 g kg−1TVS), at controlled pH 6.6. The highest medium chain fatty acid (MCFAs) level was hexanoic (cocoa, 3.5 g kg−1TVS), while octanoic reached a maximum of 2.8 g kg−1TVS for coffee at pH 6.6. At pH 5.2 MCFAs yield was relatively low. Uncontrolled pH conditions, using banana resulted in superior VFAs production compared to controlled conditions. Thus, pH became a determining variable when deciding the time and kind of carboxylic acid to be recovered. The bacterial community at the end of the chain elongation process was dominated by phyla Firmicutes, and Clostridium as the most common genera.

Medium-chain carboxylates production from plant waste: kinetic study and effect of an enriched microbiome

BackgroundThe need for addition of external electron donors such as ethanol or lactate impairs the economic viability of chain elongation (CE) processes for the production of medium-chain carboxylates (MCC). However, using feedstocks with inherent electron donors such as silages of waste biomass can improve the economics. Moreover, the use of an appropriate inoculum is critical to the overall efficiency of the CE process, as the production of a desired MCC can significantly be influenced by the presence or absence of specific microorganisms and their metabolic interactions. Beyond, it is necessary to generate data that can be used for reactor design, simulation and optimization of a given CE process. Such data can be obtained using appropriate mathematical models to predict the dynamics of the CE process.ResultsIn batch experiments using silages of sugar beet leaves, cassava leaves, and Elodea/wheat straw as substrates, caproate was the only MCC produced with maximum yields of 1.97, 3.48, and 0.88 g/kgVS, respectively. The MCC concentrations were accurately predicted with the modified Gompertz model. In a semi-continuous fermentation with ensiled sugar beet leaves as substrate and digestate from a biogas reactor as the sole inoculum, a prolonged lag phase of 7 days was observed for the production of MCC (C6–C8). The lag phase was significantly shortened by at least 4 days when an enriched inoculum was added to the system. With the enriched inoculum, an MCC yield of 93.67 g/kgVS and a productivity of 2.05 gMCC/L/d were achieved. Without the enriched inoculum, MCC yield and productivity were 43.30 g/kgVS and 0.95 gMCC/L/d, respectively. The higher MCC production was accompanied by higher relative abundances of Lachnospiraceae and Eubacteriaceae.ConclusionsEnsiled waste biomass is a suitable substrate for MCC production using CE. For an enhanced production of MCC from ensiled sugar beet leaves, the use of an enriched inoculum is recommended for a fast process start and high production performance.

Upgrading soybean dreg to caproate via intermediate of lactate and mediator of biochar

To address the environmental hazards posed by high-yield soybean dreg (SD), a high-value strategy is firstly proposed by synthesizing caproate through chain elongation (CE). Optimized conditions for lactate-rich broth as intermediate, utilizing 50 % inoculum ratio, 40 g/L substrate concentration, and pH 5, resulting in 2.05 g/L caproate from direct fermentation. Leveraging lactate-rich broth supplemented with ethanol, caproate was optimized to 2.76 g/L under a refined electron donor to acceptor of 2:1. Furthermore, incorporating 20 g/L biochar elevated caproate production to 3.05 g/L and significantly shortened the lag phase. Mechanistic insights revealed that biochar’s surface-existed quinone and hydroquinone groups exhibit potent redox characteristics, thereby facilitating electron transfer. Moreover, biochar up-regulated the abundance of key genes involved in CE process (especially fatty acids biosynthesis pathway), also enriching Lysinibacillus and Pseudomonas as an unrecognized cooperation to CE. This study paves a way for sustainable development of SD by upgrading to caproate.

Continuous chain elongation process for carbon resource recovery from excess sludge: Enhanced n-caprylate production and specific microbial functionalities

Thermal hydrolyzed sludge (THS) exhibits considerable promise in generating medium-chain fatty acids (MCFAs) through chain elongation (CE) technology. This study developed a novel continuous CE process using THS as the substrate, achieving an optimal ethanol loading rate (5.8 g COD/L/d) and stable MCFA production at 10.9 g COD/L, with a rate of 3.6 g COD/L/d. The MCFAs primarily comprised n-caproate and n-caprylate, representing 41.5 % and 54.3 % of the total MCFAs, respectively. Utilization efficiencies for ethanol and acetate were nearly complete at 100 % and 92.8 %, respectively. Key microbial taxa identified under these optimal conditions included Alcaligenes, SRB2, Sporanaerobacter, and Kurthia, which were instrumental in critical pathways such as the generation of acetyl-CoA, the initial carboxylation of acetyl-CoA, the fatty acid biosynthesis cycle, and energy metabolism. This research provides a theoretical and technical blueprint for converting waste sludge into valuable MCFAs, promoting sustainable waste-to-resource strategies.

Enhanced medium-chain fatty acid production from sewage sludge by combined electro-fermentation and anaerobic fermentation

Electro-fermentation (EF) was combined with anaerobic fermentation (AF) to promote medium-chain fatty acid (MCFA) from sewage sludge. Results showed that EF at acidification process significantly increased short-chain fatty acid (SCFA) production of by 0.5 times (82.4 mmol C/L). AF facilitated the chain elongation (CE) process by enhancing the SCFA conversion. Combined EF at acidification and AF at CE (EF-AF) achieved the highest MCFA production of 27.9 mmol C/L, which was 20 %-866 % higher than the other groups. Electrochemical analyses showed that enhanced SCFA and MCFA production was accompanied with good electrochemical performance at acidification and CE. Microbial analyses showed that EF-AF promoted MCFA production by enriching electrochemically active bacteria (EAB, Bacillus sp.). Enzyme analyses indicated that EF-AF promoted MCFA production by enriching the functional enzymes involved in Acetyl-CoA formation and the fatty acid biosynthesis (FAB) pathway. This study provided new insights into the production of MCFA from enhanced sewage sludge.

Enhancing chain-elongating microbiomes: Sequential fungi-bacteria cooperation for efficient food waste upgrading using endogenous electron donors

The upgrading of organic waste such as food waste (FW) into medium-chain carboxylates (MCC) is within a bio-based circular economy concept. An efficient chain elongation (CE) microbiome is difficult to obtain, which normally requires long-term acclimatization and an exogenous supplement of electron donor (ED, especially ethanol). In this study, the CE microbiome was rapidly shaped within 18 days and an efficient endogenous ethanol-based CE was achieved through the amendment of distiller yeast (DY) during FW fermentation. Multiple FW feeding accumulated the caproate concentration to 5.76 ± 0.33 g COD/L, which also regulated the CE process from ethanol-dependent to lactate-dependent metabolism. During the lactate-based CE process, the caproate production increased linearly (R2 = 0.82) with the rising D-lactate content. Ethanol-producing fungi yeast mainly consisted of Wickerhamomyces (66.54 %), Saccharomycopsis (5.37 %), Issatchenkia (2.21 %), unclassified_f_Metschnikowiaceae (1.73 %), and Saccharomyces (1.33 %). These yeasts were eliminated by FW feeding, which was in line with the alteration of ED preference from ethanol to lactate. Metagenome analysis indicated that the total relative abundance of lactate-based chain elongators including Ruminococcaceae bacterium CPB6, Clostridium luticellarii, Caproiciproducens galactitolivorans, and Megasphaera elsdenii was increased by 28.6 %, while the ethanol-based chain elongator Clostridium kluyveri kept stable after FW feeding. In addition, the functional genes related to the oxidation of lactate to acetyl-CoA were enriched after FW feeding, which further enhanced the subsequent CE pathway of acetyl-CoA-based reverse β oxidation rather than the fatty acids synthesis. This study provides a new insight into upgrading FW into valuable MCC without exogenous ED addition.

Influence of adding strategy of biochar on medium-chain fatty acids production from sewage sludge

Production of medium-chain fatty acids (MCFAs) from sewage sludge has dual effects on valuable sludge disposal and renewable energy generation, while low efficiency limits its application. Biochar addition is considered an effective method to improve MCFAs production. In this study, the influence of biochar adding strategies (i.e., adding biochar in acidification or chain elongation (CE) processes) on MCFAs production was explored. Results showed that by adding biochar in the acidification process, MCFAs accumulation increased by over 114%, accompanied by the highest carbon conversion efficiency (134.66%) and electron transfer efficiency of MCFAs (94.22%) by the terminal CE. Adding biochar before the acidification process better enriched CE bacteria (e.g., Paraclostridium) and strengthened the dominant metabolic pathway. In contrast, the biochar added before the CE process priorly enriched the bacteria capable of degrading organics, like unclassified_f__Dysgonomonadaceae, norank_f__norank_o__OPB41, and Acetobacterium. The differences in excessive ethanol oxidation and short-chain fatty acids accumulation induced by varied adding strategies might be responsible for this.

Enhancement of medium-chain fatty acids production from sludge anaerobic fermentation liquid under moderate sulfate reduction

In recent years, there has been a marked increase in the production of excess sludge. Chain-elongation (CE) fermentation presents a promising approach for carbon resource recovery from sludge, enabling the transformation of carbon into medium-chain fatty acids (MCFAs). However, the impact of sulfate, commonly presents in sludge, on the CE process remains largely unexplored. In this study, batch tests for CE process of sludge anaerobic fermentation liquid (SAFL) under different SCOD/SO42− ratios were performed. The moderate sulfate reduction under the optimum SCOD/SO42− of 20:1 enhanced the n-caproate production, giving the maximum n-caproate concentration, selectivity and production rate of 5.49 g COD/L, 21.4% and 4.87 g COD/L/d, respectively. The excessive sulfate reduction under SCOD/SO42− ≤ 5 completely inhibited the CE process, resulting in almost no n-caproate generation. The variations in n-caproate production under different conditions of SCOD/SO42− were all well fitted with the modified Gompertz kinetic model. Alcaligenes and Ruminococcaceae_UCG-014 were the dominant genus-level biomarkers under moderate sulfate reduction (SCOD/SO42− = 20), which enhanced the n-caproate production by increasing the generation of acetyl-CoA and the hydrolysis of difficult biodegradable substances in SAFL. The findings presented in this work elucidate a strategy and provide a theoretical framework for the further enhancement of MCFAs production from excess sludge.

Metagenomic analysis towards understanding the effects of ammonia on chain elongation process for medium chain fatty acids production

The production of medium chain fatty acids (MCFAs) through chain elongation (CE) from organic wastes/wastewater has attracted much attention, while the effects of a common inhibitor-ammonia has not been elucidated. The mechanism of ammonia affecting CE was studied by metagenomic. The lag phase duration of caproate production was increased, and the maximum caproate production rate was decreased by 43.4 % at 4 g-N/L, as compared to 0 g-N/L. And hydrochar (HC) alleviated the inhibition of ammonia at 4 g-N/L. Metagenomic analysis indicated that ammonia induced UBA4085 sp.FDU78 as the dominant microorganism, and metabolic reconstruction revealed its potential CE ability. Furthermore, ammonia inhibited the reverse β oxidation pathway and Acetyl-CoA production pathway. The tolerance of UBA4085 sp.FDU78 to ammonia was associated with the uptake of inorganic ions, energy conservation, and synthesis of osmoprotectants. The present study provided a deep-insight on the ammonia tolerance mechanism on the CE process.

Influence of acetate-to-butyrate ratio on carbon chain elongation in anaerobic fermentation

This study investigated the effect of electron acceptor (EA) distribution (acetate to butyrate ratio) on the carbon chain elongation (CCE) process. The results showed that the higher content of butyrate in the initial material led to the higher production of caproate. The maximum production of caproate was 3.74 ± 0.30 g·L−1, which was obtained when only butyrate was added as EA. Little caproate but much butyrate was produced where only acetate was added as EA. This indicated that CCE bacteria preferentially selected acetate as the EA to produce butyrate, and butyrate could be selected as EA to produce caproate only when the acetate content was much lower than butyrate. Unclassified_f_Dysgonomonadaceae, Massilibacterium, and Seramator were the predominant bacteria. Functional enzyme analysis showed that high butyrate content strengthened the fatty acid biosynthesis pathway and reverse β-oxidization pathway. The findings showed the importance of butyrate in CCE for caproate production.

Enhanced Food Waste Chain Elongation for Caproate Production: Role of Inorganic Carbon and Optimization Strategies

In this study, the role of inorganic carbon in chain elongation fermentation was investigated by examining its effects at different carbonate concentrations. The results demonstrated that higher yield of caproate (6.3±0.3 g/L - 7.4±0.2 g/L) was achieved with a NaHCO3 concentration of 2.5-5.0 g/L. Building upon these findings, a chain elongation process for food waste was developed. The caproate productions of control, CO2 and Na2CO3 trials were 4.6±0.2 g/L, 7.3±0.1 g/L, and 8.3±0.2 g/L, respectively. Chain elongation fermentation of food waste (1t TS) with Na2CO3 enhancement could synthesize 116.9 kg of caproate and 72.5 kg of carbon could be converted in caproate, accounting for 15.5% of the total organic carbon in the food waste. This study provided a comprehensive exploration of the chain elongation process for food waste, offering insights for further optimization and enhancement of food waste chain elongation performance coupling inorganic carbon utilization.

Regulation of hydraulic retention time on caproic acid production via two-phase anaerobic fermentation of Chinese cabbage waste with autopoietic electron donors

The effects of hydraulic retention time (HRT) on the performance of two-phase anaerobic fermentation for caproic acid production from Chinese cabbage waste (CCW) were investigated. In the electron donor phase, yeast was inoculated to achieve efficient autopoietic ethanol, providing electron donors for the chain elongation process. Shorter HRT led to drastic fluctuations in microorganisms, thus resulting in lower acid yields at HRT of 6 days. At HRT of 10 days, the balanced collaboration of various key bacteria avoided the accumulation of intermediate by-products, and the caproic acid production reached 4660 mg COD/L, which was 119.5% and 154.8% higher than that at HRTs of 6 and 14 days, respectively. At HRT of 14 days, the low ethanol loading rate resulted in ethanol excessive-oxidation to acetic acid. Acetic acid accounted for 41.5% of the total product, while the selectivity of caproic acid was only 15.3%. The main contributor to the production process of caproic acid was Caproiciproducens, while the Ruminalococcaceae also played a role in the process. This study provided a theoretical basis for the efficient production of caproic acid through continuous fermentation with autopoietic electron donors.

INVESTIGATING THE COMPARATIVE ANTIOXIDANT PARTITIONING IN FRIED OILS AND FRIED POTATOES: A SCIENTIFIC EVALUATION

Frying is a complex process that enhances the attractiveness and flavor of the final product. In this specific investigation, potato slices were subjected to frying using sun flower oil, and an antioxidant was introduced at a concentration of 200 ppm in accordance with FDA guidelines. Both the potato slices and the frying oil were analyzed to determine the levels of antioxidants after several frying cycles. The primary objective of this research was to evaluate the distribution of butylated hydroxyanisole (BHA), tertiary butyl hydroquinone (TBHQ), and propyl gallate (PG) in the fried potato slices and the frying oils. To conduct the experiments, batches of 50 grams of sliced potatoes were fried at 180ºC for 3-5 minutes per batch. The results showed that TBHQ demonstrated a superior ability to retain antioxidants compared to the other two types. TBHQ experienced significant degradation at frying temperatures, which likely contributes to the stability of the frying oil. The by-products generated during this degradation process impede the radical chain elongation process. As a result, the effectiveness of the antioxidants followed this order: TBHQ > BHA > PG.

Promotion of medium-chain fatty acids and long-chain alcohols production by zero-valent iron: Effect of particle sizes

The effect of zero-valent iron (ZVI) particle size on biological chain elongation (CE) was explored. The addition of ZVI particles with diameters of 50 nm-1000 µm stimulated both medium-chain fatty acids (MCFAs) and long-chain alcohols (LCAs) production, and increased carbon conversion rate of CE process by 2.8–12.2 times. ZVI particle size of 75 µm exhibited the best performance in promoting MCFAs production, achieved the highest MCFAs production of 2164.2 mg/L, which was 11.9 times of control group. ZVI particle size of 1000 µm showed the best performance in stimulating LCAs production, obtained the highest LCAs production of 434.0 mg/L, which was 33.4 times of the control group. Microbial community analysis suggested that different ZVI particle sizes significantly affected genus Clostridium_sensu_stricto_12 and Clostridium_sensu_stricto_7. Functional enzyme analysis revealed that 75 µm ZVI particles induced the strengthened reverse β oxidation pathway, while 1000 µm ZVI particles enhanced the fatty acid biosynthesis pathway.

Peracetic acid (PAA)-based pretreatment effectively improves medium-chain fatty acids (MCFAs) production from sewage sludge

Peracetic acid (PAA), known for its environmentally friendly properties as a oxidant and bactericide, is gaining prominence in decontamination and disinfection applications. The primary product of PAA oxidation is acetate that can serve as an electron acceptor (EA) for the biosynthesis of medium-chain fatty acids (MCFAs) via chain elongation (CE) reactions. Hence, PAA-based pretreatment is supposed to be beneficial for MCFAs production from anaerobic sludge fermentation, as it could enhance organic matter availability, suppress competing microorganisms and furnish EA by providing acetate. However, such a hypothesis has rarely been proved. Here we reveal that PAA-based pretreatment leads to significant exfoliation of extracellular polymeric substances (EPS) from sludge flocs and disruption of proteinic secondary structures, through inducing highly active free radicals and singlet oxygen. The production of MCFAs increases substantially to 11,265.6 mg COD L−1, while the undesired byproducts, specifically long-chain alcohols (LCAs), decrease to 723.5 mg COD L−1. Microbial activity tests further demonstrate that PAA pretreatment stimulates the CE process, attributed to the up-regulation of functional genes involved in fatty acid biosynthesis pathway. These comprehensive findings provide insights into the effectiveness and mechanisms behind enhanced MCFAs production through PAA-based technology, advancing our understanding of sustainable resource recovery from sewage sludge.

Simultaneous medium chain carboxylic acids and 1,3-propanediol production in a bioaugmented lactate-based chain elongation induced with glycerol

The objective was to investigate the impact of the bioaugmentation on chain elongation process using glycerol, lactate and lactose as substrates in an open culture fermentation. In the batch trials the highest selectivity for chain elongation product, i.e. caproate, was observed in trials inoculated with co-culture of Megasphaera elsdenii and Eubacterium limosum grown on glycerol (28.6%), and in non-bioaugmented open culture run on lactose + lactate (14.8%). The results showed that E. limosum, out of two bioaugmented strains, was able to survive in the open culture. A continuous open culture fermentation of glycerol led to caproate and 1,3-propanediol (1,3-PDO) formation, while lactate addition led to 1,3-PDO and short chain carboxylates production. Moving the process into batch mode triggered even-carbon chain elongation. Presence of E. limosum promoted odd-carbon chain elongation and valerate production. Imaging flow cytometry combined with machine learning enabled the discrimination of Eubacterium cells from other microbial strains during the process.

Permanganate (PM) pretreatment improves medium-chain fatty acids production from sewage sludge: The role of PM oxidation and in-situ formed manganese dioxide

Medium-chain fatty acids (MCFAs) production from sewage sludge is mainly restricted by the complex substrate structure, competitive metabolism and low electron transfer rate. This study proposes a novel permanganate (PM)-based strategy to promote sludge degradation and MCFAs production. Results show that PM pretreatment significantly increases MCFAs production, i.e., attaining 12,036 mg COD/L, and decreases the carbon fluxes of electron acceptor (EA)/electron donor (ED) to byproducts. Further analysis reveals that PM oxidation enhances the release and biochemical conversion of organic components via disrupting extracellular polymers (EPS) structure and reducing viable cells ratio, providing directly available EA for chain elongation (CE). The microbial activity positively correlated with MCFAs generation are apparently heightened, while the competitive metabolism of CE (i.e., methanogensis) can be completely inhibited. Accordingly, the functional bacteria related to critical bio-steps and dissimilatory manganese reduction are largely enriched. Further mechanism exploration indicates that the main contributors for sludge solubilization are 1O2 (61.6 %) and reactive manganese species (RMnS), i.e., Mn(V)/Mn(VI) (22.3 %) and Mn(III) (∼16.1 %). As the main reducing product of PM reaction, manganese dioxide (MnO2) can enable the formation of microbial aggregates, and serve as electron shuttles to facilitate the carbon fluxes to MCFAs during CE process. Overall, this strategy can achieve simultaneous hydrogen recovery, weaken competitive metabolisms and provide electron transfer accelerator for CE reactions.