Abstract: Biological routes for utilizing both carbohydrates and lignin are important to reach the ultimate goal of bioconversion of full carbon in biomass into biofuels and biochemicals. Recent biotechnology advances have shown promises toward facilitating biological transformation of lignin into lipids. Natural and engineered Rhodococcus strains (e.g., R. opacus PD630, R. jostii RHA1, and R. jostii RHA1 VanA−) have been demonstrated to utilize lignin for lipid production, and co-culture of them can promote lipid production from lignin. In this study, a co-fermentation module of natural and engineered Rhodococcus strains with significant improved lignin degradation and/or lipid biosynthesis capacities was established, which enabled simultaneous conversion of glucose, lignin, and its derivatives into lipids. Although Rhodococci sp. showed preference to glucose over lignin, nearly half of the lignin was quickly depolymerized to monomers by these strains for cell growth and lipid synthesis after glucose was nearly consumed up. Profiles of metabolites produced by Rhodococcus strains growing on different carbon sources (e.g., glucose, alkali lignin, and dilute acid flowthrough-pretreated poplar wood slurry) confirmed lignin conversion during co-fermentation, and indicated novel metabolic capacities and unexplored metabolic pathways in these organisms. Proteome profiles suggested that lignin depolymerization by Rhodococci sp. involved multiple peroxidases with accessory oxidases. Besides the β-ketoadipate pathway, the phenylacetic acid (PAA) pathway was another potential route for the in vivo ring cleavage activity. In addition, deficiency of reducing power and cellular oxidative stress probably led to lower lipid production using lignin as the sole carbon source than that using glucose. This work demonstrated a potential strategy for efficient bioconversion of both lignin and glucose into lipids by co-culture of multiple natural and engineered Rhodococcus strains. In addition, the involvement of PAA pathway in lignin degradation can help to further improve lignin utilization, and the combinatory proteomics and bioinformatics strategies used in this study can also be applied into other systems to reveal the metabolic and regulatory pathways for balanced cellular metabolism and to select genetic targets for efficient conversion of both lignin and carbohydrates into biofuels.

Abstract: Sustainable chemical production should rely on the valorization of crude renewable resources. Waste biomass refining complies with bioeconomy and circular economy initiatives. In this regard, sugar...

Abstract: Despite the potential of biotechnological processes for one-carbon (C1) bioconversion, efficient biocatalysts required for their implementation are yet to be developed. To address intrinsic limitations of native C1 biocatalysts, here we report that 2-hydroxyacyl CoA lyase (HACL), an enzyme involved in mammalian α-oxidation, catalyzes the ligation of carbonyl-containing molecules of different chain lengths with formyl-coenzyme A (CoA) to produce C1-elongated 2-hydroxyacyl-CoAs. We discovered and characterized a prokaryotic variant of HACL and identified critical residues for this newfound activity, including those supporting the hypothesized thiamine pyrophosphate-dependent acyloin condensation mechanism. The use of formyl-CoA as a C1 donor provides kinetic advantages and enables C1 bioconversion to multi-carbon products, demonstrated here by engineering an Escherichia coli whole-cell biotransformation system for the synthesis of glycolate and 2-hydroxyisobutyrate from formaldehyde and formaldehyde plus acetone, respectively. Our work establishes a new approach for C1 bioconversion and the potential for HACL-based pathways to support synthetic methylotrophy.

Abstract: Increases in global human population are leading to increasing demands for food production and waste management. Insect biomass is a sustainable alternative to traditional animal feeds when insects are produced on lignocellulosic by-products. Resources high in lignocellulose have high carbon to nitrogen ratios and require nitrogen supplementation to accelerate bioconversion. Here we report on studies that examine the influence of nitrogen supplementation of almond hull-based feedstocks on black soldier fly larvae (Hermetia illucens L.) cultivation and composition. Decreasing carbon to nitrogen ratio from 49 to 16 increased larvae harvest dry weight, specific larvae growth, and yield by 36%, 31%, and 51%, respectively. However, the decrease in carbon to nitrogen ratio decreased larvae methionine and cysteine contents by 11% and 13%, respectively. The findings demonstrate that carbon to nitrogen ratio can be managed to enhance bioconversion of lignocellulose to larvae, but that this management approach can reduce larvae amino acid content.

Abstract: The rising demand of bio-vanillin and the possibility to use microbial biotransformation to produce this compound from agroindustrial byproducts are economically attractive. However, there are still several bottlenecks, including substrate and product toxicity, formation of undesired products and genetic stability of the recombinant strains, that impede an efficient use of recombinant Escherichia coli strains to make the whole process cost effective. To overcome these problems, we developed a new E. coli strain, named FR13, carrying the Pseudomonas genes encoding feruloyl-CoA synthetase and feruloyl-CoA hydratase/aldolase integrated into the chromosome and, using resting cells, we demonstrated that the vanillin yield and selectivity were strongly affected by the physiological state of the cells, the temperature used for the growth and the recovery of the biomass and the composition and pH of the bioconversion buffer. The substrate consumption rate and the vanillin yield increased using a sodium/potassium phosphate buffer at pH 9.0 as bioconversion medium. Optimization of the bioprocess variables, using response surface methodology, together with the use of a two-phase (solid-liquid) system for the controlled release of ferulic acid allowed us to increase the vanillin yield up to 28.10 ± 0.05 mM. These findings showed that recombinant plasmid-free E. coli strains are promising candidates for the production of vanillin at industrial scale and that a reduction of the cost of the bioconversion process requires approaches that minimize the toxicity of both ferulic acid and vanillin.

Abstract: Globally, biofuel and oleochemical production based on plant oils negatively affects biodiversity. As an alternative triglyceride source, lipid production from oleaginous yeasts faces numerous challenges in feedstock availability, lipid productivity, downstream processing, and waste treatment, prohibiting the design of a cost-competitive process with regard to plant equivalents. In this study, we present a fully integrated operation for microbial oil production, which consolidates upstream and downstream processing with side-stream recycling. Co-fermentation of sugar and acetic acid was successfully implemented in fed-batch, semi-continuous, and continuous fermentation modes. Process validation was conducted at a 25 L scale with a lipid productivity of 1.2 g L−1 h−1. Cutaneotrichosporon oleaginosus cell debris was used as an inducer in Trichoderma reesei fermentation for on-site generation of yeast-specific cell-wall hydrolases. In situ hydrolase application allowed for efficient C. oleaginosus cell lysis (85% w/w) and simultaneous lipid release. A subsequent centrifugation step yielded 90% (w/w) recovery of intracellular lipids without the need for any organic solvent. The nutrient-rich water phase was applied as an internal sugar source for subsequent yeast fermentation cycles. With this yeast hydrolysate, the lipid productivity was considerably increased to 2.4 g L−1 h−1. A techno-economic analysis of the current lipid production processes estimated costs at $1.6 per kg lipid. Moreover, life cycle assessment analysis indicated an emission of 3.56 kg CO2 equivalents for every 1 kg produced yeast oil. Accordingly, we established an integrated operation for bioconversion of acetic acid and sugar to sustainable lipids at maximum productivity coupled with minimal waste generation and energy consumption.

Abstract: There is great interest in microbial conversion of methane, an abundant resource, into valuable liquid chemicals. While aerobic bioconversion of methane to liquid chemicals has been reported, studies of anaerobic methane bioconversion to liquid chemicals are rare. Here we show that a microbial culture dominated by Candidatus 'Methanoperedens nitroreducens', an anaerobic methanotrophic archaeon, anaerobically oxidizes methane to produce acetate, indirectly via reaction intermediate(s), when nitrate or nitrite is supplied as an electron acceptor under a rate-limiting condition. Isotopic labeling tests showed that acetate was produced from certain intracellular storage compounds that originated from methane. Fluorescence in situ hybridization and Nile red staining demonstrated that polyhydroxyalkanoate in M. nitroreducens was likely one of the intracellular storage compounds for acetate production, along with glycogen. Acetate is a common substrate for the production of more valuable chemicals. The microbial conversion discovered in this study potentially enables a new approach to the use of methane as a feedstock for the chemical market.

Abstract: Rice straw, an abundant agro-residue, is available for energy production. In many parts of Asian countries, it is burnt on fields causing harm to the environment. Rice straw contains lignin, cellulose, hemicelluloses, and silicates making it recalcitrant. Pretreatment processes disintegrate lignin-carbohydrate matrix for efficient bioconversion of polysaccharides to fermentable sugars. A good number of physical, biological and chemical processes have been tried but degradation of polysaccharides and subsequent fermentation is still a challenge. Alkaline pretreatment causes effective delignification and swelling of biomass. The present study was performed on alkaline pretreatment of rice straw with 1% NaOH by autoclaving for 30 min at 121 °C at 10% solid loading. It was extracted with water to remove lignins, solids separated by filtrations and washed again to neutralize the pH. Water washing also led to removal of phenolic inhibitors. High (63%) glucan enrichment was obtained with concomitant lignin loss. Dry matter loss was around 50%. Enzymatic saccharification of the pretreated solids at 5 and 10% with Accellerase® 1500 for 24 h at 50 °C gave saccharification efficiency 76 and ~ 50% respectively. Hydrolysates containing 18 and 23 gL−1 sugars, supplemented with minimal salts, yeast extract, fermented by S. cerevisiae LN for 24 h yielded ~ 2 and 4 gL−1 ethanol with fermentation efficiency 55–66%. Thus, NaOH pretreatment is a cost effective option for ethanol production from rice straw. Lignin removed in prehydrolysates can be recovered by acidification.

Abstract: This study was designed to unravel the role of Lactobacillus rhamnosus in the bioconversion of cranberry proanthocyanidins and cytotoxicity of resulting metabolites to hepatocellular carcinoma HepG2 cells. Crude (CR) and flavonol+dihydrochalcone- (FL+DHC-), anthocyanin- (AN-), proanthocyanidin- (PR-), and phenolic acid+catechin- (PA+C-) rich fractions were subjected to fermentation with L. rhamnosus at 37°C for 12, 24, and 48 h under anaerobic conditions. The major metabolites produced by bioconversion of polyphenols were 4-hydroxyphenylacetic acid, 3-(4-hydroxyphenyl)propionic acid, hydrocinnamic acid, catechol, and pyrogallol. Furthermore, cytotoxicity of the biotransformed extracts was compared to their parent extracts using human hepatocellular carcinoma HepG2 cells. The results showed that PR-biotransformed extract completely inhibited HepG2 cell proliferation in a dose- and time-dependent manner with IC50 values of 47.8 and 20.1 μg/mL at 24 and 48 h, respectively. An insight into the molecular mechanisms involved revealed that the cytotoxic effects of PR at 24 h incubation were mitochondria-controlled and not by proapoptotic caspase-3/7 dependent. The present findings suggest that the application of a bioconversion process using probiotic bacteria can enhance the pharmacological activities of cranberry proanthocyanidins by generating additional biologically active metabolites.

Abstract: Agricultural chemicals either used as nutrient inputs for soil fertility or pesticides are creating physicochemical and biological deterioration of the soils and disturbing the agro-ecosystems worldwide. Alarming concerns towards integrated agroecology demand for renewed interest in low-external input-based farming practices. These practices comprise strengthening of soil biological properties, recycling of inherent soil minerals and reuse of agricultural residual wastes. We described approaches for the bioconversion of agricultural residual wastes into value-added compost. The process involves conversion of residual waste into raw compost followed by its fortification with beneficial decomposer microorganisms to produce quality fortified compost product. Finally, incubation of fortified compost with single or consortia of beneficial microorganisms like N-fixers, P-solubilizers or K-mobilizers and biocontrol agents further enriches compost to produce bioorganic products. Bioconversion of agricultural wastes into compost using potential decomposer microorganisms and fortification of decomposed organic matter with beneficial bacterial and fungal species is of immense importance. Additional enrichment of compost with botanicals, humic acid, amino acids, mineral nutrients, phytohormones etc. may also add value to the bioinput products. In an integrated way, on-farm production of raw compost using different agricultural residual wastes and its further fortification with bioorganic farm inputs can help farmers produce value-added compost products for direct application in the crop production. Adoption of microbial bioconversion technologies and their field applications may become eco-enterprising for the rural resource-poor farming communities for enhancing their livelihood along with improving farm productivity and soil health.

Abstract: The dual roles of efficient degradation and bioconversion of a wide range of organic wastes into valuable animal protein and organic fertiliser, has led to increased interest in black soldier fly (...

Abstract: Plastic production and waste generation will continue to rise as nations worldwide grow economically. In this work, we detail a pyrolysis-based bioconversion process for polypropylene (PP) to produce value-added fatty acids (FAs). PP pellets were depolymerized by pyrolysis, generating oil that consisted of mainly branched chain fatty alcohols and alkenes. The oil was mixed with biodegradable surfactants and trace nutrients and mechanically homogenized. The resulting medium, OP4, was used for fermentation by Yarrowia lipolytica strain 78-003. Y. lipolytica assimilated > 80% of the substrate over 312 h, including 86% of the fatty alcohols. Y. lipolytica produced up to 492 mg L-1 lipids, compared with 216 mg L-1 during growth in surfactant-based control medium. C 18 compounds, including oleic acid, linoleic acid, and stearic acid, were the predominant products, followed by C 16 compounds palmitic and palmitoleic acids. Two percent of the products was C 20 compounds. The majority of the products were unsaturated FAs. Growth on hydrophobic substrates (OP4 medium, hexadecane) was compared with growth on hydrophilic substrates (glucose, starch). The resulting FA profiles revealed an absence of short-chain fatty acids during growth on hydrophobic media, findings consistent with ex novo FA biosynthesis. Overall, FA profiles by Y. lipolytica during growth in OP4 medium were similar to FA profiles while growing on natural substrates. The process described here offers an alternative approach to managing postconsumer plastic waste.

Abstract: Lignin is a potential feedstock for microbial conversion into various chemicals. However, the microbial degradation rate of native or technical lignin is low, and chemical depolymerization is needed to obtain reasonable conversion rates. In the current study, nine bacterial strains belonging to the Pseudomonas and Rhodococcus genera were evaluated for their ability to grow on alkaline-treated softwood lignin as a sole carbon source. Pseudomonas fluorescens DSM 50090 and Rhodococcus opacus DSM1069 showed the best growth of the tested species on plates with lignin. Further evaluation of P. fluorescens and R. opacus was made in liquid cultivations with depolymerized softwood Kraft lignin (DL) at a concentration of 1 g/L. Size-exclusion chromatography (SEC) showed that R. opacus consumed most of the available lower-molecular weight compounds (approximately 0.1–0.4 kDa) in the DL, but the weight distribution of larger fractions was almost unaffected. Importantly, the consumed compounds included guaiacol—one of the main monomers in the DL. SEC analysis of P. fluorescens culture broth, in contrast, did not show a large conversion of low-molecular weight compounds, and guaiacol remained unconsumed. However, a significant shift in molecular weight distribution towards lower average weights was seen after cultivation with P. fluorescens. Rhodococcus opacus and P. fluorescens were identified as two potential microbial candidates for the conversion/consumption of base-catalyzed depolymerized lignin, acting on low- and high-molecular weight lignin fragments, respectively. These findings will be of relevance for designing bioconversion of softwood Kraft lignin.

Abstract: A new bioconversion-composting process of golden needle mushroom (Flammulina velutipes) root waste was established using black soldier fly larvae (BSFL) to produce added-value biomass and organic fertilizer. The entire process included two stages: BSFL conversion and conventional composting, which lasted 40 days. After a 26-day bioconversion, a considerable quantity of BSFL was obtained, which could be used to produce the protein feed, chitin, antibacterial peptide, biodiesel and so on. The bioconversion parameters of waste reduction rate and BSFL conversion rate were 54.8 ± 0.4 and 5.06 ± 0.27%, respectively. Following the 14-day conventional composting, the mushroom root waste turned into an organic fertilizer. The moisture, organic matter, total nitrogen, total phosphorus, and total potassium concentrations of the obtained fertilizer were 29.3 ± 0.9, 76.0 ± 1.7, 2.74 ± 0.09, 2.07 ± 0.06, and 1.34 ± 0.15%, respectively. The germination index was 65.7 ± 3.2% for Chinese cabbage and 52.9 ± 1.3% for rapeseed. To accelerate bioconversion, increase output of larvae biomass and shorten the developing time of BSFL, two auxiliary materials, nitrogen source, namely, bran and kitchen wastes, were mixed with mushroom roots. Both were significantly effective, and the recommended percentage was 40%. In that case, the developing time reduced to approximately 15–16 days, and the BSFL conversion rate increased by 31.2–172.7%. Accordingly, one simple process flowchart was drawn, and the output was estimated.

Abstract: Background and objective: Polyhydroxyalkanoates have drawn significant attention as alternative to petroleum-based plastics; however, their industrial production is still hindered by the costly feed materials. Co-generation of other high-value products in addition to polyhydroxyalkanoate by the same microbial strains can be helpful in alleviating overall production cost up to 50%. This study for the first time demonstrates that polyhydroxyalkanoate and astaxanthin-rich carotenoids can be co-produced by Paracoccus sp. LL1 using waste cooking oil as substrate. Material and methods: The halophilic strain of Paracoccus sp. LL1 was grown under batch fermentation using mineral media supplemented with 1% (v v-1) waste cooking oil. Different surfactants were used to improve substrate utilization. Polyhydroxyalkanoate obtained after the fermentation was characterized by fluorescent microscopy, gas chromatography, and Fourier Transform Infra-Red spectroscopy. Results and conclusion: Oil as a substrate, led to 1.0 g l-1 poly (3-hydroxybutyrate- co -3-hydroxyvalerate) with concomitant production of 0.89 mg l-1 of carotenoids after 96 h. An enhancement of 2.7-folds in total cell dry mass was achieved when 0.1% (v v-1) Tween-80 was used as surfactant for ease in oil metabolism. Paracoccus sp. LL1 has the potential to serve as a single cell factory for bioconversion of cheap substrates into high value products. Conflict of interest: The authors declare no conflict of interest.