Two-stage microbial conversion of crude glycerol to 1,3-propanediol and polyhydroxyalkanoates after pretreatment.

With increasing demand for biodiesel, crude glycerol as a by-product in biodiesel production has been generated and oversupplied. This study, therefore, explored the pretreatment and a subsequent two-stage microbial system to convert crude glycerol into high value-added products: 1,3-propanediol (1,3-PDO) and polyhydroxyalkanoates (PHAs). After pretreatment, long chain fatty acids (LCFAs) could be effectively removed from crude glycerol to eliminate the inhibition effects on subsequent microbial process. In the anaerobic fermentation, when fed treated crude glycerol increased from 20 g/L to 100 g/L, 1,3-PDO yield decreased from 0.438 g/g to 0.345 g/g and accompanied carboxylic acids shifted from acetate and lactate dominant to lactate overwhelmingly dominant. Meanwhile, the relative abundance of Clostridiales sustained around 50% but Enterobacteriales increased from 19% to 53%. Further fed glycerol increase to 140 g/L resulted in severe substrate inhibition, which could be relieved by intermittent feeding. In aerobic process, glycerol anaerobic digestion effluent (ADE) was fed to the consortium of Bacillus megaterium and Corynebacterium hydrocarbooxydans for selectively consumption of carboxylic acids and residual glycerol from 1,3-PDO to produce PHAs as a secondary high value-added product. The consortium accumulated maximum 8.0 g/L poly (3-hydroxybutyrate) (PHB), and 1,3-PDO purity increased from initial 27.7% to almost 100% when fed with 100 g/L glycerol ADE. Overall, this study provided comprehensive and insightful information on microbial conversion of crude glycerol to high value-added products after pretreatment.

Quantum modeling simulates nutrient effect of bioplastic polyhydroxyalkanoate (PHA) production in Pseudomonas putida.

Polyhydroxyalkanoates (PHAs) could be used to make sustainable, biodegradable plastics. However, the precise and accurate mechanistic modeling of PHA biosynthesis, especially medium-chain-length PHA (mcl-PHA), for yield improvement remains a challenge to biology. PHA biosynthesis is typically triggered by nitrogen limitation and tends to peak at an optimal carbon-to-nitrogen (C/N) ratio. Specifically, simulation of the underlying dynamic regulation mechanisms for PHA bioprocess is a bottleneck owing to surfeit model complexity and current modeling philosophies for uncertainty. To address this issue, we proposed a quantum-like decision-making model to encode gene expression and regulation events as hidden layers by the general transformation of a density matrix, which uses the interference of probability amplitudes to provide an empirical-level description for PHA biosynthesis. We implemented our framework modeling the biosynthesis of mcl-PHA in Pseudomonas putida with respect to external C/N ratios, showing its optimization production at maximum PHA production of 13.81% cell dry mass (CDM) at the C/N ratio of 40:1. The results also suggest the degree of P. putida's preference in channeling carbon towards PHA production as part of the bacterium's adaptative behavior to nutrient stress using quantum formalism. Generic parameters (kD, kN and theta θ) obtained based on such quantum formulation, representing P. putida's PHA biosynthesis with respect to external C/N ratios, was discussed. This work offers a new perspective on the use of quantum theory for PHA production, demonstrating its application potential for other bioprocesses.

Incorporation of lead into pyromorphite: Effect of anion replacement on lead stabilization.

Previous studies demonstrate that the leaching of heavy metals in unreliable waste forms causes serious environmental pollution and health concerns. Thus, research is focused on identifying an effective, safe strategy for disposing of metal-laden solid waste such as lead (Pb). This study evaluated the effect of anion replacement in the structure of pyromorphite (Pb10(PO4)6Cl2, a common mineral phase for Pb sequestering) on Pb stabilization. Phosphate (PO43-) at the tetrahedral pyromorphite site was simultaneously replaced by silicate (SiO44-) and sulphate (SO42-) in a controlled thermal treatment. The lattice expanded with the incorporation of additional SiO44- and SO42-. Furthermore, the unit cell parameters of the solid solutions evolved linearly with an increase in the substitution degree (x in Pb10(SiO4)x(SO4)x(PO4)(6-2x)Cl2). This research also demonstrated that Pb distributed into amorphous in a PO43--deficient matrix, while asisite (Pb7SiO8Cl2) was formed when the matrix was dominated by SiO44- and SO42-. The leaching results showed the isomorphous substitution in the target system rendered the products less durable towards acidic attack. Moreover, the fully isomorphous-substituted product (x = 3) showed more than two orders of magnitude lower leaching resistance than the PO43--rich phase (x = 0). The lattice expansion, resulting from the isomorphous substitution, suggested that a lower dissolution energy was required in a PO43--deficient matrix. The leaching kinetics pointed to a product with a lower apparent activation energy in the leaching process. The findings of this study provide unique insight into the design and optimization of waste forms for the immobilization of heavy metals.

Genomic driven factors enhance biocatalyst-related cellulolysis potential in anaerobic digestion.

Anaerobic digestion (AD) is a promising technology to recover bioenergy from biodegradable biomass, including cellulosic wastes. Through a few fractionation/separation techniques, cellulose has demonstrated its potential in AD, but the performance of the process is rather substrate-specific, as cellulolysis bacteria are sensitive to the enzyme-substrate interactions. Cellulosome is a self-assembled enzyme complex with many functionalized modules in the bacteria which has been gradually studied, however the genomic fingerprints of the culture-specific cellulosome in AD are relatively unclear especially under processing conditions. To clarify the key factors affecting the cellulosome induced cellulolysis, this review summarized the most recent publications of AD regarding the fates of cellulose, sources and functional genes of cellulosome, and omics methods for functional analyses. Different processes for organic treatment including applying food grinds in sewer, biomass valorization, cellulose fractionation, microaeration, and enzymatic hydrolysis enhanced fermentation, were highlighted to support the sustainable development of AD technology.

Combined Effect of Activated Carbon Particles and Non-Adsorptive Spherical Beads as Fluidized Media on Fouling, Organic Removal and Microbial Communities in Anaerobic Membrane Bioreactor.

The combined effect of acrylonitrile butadiene styrene (ABS) spherical beads and granular activated carbon (GAC) particles as fluidized media on the performance of anaerobic fluidized bed membrane bioreactor (AFMBR) was investigated. GAC particles and ABS beads were fluidized together in a single AFMBR to investigate membrane fouling and organic removal efficiency as well as energy consumption. The density difference between these two similarly sized media caused the stratified bed layer where ABS beads are fluidized above the GAC along the membrane. Membrane relaxation was effective to reduce the fouling and trans-membrane pressure (TMP) below 0.25 bar could be achieved at 6 h of hydraulic retention time (HRT). More than 90% of soluble chemical oxygen demand (SCOD) was removed after 80 d operation. Biogas consisting of 65% of methane was produced by AFMBR, suggesting that combined use of GAC and ABS beads did not have any adverse effect on methane production during the operational period. Scanning Electron Microscope (SEM) examinations showed the adherence of microbes to both media. However, 16S rRNA results revealed that fewer microbes attached to ABS beads than GAC. There were also compositional differences between the ABS and GAC microbial communities. The abundance of the syntrophs and exoelectrogens population on ABS beads was relatively low compared to that of GAC. Our result implied that syntrophic synergy and possible occurrence of direct interspecies electron transfer (DIET) might be facilitated in AFMBR by GAC, while traditional methanogenic pathways were dominant in ABS beads. The electrical energy required was 0.02 kWh/m3, and it was only about 13% of that produced by AFMBR.

Metatranscriptomic evidence for classical and RuBisCO-mediated CO2 reduction to methane facilitated by direct interspecies electron transfer in a methanogenic system.

In a staged anaerobic fluidized-bed ceramic membrane bioreactor, metagenomic and metatranscriptomic analyses were performed to decipher the microbial interactions on the granular activated carbon. Metagenome bins, representing the predominating microbes in the bioreactor: syntrophic propionate-oxidizing bacteria (SPOB), acetoclastic Methanothrix concilii, and exoelectrogenic Geobacter lovleyi, were successfully recovered for the reconstruction and analysis of metabolic pathways involved in the transformation of fatty acids to methane. In particular, SPOB degraded propionate into acetate, which was further converted into methane and CO2 by M. concilii via the acetoclastic methanogenesis. Concurrently, G. lovleyi oxidized acetate into CO2, releasing electrons into the extracellular environment. By accepting these electrons through direct interspecies electron transfer (DIET), M. concilii was capable of performing CO2 reduction for further methane formation. Most notably, an alternative RuBisCO-mediated CO2 reduction (the reductive hexulose-phosphate (RHP) pathway) is transcriptionally-active in M. concilii. This RHP pathway enables M. concilii dominance and energy gain by carbon fixation and methanogenesis, respectively via a methyl-H4MPT intermediate, constituting the third methanogenesis route. The complete acetate reduction (2 mole methane formation/1 mole acetate consumption), coupling of acetoclastic methanogenesis and two CO2 reduction pathways, are thermodynamically favorable even under very low substrate condition (down to to 10-5 M level). Such tight interactions via both mediated and direct interspecies electron transfer (MIET and DIET), induced by the conductive GAC promote the overall efficiency of bioenergy processes.

Determination of monomeric composition in polyhydroxyalkanoates by liquid chromatography coupled with on-line mass spectrometry and off-line nuclear magnetic resonance.

Polyhydroxyalkanoates (PHAs) are commercially-valuable biocompatible and biodegradable polymers with many potential medical, pharmaceutical and other industrial applications. The analysis of PHA monomeric composition is especially challenging due to the broad chemical diversity of PHA monomers and lack of analytical standards to represent the chemically-diverse PHA monomer constituents. In this study, a novel strategy based on on-line liquid chromatography-mass spectrometry (LC-MS) and off-line liquid chromatography-nuclear magnetic resonance (LC-NMR) was established to quantify seven PHA monomers with available standards and used to elucidate the structures of unknown PHA monomers. The strategy was successfully applied for the determination of monomeric composition in bacterial PHAs isolated from Pseudomonads cultivated on different carbon sources after hydrolysis. The results of this work demonstrated that the newly-developed strategy was efficient, repeatable, and could have good potential to be employed for detailed analysis of PHA monomeric composition.

Microbiology and potential applications of aerobic methane oxidation coupled to denitrification (AME-D) process: A review.

Aerobic methane oxidation coupled to denitrification (AME-D) is an important link between the global methane and nitrogen cycles. This mini-review updates discoveries regarding aerobic methanotrophs and denitrifiers, as a prelude to spotlight the microbial mechanism and the potential applications of AME-D. Until recently, AME-D was thought to be accomplished by a microbial consortium where denitrifying bacteria utilize carbon intermediates, which are excreted by aerobic methanotrophs, as energy and carbon sources. Potential carbon intermediates include methanol, citrate and acetate. This mini-review presents microbial thermodynamic estimations and postulates that methanol is the ideal electron donor for denitrification, and may serve as a trophic link between methanotrophic bacteria and denitrifiers. More excitingly, new discoveries have revealed that AME-D is not only confined to the conventional synergism between methanotrophic bacteria and denitrifiers. Specifically, an obligate aerobic methanotrophic bacterium, Methylomonas denitrificans FJG1, has been demonstrated to couple partial denitrification with methane oxidation, under hypoxia conditions, releasing nitrous oxide as a terminal product. This finding not only substantially advances the understanding of AME-D mechanism, but also implies an important but unknown role of aerobic methanotrophs in global climate change through their influence on both the methane and nitrogen cycles in ecosystems. Hence, further investigation on AME-D microbiology and mechanism is essential to better understand global climate issues and to develop niche biotechnological solutions. This mini-review also presents traditional microbial techniques, such as pure cultivation and stable isotope probing, and powerful microbial techniques, such as (meta-) genomics and (meta-) transcriptomics, for deciphering linked methane oxidation and denitrification. Although AME-D has immense potential for nitrogen removal from wastewater, drinking water and groundwater, bottlenecks and potential issues are also discussed.

Bioconversion of styrene to poly(hydroxyalkanoate) (PHA) by the new bacterial strain Pseudomonas putida NBUS12.

Styrene is a toxic pollutant commonly found in waste effluents from plastic processing industries. We herein identified and characterized microorganisms for bioconversion of the organic eco-pollutant styrene into a valuable biopolymer medium-chain-length poly(hydroxyalkanoate) (mcl-PHA). Twelve newly-isolated styrene-degrading Pseudomonads were obtained and partial phaC genes were detected by PCR in these isolates. These isolates assimilated styrene to produce mcl-PHA, forming PHA contents between 0.05±0.00 and 23.10±3.25% cell dry mass (% CDM). The best-performing isolate was identified as Pseudomonas putida NBUS12. A genetic analysis of 16S rDNA and phaZ genes revealed P. putida NBUS12 as a genetically-distinct strain from existing phenotypically-similar bacterial strains. This bacterium achieved a final biomass of 1.28±0.10 g L(-1) and PHA content of 32.49±2.40% CDM. The extracted polymer was mainly comprised of 3-hydroxyhexanoate (C6 ), 3-hydroxyoctanoate (C8 ), 3-hydroxydecanoate (C10 ), 3-hydroxydodecanoate (C12 ), and 3-hydroxytetradecanoate (C14 ) monomers at a ratio of 2:42:1257:17:1. These results collectively suggested that P. putida NBUS12 is a promising candidate for the biotechnological conversion of styrene into mcl-PHA.

Enhanced styrene recovery from waste polystyrene pyrolysis using response surface methodology coupled with Box-Behnken design.

A work applied response surface methodology coupled with Box-Behnken design (RSM-BBD) has been developed to enhance styrene recovery from waste polystyrene (WPS) through pyrolysis. The relationship between styrene yield and three selected operating parameters (i.e., temperature, heating rate, and carrier gas flow rate) was investigated. A second order polynomial equation was successfully built to describe the process and predict styrene yield under the study conditions. The factors identified as statistically significant to styrene production were: temperature, with a quadratic effect; heating rate, with a linear effect; carrier gas flow rate, with a quadratic effect; interaction between temperature and carrier gas flow rate; and interaction between heating rate and carrier gas flow rate. The optimum conditions for the current system were determined to be at a temperature range of 470-505°C, a heating rate of 40°C/min, and a carrier gas flow rate range of 115-140mL/min. Under such conditions, 64.52% WPS was recovered as styrene, which was 12% more than the highest reported yield for reactors of similar size. It is concluded that RSM-BBD is an effective approach for yield optimization of styrene recovery from WPS pyrolysis.

Enhanced gas chromatography-mass spectrometry method for bacterial polyhydroxyalkanoates analysis.

A gas chromatography-mass spectrometry method for quantification of polyhydroxyalkanoates (PHAs), containing 4-carbon to 16-carbon monomers, even in the absence of standards, was developed. Strong linear correlations existed between PHA carbon number and retention time/response factor (R(2) ≥ 0.987). Based on the correlations, high recovery values, between 100.5% and 114.3%, were obtained for PHA polymers.

Caproate formation in mixed-culture fermentative hydrogen production.

Caproate always appears during fermentative H(2) production but its formation was not well explained. It possibly results from the secondary fermentation of ethanol and acetate or butyrate by some special species like Clostridium kluyveri. This study attempts to elucidate caproate formation during the fermentation H(2) production by using C. kluyveri as an example and evaluating several possible pathways of caproate formation. A detailed energetic analysis of the empirical data of an H(2)-producing reactor demonstrated that caproate can be formed from two substrates, either ethanol and acetate or ethanol and butyrate. The analysis showed that at least 5 mol ethanol per mole reaction was essential to support caproate formation under the experimental condition. The analysis also indicated that the secondary fermentation by C. kluyveri might be another pathway to spontaneously produce H(2), butyrate, and acetate in addition to the butyrate-acetate pathway. Co-production of caproate and H(2) from ethanol was thermodynamically feasible and contributed to at least 10-20% of total H(2) production in the reactor studied. It is also clarified that caproate formation is hydrogenogenic rather than hydrogenotrophic.