Biotransformation of the azo dye reactive orange 16 by Aspergillus flavus A5P1: Performance, genetic background, pathway, and mechanism.

This study reports the strain Aspergillus flavus A5P1 (A5P1), which is with the capable of degrading the azo dye reactive orange 16 (RO16). The mechanism of RO16 degradation by A5P1 was elucidated through genomic analysis, enzymatic analysis, degradation pathway analysis and oxidative stress analysis. Strain A5P1 exhibited aerobic degradation of RO16, with optimal degradation at an initial pH of 3.0. Genomic analysis indicates that strain A5P1 possesses the potential for acid tolerance and degradation of azo dye. Enzymatic analysis, combined with degradation product analysis, demonstrated that extracellular laccase, intracellular lignin peroxidase, and intracellular quinone reductase were likely key enzymes in the RO16 degradation process. Oxidative stress analysis revealed that cell stress responses may participate in the RO16 biotransformation process. The results indicated that the biotransformation of RO16 may involves biological processes such as transmembrane transport of RO16, cometabolism of the strain with RO16, and cell stress responses. These findings shed light on the biodegradation of RO16 by A5P1, indicating A5P1's potential for environmental remediation.

Improving efficiency and reducing enzyme inactivation during lipase-mediated epoxidation of α-pinene in a double-phase reaction system.

Chemoenzymatic epoxidation of olefin mediated by lipase is a green and environmentally friendly alternative process. However, the mass transfer barrier and lipase deactivation caused by the traditional organic-water biphasic reaction system have always been the focus of researchers' attention. To overcome these issues, we investigated the effects of reaction temperature and two important substrates (H2O2 and acyl donor) on the epoxidation reaction and interfacial mass transfer. As a result, we determined the optimal reaction conditions: a temperature of 30 °C, 30 wt-% H2O2 as the oxygen source, and 1 M lauric acid as the oxygen carrier. Additionally, by simulating the conditions of shaking flask reactions, we designed a batch reactor and added a metal mesh to effectively block the direct contact between high-concentration hydrogen peroxide and the enzyme. Under these optimal conditions, the epoxidation reaction was carried out for 5 h, and the product yield reached a maximum of 93.2%. Furthermore, after seven repetitive experiments, the lipase still maintained a relative activity of 51.2%.

Glutaraldehyde-crosslinked Rhizopus oryzae whole cells show improved catalytic performance in alkene epoxidation.

BACKGROUND: Existing methods for alkene epoxidation are based on lipase-catalysed perhydrolysis. However, the inactivation of the expensive lipase enzyme is problematic for enzymatic epoxidation at large scales due to the use of hydrogen peroxide and peracids at high concentrations in the reaction. The immobilisation of whole cells appears to be a promising approach to alleviate this problem. RESULTS: A green oxidation system containing hydrogen peroxide, Na3C6H5O7, an acyl donor, and glutaraldehyde (GA)-crosslinked cells of Rhizopus oryzae was developed for the epoxidation of alkenes. GA-crosslinked cells of Rhizopus oryzae were adopted as a biocatalyst into the epoxidation system. A variety of alkenes were oxidised with this system, with a 56-95% analytical yield of the corresponding epoxides. The catalytic performance of the crosslinked treated cells was substantially improved compared to that of the untreated cells and the initial reaction rate increased from 126.71 to 234.72 mmol/L/h, retaining 83% yields even after four batches of reactions. The addition of 3.5 mmol Na3C6H5O7 not only acts as an acid-trapping reagent to eliminate the negative effect of the carboxylic acid on the alkene oxide but also forms a saturated salt solution with the aqueous phase, affecting the concentration of H2O2 in the three phases and thus the epoxidation reaction. Organic solvents with a logP value > 0.68 were good at producing hydroxy peracids; however, this method is only suitable for oxidation in a two-liquid phase. CONCLUSIONS: Compared with other lipase biocatalysts, the GA-crosslinked whole-cell biocatalyst is inexpensive, readily available, and highly stable. Therefore, it can be considered promising for industrial applications.

Metagenomic insights into role of red mud in regulating fate of compost antibiotic resistance genes mediated by both direct and indirect ways.

In this study, the amendment of red mud (RM) in dairy manure composting on the fate of antibiotic resistance genes (ARGs) by both direct (bacteria community, mobile genetic elements and quorum sensing) and indirect ways (environmental factors and antibiotics) was analyzed. The results showed that RM reduced the total relative abundances of 10 ARGs and 4 mobile genetic elements (MGEs). And the relative abundances of total ARGs and MGEs decreased by 53.48% and 22.30% in T (with RM added) on day 47 compared with day 0. Meanwhile, the modification of RM significantly increased the abundance of lsrK, pvdQ and ahlD in quorum quenching (QQ) and decreased the abundance of luxS in quorum sensing (QS) (P < 0.05), thereby attenuating the intercellular genes frequency of communication. The microbial community and network analysis showed that 25 potential hosts of ARGs were mainly related to Firmicutes, Proteobacteria and Actinobacteria. Redundancy analysis (RDA) and structural equation model (SEM) further indicated that RM altered microbial community structure by regulating antibiotic content and environmental factors (temperature, pH, moisture content and organic matter content), which then affected horizontal gene transfer (HGT) in ARGs mediated by QS and MGEs. These results provide new insights into the dissemination mechanism and removal of ARGs in composting process.

Multifunctional wound dressing for rapid hemostasis, bacterial infection monitoring and photodynamic antibacterial therapy.

Wound management is a major global issue, and there is a growing challenge to develop more effective hemostatic dressings to control bleeding and prevent pathogen infections. In this study, a multifunctional wound dressing was developed to meet the clinical need. The hemostatic layer of wound dressing can quickly stop the bleeding. Meanwhile, the detection layer is used for real-time fluorescence monitoring of the bacterial colonization. When infection occurs, wound dressing is further subjected to illumination for in-situ photodynamic antibacterial treatment. In the rabbit ear artery hemostasis model, the hemostasis time of the wound dressing was 1 s. The detection limit of the wound dressing was 1.4 × 105 CFU/cm2 for Escherichia coli, 5.9 × 105 CFU/cm2 for Staphylococcus aureus, and 3.8 × 106 CFU/cm2 for Pseudomonas aeruginosa, respectively. Compared with the control group, an enhanced wound closure (up to 97.3%) were observed in mice treated with the wound dressing. In vitro and in vivo experiment results suggested that the wound dressing was effective in killing pathogenic bacterial and exhibited good biological compatibility, and induced no inflammatory reaction. The proposed design prevents massive bleeding and wound infection, and further promotes wound healing. STATEMENT OF SIGNIFICANCE: In this work, we developed a multifunctional wound dressing, capable of rapid hemostasis, colorimetric monitoring of bacterial infection, and in situ photodynamic antibacterial. The hemostatic layer can quickly stop the bleeding due to its large specific surface area and adsorption pore size for platelet at bleeding site. Meanwhile, the detection layer can intelligently monitor the bacterial infection and respond to report bacterial infection by emitting fluorescence. When infection occurs, wound dressing can be used for in-situ photodynamic antibacterial treatment. In vitro and in vivo results showed that the wound dressing was biocompatible, prevented massive bleeding and wound infection, and further promoted wound healing.

Simultaneous leaching of multiple heavy metals from a soil column by extracellular polymeric substances of Aspergillus tubingensis F12.

The use of the biological agents for leaching heavy metals from contaminated soils is a very promising method that is both efficient and eco-friendly. In this study, a fungus Aspergillus tubingensis F12 was reported to possess a strong adsorption capacity for various heavy metal ions and shown to adsorb 90.8% Pb, 68.4% Zn, 64.5% Cr, 13.1% Cu, 12.9% Ni, and 6.9% Cd in aqueous solution. As extracellular polymeric substance (EPS) was found to play a leading role in the adsorption of metal ions, we applied EPS as a leaching agent to simultaneously remove six metals from soil in a column leaching experiment. The flow rate, initial solution pH, initial EPS concentration, and ionic strength were investigated using response surface methodology. The minimum and maximum metal leaching capacities were determined to be 0.089 mg/g and 3.703 mg/g, respectively. Verified by Fourier transform infrared spectroscopy, scanning electron microscope energy dispersive X-ray spectroscopy, and X-ray photoelectron spectroscopy, we made the preliminary deductions that ion exchange determines the leaching capacity limit and that biosorption plays a large role in reaching that limit. Additionally, the redox behaviour of Cu produced more carboxyl groups, which increased the adsorption of heavy metals. The ecological impact of this method was also examined; we found that the influences of leaching with EPS on soil properties and microbial community structure were slight. Therefore, the reported leaching process might have application prospects for metal removal from soil.

Preparation of PVDF/CdS/Bi2WO6/ZnO hybrid membrane with enhanced visible-light photocatalytic activity for degrading nitrite in water.

In this work, a visible light-driven ternary heterojunction photocatalyst, CdS/Bi2WO6/ZnO, was synthesized using hydrothermal, ultrasonic dispersion, and deposition precipitation methods. The results show that photocatalysts with flower-like heterostructures were obtained, which could efficiently separate electron-hole pairs, and the photocatalytic activity was thereby significantly enhanced. Furthermore, CdS/Bi2WO6/ZnO and polyvinylidene fluoride (PVDF) were used to fabricate hybrid membranes via a phase-conversion method. The samples were characterized using SEM, TEM, EDX, XRD, DRS, XPS, PL, and N2 adsorption-desorption isotherms, and the transient photocurrent response. The photocatalytic activity of the hybrid membrane was evaluated, and 92.58% of the nitrite was converted into non-toxic substances within 4 h under simulated sunlight irradiation. This result indicated that the photocatalyst exhibited a good photocatalytic activity after immobilization. The possible mechanism was elucidated by studying the product during the photocatalytic degradation, and the effects of different pH values, electron scavengers, and hole scavengers on the photocatalytic performance were further investigated.

A novel neutral and thermophilic endoxylanase from Streptomyces ipomoeae efficiently produced xylobiose from agricultural and forestry residues.

Endoxylanases capable of producing high ratios of xylobiose from agricultural and forestry residues in neutral and high temperature conditions are attractive for the prebiotic and alternative sweetener industries. In this study, a putative glycosyl hydrolase gene from Streptomyces ipomoeae was cloned and expressed in Escherichia coli. The recombinant enzyme, named as SipoEnXyn10A, hydrolyzed beechwood xylan in endo-action mode releasing xylobiose as its main end product. It was most active at pH 6.5 and 75-80 °C and showed remarkable stability at 65 °C. The xylobiose yield from 10 g corncob and moso bamboo reached 1.123 ±â€¯0.021 and 0.229 ±â€¯0.005 g, respectively, at pH 6.5 and 70 °C, whichwas higher than other reports using the same material. Moreover, high ratios of xylobiose in the xylose-based product of about 85% were obtained from corncob, moso bamboo sawdust, cassava stem and Chinese fir sawdust. These results demonstrated that SipoEnXyn10A has potential for industrial application.

Immobilization of Candida cylindracea Lipase by Covalent Attachment on Glu-Modified Bentonite.

Alkaline Ca-bentonite, obtained upon acid activation and base load of natural bentonite, has a good anion exchange capability. Glu-modified alkaline Ca-bentonites were further prepared by covalent binding with glutamic acid for the immobilization of lipase OF from Candida cylindracea. The obtained immobilized lipase demonstrated a significantly higher catalytic activity than that of unmodified alkaline Ca-bentonite, giving a specific activity of 62.1 U mg-1 protein, twice that of the unmodified carrier, and a total activity of 391.2 U g-1 support, retaining ~ 82.3% of the activity after being reused five times for olive oil emulsion hydrolysis. X-ray diffraction and Fourier transform infrared spectroscopy assays demonstrated the successful immobilization of the lipase on the surface of the bentonite. Upon immobilization, the thermostability of the lipase improved remarkably. At 50 °C, free lipase retained only 6.0% of its initial activity at 6 h, in comparison with 15% for Ca-Bent-lipase and 50% for Glu-Ca-Bent-lipase after 8 h. The Glu-Ca-Bent-lipase is proved as an effective biocatalyst for the biodiesel preparation, improving the transesterification reaction conversion from 52.8% in the condition of free lipase to 99.9% and keeping at 56.2% after being reused five times, while the free lipase was inactive upon two reuses. The above results provide a new route in the use of inexpensive bentonite for the enzyme immobilization.

Synergic effect of adsorption and biodegradation enhance cyanide removal by immobilized Alcaligenes sp. strain DN25.

A high efficiency and stability polyurethane-foam (PUF)-immobilized cell system was constructed to remove cyanide based on simultaneous adsorption and biodegradation (SAB). The performance of the PUF-immobilized system was evaluated by comparison with the freely suspended cell system. The SAB system exhibited more effective and robust, and could still remain degradation activity even at 40 °C or pH 11.0. The SAB system completely removed 500 mg CN-/L within 8 h at 30 °C, pH 8.0, and 120 rpm, whereas 12 h were required for the free cells system. Moreover, the SAB system showed apparent superiority in removing higher concentration cyanide up to 1200 mg CN-/L. A continuously stirred tank bioreactor (CSTR) was successfully designed and steadily operated with approximately 85% of the total average removal efficiency for 52 days at an influent cyanide concentration of 100-200 mg/L, which demonstrated a favorable reliability. Cyanide removal process could be well described using a pseudo first-order model, and the higher apparent rate constants (k) of the immobilized cells showed the synergic effect of adsorption and biodegradation significantly enhanced cyanide removal. Preliminarily, it was found that the foam characteristic might play a not negligible role on the cyanide-degrading enzyme expression of strain DN25 in the SAB system.

Purification and Characterization of a Novel ß-Cypermethrin-Degrading Aminopeptidase from Pseudomonas aeruginosa GF31.

In this study, a novel ß-cypermethrin-degrading enzyme was isolated and purified by 32.8 fold from the extracellular cell-free filtrate of Pseudomonas aeruginosa GF31with the protein recovery of 26.6%. The molecular mass of the enzyme was determined to be 53 kDa. The optimum temperature for the activity was surprisingly 60 °C, and moreover, the purified enzyme showed a good pH stability, maintaining over 85% of its initial activity in the pH 5.0-9.0 range. Most of the common metal ions exhibited little influence on the activity except for Hg2+, Ag+, and Cu2+. After the complete gene sequence of the degrading enzyme was obtained by subcloning, sequence analyses as well as enzymatic properties demonstrated that the islolated enzyme should be an aminopeptidase. This is the first reported aminopeptidase for pyrethroid hydrolase, providing new potential enzyme resources for the degradation of this type of pesticide.

Biodegradation and extracellular enzymatic activities of Pseudomonas aeruginosa strain GF31 on ß-cypermethrin.

Pseudomonas aeruginosa strain GF31, isolated from a contaminated soil, can effectively degrade ß-cypermethrin (ß-CP), as well as fenpropathrin, fenvalerate, and cyhalothrin. The highest level of degradation (81.2 %) was achieved with the addition of peptone. Surprisingly, the enzyme responsible for degradation was mainly localized to the extracellular areas of the bacteria, in contrast to the other known pyrethroid-degrading enzymes, which are intracellular. Although intact bacterial cells function at about 30 °C for biodegradation, similar to other degrading strains, the crude extracellular extract of strain GF31 remained biologically active at 60 °C. Moreover, the extract fraction showed good storage stability, maintaining >50 % of its initial activity following storage at 25 °C for at least 20 days. Significant differences in the characteristics of the crude GF31 extracellular extract compared with the known pyrethroid-degrading enzymes indicate the presence of a novel pyrethroid-degrading enzyme. Furthermore, the identification of 3-phenoxybenzoic acid and 2,2-dimethylcyclopropanecarboxylate from the degradation products suggests the possibility that ß-CP degradation by both the strain and the crude extracellular fraction is achieved through a hydrolysis pathway. Further degradation of these two metabolites may lead to the development of an efficient method for the mineralization of these types of pollutants.

A novel non-hydrolytic protein from Pseudomonas oryzihabitans enhances the enzymatic hydrolysis of cellulose.

Several kinds of protein such as the expansin, expansin-like proteins and LPMOs (lytic polysaccharide monooxygenases) are known to exert enhancement effects on cellulase activity. In this study, a novel cellulase synergistic protein named POEP1 was purified from the culture filtrate of Pseudomonas oryzihabitans CGMCC 6169, and was homogeneous on SDS-PAGE with a molecular weight of 60kDa. Mass spectrometry analysis indicated that it was an unknown protein without sequence similarity to the expansin and expansin-like proteins. Evaluation of the enzymatic hydrolysis of filter paper revealed that POEP1 had no cellulase activity but displayed high synergistic activity of 364% at a cellulase concentration of 0.1FPU/g of filter paper. When a mixture containing 0.6FPU cellulase and 700µg POEP1 per g of cellulose was evaluated, the maximal sugar yield was achieved, which was 2.2-fold greater than that with the cellulase alone. POEP1 was found to have functional similarity to the expansin and expansin-like proteins, which could decrease both the hydrogen-bond intensity and crystallinity, and cause the filter paper disruption. This study provided evidence for the existence of novel bacterial proteins in nature serving the same function as expansin and expansin-like proteins.