Uptake and Biotransformation of Guvermectin in Three Crops after In Vivo and In Vitro Exposure.

Guvermectin, as a novel nucleoside-like biopesticide, could increase the rice yield excellently, but the potential environmental behaviors remain unclear, which pose potential health risks. Therefore, the uptake and biotransformation of guvermectin in three types of crops (rice, lettuce, and carrot) were first evaluated with a hydroponic system. Guvermectin could be rapidly absorbed and reached equilibrium in roots (12-36 h) and shoots (24-60 h) in three plants, and guvermectin was also vulnerable to dissipation in roots (t1/2 1.02-3.65 h) and shoots (t1/2 9.30-17.91 h). In addition, 8 phase I and 2 phase II metabolites, transformed from guvermectin degradation in vivo and in vitro exposure, were identified, and one was confirmed as psicofuranine, which had antibacterial and antitumor properties; other metabolites were nucleoside-like chemicals. Molecular simulation and quantitative polymerase chain reaction further demonstrated that guvermectin was metabolized by the catabolism pathway of an endogenous nucleotide. Guvermectin had similar metabolites in three plants, but the biotransformation ability had a strong species dependence. In addition, all the metabolites exhibit neglectable toxicities (bioconcentration factor <2000 L/kg b.w., LC50,rat > 5000 mg/kg b.w.) by prediction. The study provided valuable evidence for the application of guvermectin and a better understanding of the biological behavior of nucleoside-like pesticides.

Identification of New Hepatic Metabolites of Miconazole by Biological and Electrochemical Methods Using Ultra-High-Performance Liquid Chromatography Combined with High-Resolution Mass Spectrometry.

The main objective of this study was to investigate the metabolism of miconazole, an azole antifungal drug. Miconazole was subjected to incubation with human liver microsomes (HLM) to mimic phase I metabolism reactions for the first time. Employing a combination of an HLM assay and UHPLC-HRMS analysis enabled the identification of seven metabolites of miconazole, undescribed so far. Throughout the incubation with HLM, miconazole underwent biotransformation reactions including hydroxylation of the benzene ring and oxidation of the imidazole moiety, along with its subsequent degradation. Additionally, based on the obtained results, screen-printed electrodes (SPEs) were optimized to simulate the same biotransformation reactions, by the use of a simple, fast, and cheap electrochemical method. The potential toxicity of the identified metabolites was assessed using various in silico models.

Continuous selenite biotransformation and biofuel production by marine diatom in the presence of fulvic acid.

In this study, the biochemical response of Phaeodactylum tricornutum to varying concentrations of inorganic selenium (Se) was investigated. It was observed that, when combined with fulvic acid, P. tricornutum exhibited enhanced uptake and biotransformation of inorganic Se, as well as increased microalgal lipid biosynthesis. Notably, when subjected to moderate (5 and 10 mg/L) and high (20 and 40 mg/L) concentrations of selenite under fulvic acid treatment, there was a discernible redirection of carbon flux towards lipogenesis and protein biosynthesis from carbohydrates. In addition, the key parameters of microalgae-based biofuels aligned with the necessary criteria outlined in biofuel regulations. Furthermore, the Se removal capabilities of P. tricornutum, assisted by fulvic acid, were coupled with the accumulation of substantial amounts of organic Se, specifically SeCys. These findings present a viable and successful approach to establish a microalgae-based system for Se uptake and biotransformation.

Effects of nitrate reduction on the biotransformation of 1H-1,2,4-triazole: Mechanism and community evolution.

Due to the refractory of 1 H-1,2,4-triazole (TZ), conventional anaerobic biological treatment technology is usually restricted by low removal efficiency and poor system stability. In this study, TZ biodegradation and nitrate reduction was coupled to improve the removal efficiency of TZ from polluted wastewater. Batch assay was performed with pure culture strain Raoultella sp. NJUST42, which was reported to have the capability to degrade TZ in our previous study. Based on batch assay result, complete removal of TZ could be achieved in the presence of nitrate, whereas only 50% of TZ could be removed in the control system. Long-term stability experiment indicated that the relative abundance of microorganisms (Bacteroidetes_vadinHA17, Georgenia, Anaerolinea, etc) was obviously enhanced under nitrate reduction condition. During long-term period, major intermediates for TZ biodegradation such as [1,2,4]Triazolidine-3,5-diol, hydrazine dibasic carboxylic acid and carbamic acid were detected. A novel TZ biotransformation approach via hydration, TZ-ring cleavage, deamination and oxidation was speculated. PICRUSt1 and KEGG pathway analyses indicated that hydration (dch), oxidation (adhD, oah, pucG, fdhA) of TZ and nitrate reduction (Nar, napA, nrfA, nirBK, norB, nosZ) were significantly enhanced in the presence of nitrate. Moreover, the significant enrichment of TCA cycle (gab, sdh, fum, etc.) indicated that carbon and energy metabolism were facilitated with the addition of nitrate, thus improved TZ catabolism. The proposed mechanism demonstrated that TZ biodegradation coupled with nitrate reduction would be a promising approach for efficient treatment of wastewater contaminated by TZ.

Mechanism of tartaric acid mediated dissipation and biotransformation of tetrabromobisphenol A and its derivatives in soil.

Biotransformation is a major dissipation process of tetrabromobisphenol A and its derivatives (TBBPAs) in soil. The biotransformation and ultimate environmental fate of TBBPAs have been widely studied, yet the effect of root exudates (especially low-molecular weight organic acids (LMWOAs)) on the fate of TBBPAs is poorly documented. Herein, the biotransformation behavior and mechanism of TBBPAs in bacteriome driven by LMWOAs were comprehensively investigated. Tartaric acid (TTA) was found to be the main component of LMWOAs in root exudates of Helianthus annus in the presence of TBBPAs, and was identified to play a key role in driving shaping bacteriome. TTA promoted shift of the dominant genus in soil bacteriome from Saccharibacteria_genera_incertae_sedis to Gemmatimonas, with a noteworthy increase of 24.90-34.65% in relative abundance of Gemmatimonas. A total of 28 conversion products were successfully identified, and ß-scission was the principal biotransformation pathway for TBBPAs. TTA facilitated the emergence of novel conversion products, including 2,4-dibromophenol, 3,5-dibromo-4-hydroxyacetophenone, para-hydroxyacetophenone, and tribromobisphenol A. These products were formed via oxidative skeletal cleavage and debromination pathways. Additionally, bisphenol A was observed during the conversion of derivatives. This study provides a comprehensive understanding about biotransformation of TBBPAs driven by TTA in soil bacteriome, offering new insights into LMWOAs-driven biotransformation mechanisms.

In vitro and in vivo screening of bacterial species from contaminated soil for heavy metal biotransformation activity.

Heavy metals (HMs) are widely used in various industries. High concentrations of HMs can be severely toxic to plants, animals and humans. Microorganism-based bioremediation has shown significant potential in degrading and detoxifying specific HM contaminants. In this study, we cultivated a range of bacterial strains in liquid and solid nutrient medium containing different concentrations of different HMs to select and analyze bacteria capable of transforming HMs. The bacterial strains most resistant to selected HMs and exhibiting the ability to remove HMs from contaminated soils were identified. Then, the bacterial species capable of utilizing HMs in soil model experiments were selected, and their ability to transform HMs was evaluated. This study has also generated preliminary findings on the use of plants for further removal of HMs from soil after microbial bioremediation. Alcaligenes faecalis, Delftia tsuruhatensis and Stenotrophomonas sp. were selected for their ability to grow in and utilize HM ions at the maximum permissible concentration (MPC) and two times the MPC. Lysinibacillus fusiformis (local microflora) can be used as a universal biotransformation tool for many HM ions. Brevibacillus parabrevis has potential for the removal of lead ions, and Brevibacillus reuszeri and Bacillus safensis have potential for the removal of arsenic ions from the environment. The bacterial species have been selected for bioremediation to remove heavy metal ions from the environment.

In Silico and Chromatographic Methods for Analysis of Biotransformation of Prospective Neuroprotective Pyrrole-Based Hydrazone in Isolated Rat Hepatocytes.

In the current study, chromatographic and in silico techniques were applied to investigate the biotransformation of ethyl 5-(4-bromophenyl)-1-(2-(2-(2-hydroxybenzylidene) hydrazinyl)-2-oxoethyl)-2-methyl-1H-pyrrole-3-carboxylate (11b) in hepatocytic media. The initial chromatographic procedure was based on the employment of the conventional octadecyl stationary phase method for estimation of the chemical stability. Subsequently, a novel and rapid chromatographic approach based on a phenyl-hexyl column was developed, aiming to separate the possible metabolites. Both methods were performed on a Dionex 3000 ThermoScientific (ACM 2, Sofia, Bulgaria) device equipped with a diode array detector set up at 272 and 279 nm for analytes detection. An acetonitrile: phosphate buffer of pH 3.5: methanol (17:30:53 v/v/v) was eluted isocratically as a mobile phase with a 1 mL/min flow rate. A preliminary purification from the biological media was achieved by protein precipitation with methanol. A validation procedure was carried out, where the method was found to correspond to all ICH (Q2) and M10 set criteria. Additionally, an in silico-based approach with the online server BioTransformer 3.0 was applied in an attempt to predict the possible metabolites of the title compound 11b. It was hypothesized that four CYP450 isoforms (1A2, 2C9, 3A4, and 2C8) were involved in the phase I metabolism, resulting in the formation of 12 metabolites. Moreover, docking studies were conducted to evaluate the formation of stable complexes between 11b and the aforementioned isoforms. The obtained data indicated three metabolites as the most probable products, two of which (M9_11b and M10_11b) were synthesized by a classical approach for verification. Finally, liquid chromatography with a mass detector was implemented for comprehensive and summarized analysis, and the obtained results revealed that the metabolism of the 11b proceeds possibly with the formation of glucuronide and glycine conjugate of M11_11b.

ATP-free in vitro biotransformation of starch-derived maltodextrin into poly-3-hydroxybutyrate via acetyl-CoA.

In vitro biotransformation (ivBT) facilitated by in vitro synthetic enzymatic biosystems (ivSEBs) has emerged as a highly promising biosynthetic platform. Several ivSEBs have been constructed to produce poly-3-hydroxybutyrate (PHB) via acetyl-coenzyme A (acetyl-CoA). However, some systems are hindered by their reliance on costly ATP, limiting their practicality. This study presents the design of an ATP-free ivSEB for one-pot PHB biosynthesis via acetyl-CoA utilizing starch-derived maltodextrin as the sole substrate. Stoichiometric analysis indicates this ivSEB can self-maintain NADP+/NADPH balance and achieve a theoretical molar yield of 133.3%. Leveraging simple one-pot reactions, our ivSEBs achieved a near-theoretical molar yield of 125.5%, the highest PHB titer (208.3 mM, approximately 17.9 g/L) and the fastest PHB production rate (9.4 mM/h, approximately 0.8 g/L/h) among all the reported ivSEBs to date, and demonstrated easy scalability. This study unveils the promising potential of ivBT for the industrial-scale production of PHB and other acetyl-CoA-derived chemicals from starch.

Potential routes of plastics biotransformation involving novel plastizymes revealed by global multi-omic analysis of plastic associated microbes.

Despite increasing efforts across various disciplines, the fate, transport, and impact of synthetic plastics on the environment and public health remain poorly understood. To better elucidate the microbial ecology of plastic waste and its potential for biotransformation, we conducted a large-scale analysis of all publicly available meta-omic studies investigating plastics (n = 27) in the environment. Notably, we observed low prevalence of known plastic degraders throughout most environments, except for substantial enrichment in riverine systems. This indicates rivers may be a highly promising environment for discovery of novel plastic bioremediation products. Ocean samples associated with degrading plastics showed clear differentiation from non-degrading polymers, showing enrichment of novel putative biodegrading taxa in the degraded samples. Regarding plastisphere pathogenicity, we observed significant enrichment of antimicrobial resistance genes on plastics but not of virulence factors. Additionally, we report a co-occurrence network analysis of 10 + million proteins associated with the plastisphere. This analysis revealed a localized sub-region enriched with known and putative plastizymes-these may be useful for deeper investigation of nature's ability to biodegrade man-made plastics. Finally, the combined data from our meta-analysis was used to construct a publicly available database, the Plastics Meta-omic Database (PMDB)-accessible at plasticmdb.org. These data should aid in the integrated exploration of the microbial plastisphere and facilitate research efforts investigating the fate and bioremediation potential of environmental plastic waste.

Biotransformation of fluorinated drugs and xenobiotics by the model fungus Cunninghamella elegans.

Some species of the genus Cunninghamella (C. elegans, C. echinulata and C. blaskesleeana) produce the same phase I and phase II metabolites when incubated with xenobiotics as mammals, and thus are considered microbial models of mammalian metabolism. This had made these fungi attractive for metabolism studies with drugs, pesticides and environmental pollutants. As a substantial proportion of pharmaceuticals and agrochemicals are fluorinated, their biotransformation has been studied in Cunninghamella fungi and C. elegans in particular. This article details the methods employed for cultivating the fungi in planktonic and biofilm cultures, and extraction and analysis of fluorinated metabolites. Furthermore, protocols for the heterologous expression of Cunninghamella cytochromes P450 (CYPs), which are the enzymes associated with phase I metabolism, are described.

Insight into chemically reactive metabolites of aliphatic amine pollutants: A de novo prediction strategy and case study of sertraline.

The uncommon metabolic pathways of organic pollutants are easily overlooked, potentially leading to idiosyncratic toxicity. Prediction of their biotransformation associated with the toxic effects is the very purpose that this work focuses, to develop a de novo method to mechanistically predict the reactive toxicity pathways of uncommon metabolites from start aliphatic amine molecules, which employed sertraline triggered by CYP450 enzymes as a model system, as there are growing concerns about the effects on human health posed by antidepressants in the aquatic environment. This de novo prediction strategy combines computational and experimental methods, involving DFT calculations upon sequential growth, in vitro and in vivo assays, dissecting chemically reactive mechanism relevant to toxicity, and rationalizing the fundamental factors. Significantly, desaturation and debenzylation-aromatization as the emerging metabolic pathways of sertraline have been elucidated, with the detection of DNA adducts of oxaziridine metabolite in mice, highlighting the potential reactive toxicity. Molecular orbital analysis supports the reactivity preference for toxicological-relevant C-N desaturation over N-hydroxylation of sertraline, possibly extended to several other aliphatic amines based on the Bell-Evans-Polanyi principle. It was further validated toward some other wide-concerned aliphatic amine pollutants involving atrazine, ε-caprolactam, 6PPD via in silico and in vitro assays, thereby constituting a complete path for de novo prediction from case study to general applications.

Efficient biotransformation of naringenin to naringenin α-glucoside, a novel α-glucosidase inhibitor, by amylosucrase from Deinococcus wulumuquiensis.

Amylosucrase (ASase) efficiently biosynthesizes α-glucoside using flavonoids as acceptor molecules and sucrose as a donor molecule. Here, ASase from Deinococcus wulumuqiensis (DwAS) biosynthesized more naringenin α-glucoside (NαG) with sucrose and naringenin as donor and acceptor molecules, respectively, than other ASases from Deinococcus sp. The biotransformation rate of DwAS to NαG was 21.3% compared to 7.1-16.2% for other ASases. Docking simulations showed that the active site of DwAS was more accessible to naringenin than those of others. The 217th valine in DwAS corresponded to the 221st isoleucine in Deinococcus geothermalis AS (DgAS), and the isoleucine possibly prevented naringenin from accessing the active site. The DwAS-V217I mutant had a significantly lower biosynthetic rate of NαG than DwAS. The kcat/Km value of DwAS with naringenin as the donor was significantly higher than that of DgAS and DwAS-V217I. In addition, NαG inhibited human intestinal α-glucosidase more efficiently than naringenin.

Unveiling behaviors of 8:2 fluorotelomer sulfonic acid (8:2 FTSA) in Arabidopsis thaliana: Bioaccumulation, biotransformation and molecular mechanisms of phytotoxicity.

8:2 fluorotelomer sulfonic acid (8:2 FTSA) has been commonly detected in the environment, but its behaviors in plants are not sufficiently known. Here, the regular and multi-omics analyses were used to comprehensively investigate the bioaccumulation, biotransformation, and toxicity of 8:2 FTSA in Arabidopsis thaliana. Our results demonstrated that 8:2 FTSA was taken up by A. thaliana roots and translocated to leaves, stems, flowers, and seeds. 8:2 FTSA could be successfully biotransformed to several intermediates and stable perfluorocarboxylic acids (PFCAs) catalyzed by plant enzymes. The plant revealed significant growth inhibition and oxidative damage under 8:2 FTSA exposure. Metabolomics analysis showed that 8:2 FTSA affected the porphyrin and secondary metabolisms, resulting in the promotion of plant photosynthesis and antioxidant capacity. Transcriptomic analysis indicated that differentially expressed genes (DEGs) were related to transformation and transport processes. Integrative transcriptomic and metabolomic analysis revealed that DEGs and differentially expressed metabolites (DEMs) in plants were predominantly enriched in the carbohydrate metabolism, amino acid metabolism, and lipid metabolism pathways, resulting in greater energy consumption, generation of more nonenzymatic antioxidants, alteration of the cellular membrane composition, and inhibition of plant development. This study provides the first insights into the molecular mechanisms of 8:2 FTSA stress response in plants.

Transcriptomics aids in uncovering the metabolic shifts and molecular machinery of Schizochytrium limacinum during biotransformation of hydrophobic substrates to docosahexaenoic acid.

BACKGROUND: Biotransformation of waste oil into value-added nutraceuticals provides a sustainable strategy. Thraustochytrids are heterotrophic marine protists and promising producers of omega (ω) fatty acids. Although the metabolic routes for the assimilation of hydrophilic carbon substrates such as glucose are known for these microbes, the mechanisms employed for the conversion of hydrophobic substrates are not well established. Here, thraustochytrid Schizochytrium limacinum SR21 was investigated for its ability to convert oils (commercial oils with varying fatty acid composition and waste cooking oil) into ω-3 fatty acid; docosahexaenoic acid (DHA). RESULTS: Within 72 h SR21 consumed ~ 90% of the oils resulting in enhanced biomass (7.5 g L- 1) which was 2-fold higher as compared to glucose. Statistical analysis highlights C16 fatty acids as important precursors of DHA biosynthesis. Transcriptomic data indicated the upregulation of multiple lipases, predicted to possess signal peptides for secretory, membrane-anchored and cytoplasmic localization. Additionally, transcripts encoding for mitochondrial and peroxisomal ß-oxidation along with acyl-carnitine transporters were abundant for oil substrates that allowed complete degradation of fatty acids to acetyl CoA. Further, low levels of oxidative biomarkers (H2O2, malondialdehyde) and antioxidants were determined for hydrophobic substrates, suggesting that SR21 efficiently mitigates the metabolic load and diverts the acetyl CoA towards energy generation and DHA accumulation. CONCLUSIONS: The findings of this study contribute to uncovering the route of assimilation of oil substrates by SR21. The thraustochytrid employs an intricate crosstalk among the extracellular and intracellular molecular machinery favoring energy generation. The conversion of hydrophobic substrates to DHA can be further improved using synthetic biology tools, thereby providing a unique platform for the sustainable recycling of waste oil substrates.

Using Network Analysis and Predictive Functional Analysis to Explore the Fluorotelomer Biotransformation Potential of Soil Microbial Communities.

Microbial transformation of per- and polyfluoroalkyl substances (PFAS), including fluorotelomer-derived PFAS, by native microbial communities in the environment has been widely documented. However, few studies have identified the key microorganisms and their roles during the PFAS biotransformation processes. This study was undertaken to gain more insight into the structure and function of soil microbial communities that are relevant to PFAS biotransformation. We collected 16S rRNA gene sequencing data from 8:2 fluorotelomer alcohol and 6:2 fluorotelomer sulfonate biotransformation studies conducted in soil microcosms under various redox conditions. Through co-occurrence network analysis, several genera, including Variovorax, Rhodococcus, and Cupriavidus, were found to likely play important roles in the biotransformation of fluorotelomers. Additionally, a metagenomic prediction approach (PICRUSt2) identified functional genes, including 6-oxocyclohex-1-ene-carbonyl-CoA hydrolase, cyclohexa-1,5-dienecarbonyl-CoA hydratase, and a fluoride-proton antiporter gene, that may be involved in defluorination. This study pioneers the application of these bioinformatics tools in the analysis of PFAS biotransformation-related sequencing data. Our findings serve as a foundational reference for investigating enzymatic mechanisms of microbial defluorination that may facilitate the development of efficient microbial consortia and/or pure microbial strains for PFAS biotransformation.

Substrate promiscuity of xenobiotic-transforming hydrolases from stream biofilms impacted by treated wastewater.

Organic contaminants enter aquatic ecosystems from various sources, including wastewater treatment plant effluent. Freshwater biofilms play a major role in the removal of organic contaminants from receiving water bodies, but knowledge of the molecular mechanisms driving contaminant biotransformations in complex stream biofilm (periphyton) communities remains limited. Previously, we demonstrated that biofilms in experimental flume systems grown at higher ratios of treated wastewater (WW) to stream water displayed an increased biotransformation potential for a number of organic contaminants. We identified a positive correlation between WW percentage and biofilm biotransformation rates for the widely-used insect repellent, N,N-diethyl-meta-toluamide (DEET) and a number of other wastewater-borne contaminants with hydrolyzable moieties. Here, we conducted deep shotgun sequencing of flume biofilms and identified a positive correlation between WW percentage and metagenomic read abundances of DEET hydrolase (DH) homologs. To test the causality of this association, we constructed a targeted metagenomic library of DH homologs from flume biofilms. We screened our complete metagenomic library for activity with four different substrates, including DEET, and a subset thereof with 183 WW-related organic compounds. The majority of active hydrolases in the metagenomic library preferred aliphatic and aromatic ester substrates while, remarkably, only a single reference enzyme was capable of DEET hydrolysis. Of the 626 total enzyme-substrate combinations tested, approximately 5% were active enzyme-substrate pairs. Metagenomic DH family homologs revealed a broad substrate promiscuity spanning 22 different compounds when summed across all enzymes tested. We biochemically characterized the most promiscuous and active enzymes identified based on metagenomic analysis from uncultivated Rhodospirillaceae and Planctomycetaceae. In addition to characterizing new DH family enzymes, we exemplified a framework for linking metagenome-guided hypothesis generation with experimental validation. Overall, this study expands the scope of known enzymatic contaminant biotransformations for metagenomic hydrolases from WW-receiving stream biofilm communities.

Unravelling the role of the group 6 soluble di-iron monooxygenase (SDIMO) SmoABCD in alkane metabolism and chlorinated alkane degradation.

Soluble di-iron monooxygenases (SDIMOs) are multi-component enzymes catalysing the oxidation of various substrates. These enzymes are characterized by high sequence and functional diversity that is still not well understood despite their key role in biotechnological processes including contaminant biodegradation. In this study, we analysed a mutant of Rhodoccocus aetherivorans BCP1 (BCP1-2.10) characterized by a transposon insertion in the gene smoA encoding the alpha subunit of the plasmid-located SDIMO SmoABCD. The mutant BCP1-2.10 showed a reduced capacity to grow on propane, lost the ability to grow on butane, pentane and n-hexane and was heavily impaired in the capacity to degrade chloroform and trichloroethane. The expression of the additional SDIMO prmABCD in BCP1-2.10 probably allowed the mutant to partially grow on propane and to degrade it, to some extent, together with the other short-chain n-alkanes. The complementation of the mutant, conducted by introducing smoABCD in the genome as a single copy under a constitutive promoter or within a plasmid under a thiostreptone-inducible promoter, allowed the recovery of the alkanotrophic phenotype as well as the capacity to degrade chlorinated n-alkanes. The heterologous expression of smoABCD allowed a non-alkanotrophic Rhodococcus strain to grow on pentane and n-hexane when the gene cluster was introduced together with the downstream genes encoding alcohol and aldehyde dehydrogenases and a GroEL chaperon. BCP1 smoA gene was shown to belong to the group 6 SDIMOs, which is a rare group of monooxygenases mostly present in Mycobacterium genus and in a few Rhodococcus strains. SmoABCD originally evolved in Mycobacterium and was then acquired by Rhodococcus through horizontal gene transfer events. This work extends the knowledge of the biotechnologically relevant SDIMOs by providing functional and evolutionary insights into a group 6 SDIMO in Rhodococcus and demonstrating its key role in the metabolism of short-chain alkanes and degradation of chlorinated n-alkanes.

Biotransformation activities of fungal strain apiotrichum sp. IB-1 to ibuprofen and naproxen.

Ibuprofen (IBU) and naproxen (NPX), as widely prescribed non-steroidal anti-inflammatory drugs (NSAIDs), are largely produced and consumed globally, leading to frequent and ubiquitous detection in various aqueous environments. Previously, the microbial transformation of them has been given a little attention, especially with the isolated fungus. A yeast-like Apiotrichum sp. IB-1 has been isolated and identified, which could simultaneously transform IBU (5 mg/L) and NPX (2.5 mg/L) with maximum efficiencies of 95.77% and 88.31%, respectively. For mono-substrate, the transformation efficiency of IB-1 was comparable to that of co-removal conditions, higher than most of isolates so far. IBU was oxidized mainly through hydroxylation (m/z of 221, 253) and NPX was detoxified mainly via demethylation (m/z of 215) as shown by UPLC-MS/MS results. Based on transcriptome analysis, the addition of IBU stimulated the basic metabolism like TCA cycle. The transporters and respiration related genes were also up-regulated accompanied with higher expression of several dehydrogenase, carboxylesterase, dioxygenase and oxidoreductase encoding genes, which may be involved in the transformation of IBU. The main functional genes responsible for IBU and NPX transformation for IB-1 should be similar in view of previous studies, which needs further confirmation. This fungus would be useful for potential bioremediation of NSAIDs pollution and accelerate the discovery of functional oxidative genes and enzymes different from those of bacteria.

Evaluating efficacy of Pseudomonas sp. EN-4 to lower the toxic potential of 4-bromophenol and assessing its competency in simulated microcosm.

An indigenous bacterium Pseudomonas sp. EN-4 had been reported earlier for its ability to co-metabolise 4-bromophenol (4-BP), in presence of phenol (100 mg/L) as co-substrate. The present study was undertaken to validate the efficacy of biotransformation by comparing the toxicity profiles of untreated and EN-4 transformed samples of 4-BP, using both plant and animal model. The toxicity studies in Allium cepa (A. cepa) indicated to lowering of mitotic index (MI) from 12.77% (water) to 3.33% in A. cepa bulbs exposed to 4-BP + phenol, which reflects the cytotoxic nature of these compounds. However, the MI value significantly improves to 11.36% in its biologically treated counterpart, indicating normal cell growth. This was further supported by significant reduction in chromosomal aberrations in A. cepa root cells exposed to biologically treated samples of 4-BP as compared to untreated controls. The oxidative stress assessed by comparing the activity profiles of different marker enzymes showed that the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX) and guaiacol peroxidase (GPX) were reduced by 56%, 72%, and 37% respectively, in EN-4 transformed samples of 4-BP + phenol compared to its untreated counterpart. Similar trends were evident in the comet assay of fish (Channa punctatus) blood cells exposed to untreated and biologically treated samples of 4-BP. The comparative studies showed significant reduction in tail length (72.70%) and % tail intensity (56.15%) in fish blood cells exposed to EN-4 treated 4-BP + phenol, compared to its untreated counterpart. The soil microcosm studies validated the competency of the EN-4 cells to establish and transform 4-BP in soil polluted with 4-BP (20 mg/kg) and 4-BP + phenol (20 + 100 mg/kg). The isolate EN-4 achieved 98.08% transformation of 4-BP in non-sterile microcosm supplemented with phenol, indicating to potential of EN-4 cells to establish along with indigenous microflora.

Establishment of efficient 5-hydroxyvaleric acid production system by regenerating alpha-ketoglutaric acid and its application in poly(5-hydroxyvaleric acid) production.

5-hydroxyvaleric acid (5-HV) is a versatile C5 intermediate of bio-based high-value chemical synthesis pathways. However, 5-HV production faces a few shortcomings involving the supply of cofactors, especially α-ketoglutaric acid (α-KG). Herein, we established a two-cell biotransformation system by introducing L-glutamate oxidase (GOX) to regenerate α-KG. Additionally, the catalase KatE was adapted to inhibit α-KG degradation by the H2O2 produced during GOX reaction. We searched for the best combination of genes and vectors and optimized the biotransformation conditions to maximize GOX effectiveness. Under the optimized conditions, 5-HV pathway with GOX showed 1.60-fold higher productivity than that of without GOX, showing 11.3 g/L titer. Further, the two-cell system with GOX and KatE was expanded to produce poly(5-hydroxyvaleric acid) (P(5HV)), and it reached at 412 mg/L of P(5HV) production and 20.5% PHA contents when using the biotransformation supernatant. Thus, the two-cell biotransformation system with GOX can potentially give the practical and economic alternative of 5-HV production using bio-based methods. We also propose direct utilization of 5-HV from bioconversion for P(5HV) production.