Comparative analysis of SilA-laccase mediated degradation of ciprofloxacin, norfloxacin and ofloxacin and interpretation of the possible catalytic mechanism.

Fluoroquinolones (FQs) are the most commonly used antimicrobial drugs and regardless of their advantages in the healthcare sector, the pollution of these antimicrobial drugs in the environment has big concerns about human and environmental health. The presence of these antibiotic drugs even at the lowest concentrations in the environment has resulted in the emergence and spread of antibiotic resistance. Hence, it is necessary to remediate these pollutants from the environment. Previously alkaline laccase (SilA) from Streptomyces ipomoeae has been demonstrated to show degrading potentials against two of the FQs, Ciprofloxacin (CIP) and Norfloxacin (NOR); however, the molecular mechanism was not elucidated in detail. In this study, we have analyzed the possible molecular catalytic mechanism of FQ degrading SilA-laccase for the degradation of the FQs, CIP, NOR and Ofloxacin (OFL) using three-dimensional protein structure modeling, molecular docking and molecular dynamic (MD) studies. The comparative protein sequence analysis revealed the presence of tetrapeptide conserved catalytic motif, His102-X-His104-Gly105. After evaluating the active site of the enzyme in depth using CDD, COACH and S-site tools, we have identified the catalytic triad composed of three conserved amino acid residues, His102, Val103 and Tyr108 with which ligands interacted during the catalysis process. By analyzing the MD trajectories, it is revealed that the highest degradation potential of SilA is for CIP followed by NOR and OFL. Ultimately, this study provides the possible comparative catalytic mechanism for the degradation of CIP, NOR and OFL by the SilA enzyme.Communicated by Ramaswamy H. Sarma.

Combined toxic effects of nanoplastics and norfloxacin on antioxidant and immune genes in mussels.

Nanoplastics (NPs) and antibiotics (ABs) are two of the emerging marine contaminants that have drawn the most attention in recent years. Given the necessity of figuring out the effects of plastic and antibiotic contamination on marine organism life and population in the natural environment, it is essential to apply rapid and effective biological indicators to evaluate their comprehensive toxic effects. In this study, using mussel (Mytilus coruscus) as a model, we investigated the combined toxic effects of NP (80 nm polystyrene beads) and AB (Norfloxacin, NOR) at environmental-relevant concentrations on antioxidant and immune genes. In terms of the antioxidant genes, NPs significantly increased the relative expression of Cytochrome P450 3A-1 (CYP3A-1) under various concentrations of NOR conditions, but they only significantly increased the relative expression of CYP3A-2 in the high concentration (500 µg L-1 NOR) co-exposure group. In the NP-exposure group which exposed to no or low concentrations of NOR, nuclear factor erythroid 2-related factor 2 (Nrf2) was upregulated. In terms of the immune genes, interleukin-1 receptor-associated kinase (IRAK) -1 showed a significant increase in the low-concentration NOR group while a significant inhibition in the high-concentration NOR group. Due to the presence of NPs, exposure to NOR resulted in a significant increase in both IRAK-4 and heat shock protein (HSP) 70. Our findings indicate that polystyrene NPs can exacerbate the effects of NOR on the anti-oxidant and immune defense performance of mussels. This study delves into the toxic effects of NPs and ABs from a molecular perspective. Given the expected increase in environmental pollution due to NPs and ABs, future research is needed to investigate the potential synergistic effect of NPs and ABs on other organisms.

Synthesis, structural, characterization, antibacterial and antibiotic modifying activity, ADMET study, molecular docking and dynamics of chalcone (E)-1-(4-aminophenyl)-3-(4-nitrophenyl)prop-2-en-1-one in strains of Staphylococcus aureus carrying NorA and MepA efflux pumps.

Chalcones have an open chain flavonoid structure that can be obtained from natural sources or by synthesis and are widely distributed in fruits, vegetables, and tea. They have a simple and easy to handle structure due to the α-ß-unsaturated bridge responsible for most biological activities. The facility to synthesize chalcones combined with its efficient in combating serious bacterial infections make these compounds important agents in the fight against microorganisms. In this work, the chalcone (E)-1-(4-aminophenyl)-3-(4-nitrophenyl)prop-2-en-1-one (HDZPNB) was characterized by spectroscopy and electronic methods. In addition, microbiological tests were performed to investigate the modulator potential and efflux pump inhibition on S. aureus multi-resistant strains. The modulating effect of HDZPNB chalcone in association with the antibiotic norfloxacin, on the resistance of the S. aureus 1199 strain, resulted in increase the MIC. In addition, when HDZPNB was associated with ethidium bromide (EB), it caused an increase in the MIC value, thus not inhibiting the efflux pump. For the strain of S. aureus 1199B, carrying the NorA pump, the HDZPNB associated with norfloxacin showed no modulatory, and when the chalcone was used in association with EB, it had no inhibitory effect on the efflux pump. For the tested strain of S. aureus K2068, which carries the MepA pump, it can be observed that the chalcone together the antibiotic resulted in an increase the MIC. On the other hand, when chalcone was used in association with EB, it caused a decrease in bromide MIC, equal to the reduction caused by standard inhibitors. Thus, these results indicate that the HDZPNB could also act as an inhibitor of the S. aureus gene overexpressing pump MepA. The molecular docking reveals that chalcone has a good binding energies -7.9 for HDZPNB/MepA complexes, molecular dynamics simulations showed that Chalcone/MetA complexes showed good stability of the structure in an aqueous solution, and ADMET study showed that the chalcone has a good oral bioavailability, high passive permeability, low risk of efflux, low clearance rate and low toxic risk by ingestion. The microbiological tests show that the chalcone can be used as a possible inhibitor of the Mep A efflux pump.Communicated by Ramaswamy H. Sarma.

Functional Proteomics Analysis of Norfloxacin-Resistant Edwardsiella tarda.

Multidrug-resistant Edwardsiella tarda threatens both sustainable aquaculture and human health, but the control measure is still lacking. In this study, we adopted functional proteomics to investigate the molecular mechanism underlying norfloxacin (NOR) resistance in E. tarda. We found that E. tarda had a global proteomic shift upon acquisition of NOR resistance, featured with increased expression of siderophore biosynthesis and Fe3+-hydroxamate transport. Thus, either inhibition of siderophore biosynthesis with salicyl-AMS or treatment with another antibiotic, kitasamycin (Kit), which was uptake through Fe3+-hydroxamate transport, enhanced NOR killing of NOR-resistant E. tarda both in vivo and in vitro. Moreover, the combination of NOR, salicyl-AMS, and Kit had the highest efficacy in promoting the killing effects of NOR than any drug alone. Such synergistic effect not only confirmed in vitro and in vivo bacterial killing assays but also applicable to other clinic E. tarda isolates. Thus, our data suggest a proteomic-based approach to identify potential targets to enhance antibiotic killing and propose an alternative way to control infection of multidrug-resistant E. tarda.

Quantitative Proteomics Reveal the Inherent Antibiotic Resistance Mechanism against Norfloxacin Resistance in Aeromonas hydrophila.

Recently, the prevalence of Aeromonas hydrophila antibiotic-resistant strains has been reported in aquaculture, but its intrinsic antibiotic resistance mechanisms are largely unknown. In the present study, a label-free proteomics technology was used to compare the differential protein abundances in response to norfloxacin (NOR) stress in A. hydrophila. The results showed that there were 186 proteins decreasing and 220 proteins increasing abundances in response to NOR stress. Bioinformatics analysis showed that the differentially expressed proteins were enriched in several biological processes, such as sulfur metabolism and homologous recombination. Furthermore, the antibiotic sensitivity assays showed that the deletion of AHA_0904, cirA, and cysI significantly decreased the resistance against NOR, whereas ΔAHA_1239, ΔcysA, ΔcysD, and ΔcysN significantly increased the resistance against NOR. Our results provide insights into NOR resistance mechanisms and indicate that AHA_0904, cirA, AHA_1239, and sulfur metabolism may play important roles in NOR resistance in A. hydrophila.

The different toxicological effects and removal efficiencies of norfloxacin and sulfadiazine in culturing Arthrospira (Spirulina) platensis.

Norfloxacin (NFX) and sulfadiazine (SDZ) are two widely used antibiotics belonging to fluoroquinolone and sulfonamide groups, respectively, and have become the commonly detected micropollutants in aquatic environments. However, only few works have been conducted to investigate the highly probable inhibition of these antibiotic pollutants to Arthrospira platensis, which is an important species of cyanobacteria that is one of primary producers in aquatic ecosystems and should be remarkably sensitive to environmental pollutants due to its prokaryotic characteristics. Hence, the toxicological effects and removal efficiencies of NFX and SDZ in culturing A. platensis were studied by analyzing the biomass growth, photosynthetic pigments, primary biocomponents, and antibiotics concentration. The corresponding variations of these characteristics showed the higher sensitivity of A. platensis to NFX than to SDZ, indicating the specifically targeted effect of NFX on A. platensis, which could be confirmed in silico by the higher binding affinity of NFX with the critical enzyme. The obtained results illustrated the roles of NFX and SDZ on the growth of A. platensis, thus providing the great support in employing A. platensis to reduce hazards from contaminated water and recover biomass resources.

A comprehensive description of the TolC effect on the antimicrobial susceptibility profile in Enterobacter bugandensis.

Background: Enterobacter bugandensis is an emerging human pathogen in which multidrug resistant strains have been continuously isolated from various environments. Thus, this organism possesses the potential to pose challenges in human healthcare. However, the mechanisms, especially the efflux pumps, responsible for the multidrug resistance in E. bugandensis remain to be well elucidated. Methods: The Enterobacter strain CMCC(B) 45301 was specifically identified using whole genome sequencing. The specific CMCC(B) 45301 homologues of the TolC dependent efflux-pump genes characterized in Escherichia coli were identified. The tolC deletion mutant in CMCC(B) 45301 was constructed and subjected to susceptibility tests using 26 different antimicrobial agents, along with the wild type strain. The synergistic effects combining the Bacillus crude extract (BCE) and several other TolC-affected compounds against CMCC(B) 45301 were assayed. Results: We reclassified the Enterobacter CMCC(B) 45301 strain from species cloacae to bugandensis, on the basis of its whole genome sequence. We found that the CMCC(B) 45301 TolC, AcrAB, AcrD, AcrEF, MdtABC, EmrAB, and MacAB exhibit high similarity with their respective homologues in E. coli and Enterobacter cloacae. Our results for the susceptibility tests revealed that lacking tolC causes 4- to 256-fold decrease in the minimal inhibitory concentrations of piperacillin, gentamicin, kanamycin, tetracycline, norfloxacin, ciprofloxacin, chloramphenicol, and erythromycin against CMCC(B) 45301. In addition, the inhibition zones formed by cefuroxime, cefoperazone, amikacin, streptomycin, minocycline, doxycycline, levofloxacin, florfenicol, trimethoprim-sulfamethoxazole, azithromycin, lincomycin, and clindamycin for the tolC mutant were larger or more obvious than that for the parent. Our data suggested the important role played by TolC in CMCC(B) 45301 susceptibility to common antibiotic families covering ß-lactam, aminoglycoside, tetracycline, fluoroquinolone, phenicol, folate pathway antagonist, macrolide, and lincosamide. Deletion for tolC also increased the susceptibility of CMCC(B) 45301 to berberine hydrochloride and BCE, two natural product-based agents. Finally, we found that erythromycin, norfloxacin, and ciprofloxacin can potentiate the antibacterial activity of BCE against CMCC(B) 45301. Discussion: The present study elaborated the comprehensive TolC effect on the antimicrobial susceptibility profile in E. bugandensis, which might contribute to the development of more therapeutic options against this nosocomial pathogen.

A NiCoT family metal transporter of Mycobacterium tuberculosis (Rv2856/NicT) behaves as a drug efflux pump that facilitates cross-resistance to antibiotics.

Metals often act as a facilitator in the proliferation and persistence of antibiotic resistance. Efflux pumps play key roles in the co-selection of metal and antibiotic resistance. Here, we report the ability of a putative nickel/cobalt transporter (NiCoT family), Rv2856 or NicT of Mycobacterium tuberculosis (Mtb), to transport metal and antibiotics and identified some key amino acid residues that are important for its function. Ectopic expression of NicT in Escherichia coli CS109 resulted in the increase of intracellular nickel uptake. Additionally, enhanced tolerance towards several antibiotics (norfloxacin, sparfloxacin, ofloxacin, gentamicin, nalidixic acid and isoniazid) was observed with NicT overexpression in E. coli and Mycobacterium smegmatis. A comparatively lower intracellular accumulation of norfloxacin upon NicT overexpression than that of the cells without NicT indicated the involvement of NicT in an active efflux process. Although expression of NicT did not alter the sensitivity towards kanamycin, doxycycline, tetracycline, apramycin, neomycin and ethambutol, the presence of a sub-inhibitory dose of Ni2+ resulted in the manifestation of low-level tolerance towards these drugs. Further, substitution of four residues (H77I, D82I, H83L and D227I) in the conserved regions of NicT by isoleucine and leucine resulted in reduced to nearly complete loss of the transport function for both metals and antimicrobials. Therefore, the study suggests that nickel transporter Rv2856/NicT may actively export different drugs and the presence of nickel might drive the cross-resistance to some of the antibiotics.

Validation of a random Vibrio parahaemolyticus genomic library by selection of quinolone resistance in a heterologous host.

Vibrio parahaemolyticus is a shellfish-borne pathogen that is a highly prevalent causative agent of inflammatory gastroenteritis in humans. Genomic libraries have proven useful for the identification of novel gene functions in many bacterial species. In this study we prepared a library containing 40 kb fragments of randomly sheared V. parahaemolyticus genomic DNA and introduced this into Escherichia coli HB101 using a commercially available low copy cosmid system. In order to estimate coverage and suitability of the library and potentially identify novel antimicrobial resistance determinants, we screened for the acquisition of resistance to the fluoroquinolone norfloxacin - a phenotype exhibited by V. parahaemolyticus but not the heterologous E. coli host. Upon selection on solid medium containing norfloxacin, 0.52% of the library population was resistant, consistent with the selection of a single resistance locus. End-sequencing identified six distinct insert fragments. All clones displayed fourfold increased norfloxacin MIC compared with E. coli HB101 carrying an empty vector. The common locus contained within resistant clones included qnr, a previously described quinolone resistance gene. These results indicate that the library was unbiased, of sufficient coverage and that heterologous expression was possible. While we hope that this library proves useful for identifying the genetic determinants of complex phenotypes such as those related to virulence, not all norfloxacin resistance genes were detected in our screen. As such, we discuss the benefits and limitations of this approach for identifying the genetic basis of uncharacterized bacterial phenotypes.

Effects of long-term exposure of norfloxacin on the HPG and HPT axes in juvenile common carp.

Currently, there is a relatively lack of relevant research on the interference effect of quinolone antibiotics on the endocrine of aquatic animals. In this study, the toxicity of norfloxacin (NOR) on the endocrine system of juvenile common carp (Cyprinus carpio) was evaluated, as well as the hematocyte parameters. Specifically, two important endocrine axes were assessed: the hypothalamus-pituitary-thyroid (HPT) axis and hypothalamus-pituitary-gonadal (HPG) axis. Norfloxacin was used as a representative of quinolone antibiotics. According to the concentration of water pollution areas and considering the bad situation that may be caused by wastewater discharge, a control, 100 ng/L NOR, and 1 mg/L NOR treatment groups were set up. The juvenile carp, as the test animal, was subjected to an exposure experiment for 42 days. Thyroid hormones (T3 and T4) and related genes in HPT axis and sex hormones (11-ketotestosterone [11-KT] and progesterone [PROG]) and related genes in HPG axis and blood count are tested. It was found that the T4 iodine level and conversion process were enhanced after NOR treatment, which in turn led to the increase of T3 content and biological activity in the blood. One hundred nanograms per liter NOR can inhibit the level of sex hormones and inhibit the expression of HPG axis-related genes. In the 1 mg/L NOR treatment group, long-term exposure over a certain concentration range may lead to the development of adaptive mechanisms, making the changes in hormones and related genes insignificant. In conclusion, this study provides reference data for the endocrine interference of quinolone antibiotics on aquatic organisms, and has ecological significance for assessing the health of fish populations of quinolone antibiotics. However, the specific sites and mechanisms of action related to the effects of NOR on the endocrine system remain unclear and require further study.

In silico and in vitro evaluation of efflux pumps inhibition of α,ß-amyrin.

The use of the bacterial efflux pump mechanism to reduce the concentrations of antibiotics in the intracellular to the extracellular region is one of the main mechanisms by which bacteria acquire resistance to antibiotics. The present study aims to evaluate the antibacterial activity of the α,ß-amyrin mixture isolated from Protium heptaphyllum against the multidrug-resistant strains of Escherichia coli 06 and Staphylococcus aureus 10, and to verify the inhibition of the efflux resistance mechanisms against the strains of S. aureus 1199B and K2068, carrying the NorA and MepA efflux pumps, respectively. The α,ß-amyrin did not show clinically relevant direct bacterial activity. However, the α,ß-amyrin when associated with the gentamicin antibiotic presented synergistic effect against the multidrug-resistant bacterial strain of S. aureus 10. In strains with efflux pumps, α,ß-amyrin was able to inhibit the action of the efflux protein NorA against Ethidium Bromide. However, this inhibitory effect was not observed in the MepA efflux pump. In addition, when evaluating the effect of standard efflux pump inhibitors, clorptomazine and CCCP, α,ß-amyrin showed a decrease in MIC, demonstrating the presence of the efflux mechanism through synergism. Docking studies indicate that α, ß-amyrin have a higher affinity energy to MepA, and NorA than ciprofloxacin and norfloxacin. Also, α, ß-amyrin bind to the same region of the binding site as these antibiotics. It was concluded that the α, ß-amyrin has the potential to increase antibacterial activity with the association of antibiotics, together with the ability to be a strong candidate for an efflux pump inhibitor.Communicated by Ramaswamy H. Sarma.

Investigation of major amino acid residues of anti-norfloxacin monoclonal antibodies responsible for binding with fluoroquinolones.

It is important to understand the amino acid residues that govern the properties of the binding between antibodies and ligands. We studied the binding of two anti-norfloxacins, anti-nor 132 and anti-nor 155, and the fluoroquinolones norfloxacin, enrofloxacin, ciprofloxacin, and ofloxacin. Binding cross-reactivities tested by an indirect competitive enzyme-linked immunosorbent assay indicated that anti-nor 132 (22-100%) had a broader range of cross-reactivity than anti-nor 155 (62-100%). These cross-reactivities correlated with variations in the numbers of interacting amino acid residues and their positions. Molecular docking was employed to investigate the molecular interactions between the fluoroquinolones and the monoclonal antibodies. Homology models of the heavy chain and light chain variable regions of each mAb 3D structure were docked with the fluoroquinolones targeting the crucial part of the complementarity-determining regions. The fluoroquinolone binding site of anti-nor 155 was a region of the HCDR3 and LCDR3 loops in which hydrogen bonds were formed with TYR (H:35), ASN (H:101), LYS (H:106), ASN (L:92), and ASN (L:93). These regions were further away in anti-nor 132 and could not contact the fluoroquinolones. Another binding region consisting of HIS (L:38) and ASP (H:100) was found for norfloxacin, enrofloxacin, and ciprofloxacin, whereas only ASP (H:100) was found for ofloxacin.

Light modulates the effect of antibiotic norfloxacin on photosynthetic processes of Microcystis aeruginosa.

Norfloxacin is one of the widely used antibiotics, often detected in aquatic ecosystems, and difficultly degraded in the environment. However, how norfloxacin affects the photosynthetic process of freshwater phytoplankton is still largely unknown, especially under varied light conditions. In this study, we investigated photosynthetic mechanisms of Microcystis aeruginosa in responses to antibiotic norfloxacin (0-50 µg/L) for 72 h under low (LL; 50 µmol photons m-2 s-1) and high (HL; 250 µmol photons m-2 s-1) growth light regimes. We found that environmentally related concentrations of norfloxacin inhibited the growth rate and operational quantum yield of photosynthesis system II (PSII) of M. aeruginosa more under HL than under LL, suggesting HL increased the toxicity of norfloxacin to M. aeruginosa. Further analyses showed that norfloxacin deactivated PSII reaction centers under both growth light regimes with increased minimal fluorescence yields only under HL, suggesting that norfloxacin not only damaged reaction centers of PSII, but also inhibited energy transfer among phycobilisomes in M. aeruginosa under HL. However, non-photosynthetic quenching decreased in the studied species by norfloxacin exposure under both growth light regimes, suggesting that excess energy might not be efficiently dissipated as heat. Also, we found that reactive oxygen species (ROS) content increased under norfloxacin treatments with a higher ROS content under HL compared to LL. In addition, HL increased the absorption of norfloxacin by M. aeruginosa, which could partly explain the high sensitivity to norfloxacin of M. aeruginosa under HL. This study firstly reports that light can strongly affect the toxicity of norfloxacin to M. aeruginosa, and has vitally important implications for assessing the toxicity of norfloxacin to aquatic microorganisms.

Synthesis of Novel Ternary Dual Z-scheme AgBr/LaNiO3/g-C3N4 Composite with Boosted Visible-Light Photodegradation of Norfloxacin.

Promoting the separation of photogenerated charges and enhanced optical absorption capacity is the main means to modify photocatalytic capacities to advance semiconductor photocatalyst applications. For the first time, a novel ternary photocatalyst for dual Z-scheme system AgBr/LaNiO3/g-C3N4 (ALG) was prepared via a modest ultrasound-assisted hydrothermal method. The results indicated that LaNiO3 nanoballs and AgBr nanoparticles were successfully grown on the surface of g-C3N4 nanosheets. A dual Z-scheme photocatalytic reaction system could be constructed based on the energy band matching within AgBr, LaNiO3 and g-C3N4. Metallic Ag during the photocatalytic reaction process acted as the active electrons transfer center to enhance the photocatalytic charge pairs separation. The chemical composition of ALG was optimized and composites with 3% AgBr, 30% LaNiO3 and 100% g-C3N4 which was noted as 3-ALG displayed the best photocatalytic performance. A total of 92% of norfloxacin (NOR) was photodegraded within two hours over ALG and the photodegradation rate remained >90% after six cycles. The main active species during the degradation course were photogenerated holes, superoxide radical anion and hydroxyl radical. A possible mechanism was proposed based on the synergetic effects within AgBr, LaNiO3 and g-C3N4. This work would offer a credible theoretical basis for the application of dual Z-scheme photocatalysts in environment restoration.

Shuttle-like CeO2/g-C3N4 composite combined with persulfate for the enhanced photocatalytic degradation of norfloxacin under visible light.

In this work, the shuttle-like CeO2 modified g-C3N4 composite was synthesized and was combined with persulfate (PS) for the efficient photocatalytic degradation of norfloxacin (NOR) under visible light. Scanning and transmission electron microscopy (SEM and TEM), X-ray diffraction (XRD), UV-vis diffuse reflectance spectroscopy (DRS) and photoluminescence (PL) emission spectra were used to characterize the structural and optical properties of the as-prepared catalysts. Active species trapping experiments demonstrated that additional sulfate radicals (·SO4-) formed upon the addition of PS which could cooperate with superoxide radicals (O2-), holes (h+) and hydroxyl radicals (OH) to decompose NOR. Singlet oxygen (1O2) was also formed during the reaction and acted as an important active species. The degradation products of NOR were also identified and analyzed by using LC-MS technology, and the possible degradation mechanism and pathways were proposed and discussed. This work indicated that the shuttle-like CeO2 modified g-C3N4 coupled with PS displayed promising applications in the field of pharmaceutical wastewater purification.

Molecular Modification of Fluoroquinolone-Biodegrading Enzymes Based on Molecular Docking and Homology Modelling.

To improve the biodegradation efficiency of fluoroquinolone antibiotics during sewage treatment, fluoroquinolone aerobic, anaerobic and facultative degrading enzymes for fluoroquinolone degradation were modified by molecular docking and homology modelling. First, amino acid residues of the binding sites of degrading enzymes for the target fluoroquinolones ciprofloxacin (CIP), norfloxacin (NOR) and ofloxacin (OFL) were analysed by the molecular docking method. The hydrophobic amino acid residues within 5 Å of the target fluoroquinolone molecules were selected as the modification sites. The hydrophobic amino acid residues at the modified sites were replaced by the hydrophilic amino acid residues, and 150 amino acid sequence modification schemes of the degrading enzymes were designed. Subsequently, a reconstruction scheme of the degrading enzyme amino acid sequence reconstruction scheme was submitted to the SWISS-MODEL server and a selected homology modelling method was used to build a new structure of the degrading enzyme. At the same time, the binding affinities between the novel degrading enzymes and the target fluoroquinolones (represented by the docking scoring function) were evaluated by the molecular docking method. It was found that the novel enzymes can simultaneously improve the binding affinities for the three target fluoroquinolones, and the degradation ability of the six modification schemes was increased by more than 50% at the same time. Among the novel enzymes, the affinity effect of the novel anaerobic enzyme (6-1) with CIP, NOR and OFL was significantly increased, with increases of 129.24%, 165.06% and 169.59%, respectively, followed by the facultative enzyme and aerobic enzyme. In addition, the designed degrading enzymes had certain selectivity for the degradation of the target quinolone. Among the novel enzymes, the binding affinities of the novel anaerobic enzyme (6-3) and CIP, the novel aerobic enzyme (3-6) and NOR, and the novel facultative enzyme (13-6) and OFL were increased by 149.71%, 178.57% and 297.12% respectively. Calculations using the Gaussian09 software revealed that the degradation reaction barrier of the novel degrading enzyme (7-1) and CIP NOR and OFL decreased by 37.65 kcal·mol-1, 6.28 kcal·mol-1 and 6.28 kcal·mol-1, respectively, which would result in efficient degradation of the target fluoroquinolone molecules. By analysing the binding affinity of the degrading enzymes before and after the modification with methanol, it was further speculated that the degradation effect of the modified aerobic degrading enzymes on organic matter was lower than that before the modification, and the increase or decrease in the degradation effect was less than 10%. The mechanism analysis found that the interaction between the modified amino acid residues of the degrading enzymes and the fluoroquinolone molecules increased. The average distance between the amino acid residues and the fluoroquinolone molecules represented a comprehensive affinity effect, and its value was positively correlated with the degradation effect of the novel degrading enzymes.

Bio-Electron-Fenton (BEF) process driven by sediment microbial fuel cells (SMFCs) for antibiotics desorption and degradation.

A sediment microbial fuel cell electro-Fenton (SMFC-E-Fenton) system was proposed in this study by utilizing the biological electrons produced from an SMFC to power an E-Fenton process. Antibiotics with different absorbability in both aqueous and solid phase can be removed by this system at room temperature and pressure condition, without external power or other chemical reagents. γ-FeOOH graphene polyacrylamide carbonized aerogel (γ-FeOOH GPCA) with high electrochemically active surface area (EASA), good conductivity and stable electrochemical activity were used as the cathode. After 40 h treatment, the total degradation rate of sulfamethoxazole (SMX) and norfloxacin (NOR) were 97.4 ±â€¯2.9% and 96.1 ±â€¯3.0%, respectively. Compared with the sludge digestion system, the residual SMX and NOR in sludge declined from 10.2 ±â€¯1.5% to 1.1 ±â€¯1.2% and from 31.3 ±â€¯1.8% to 3.1 ±â€¯1.3%, respectively. The E-Fenton process can also promote electricity production and sludge reduction efficiency of SMFC, as the maximum power density SMFC-E-Fenton system reached 472.21 ±â€¯11.5 mW m-2 and 431.39 ±â€¯15.6 mWm-2, respectively. 6.2 ±â€¯0.3% and 5.7 ±â€¯0.8% of the initial sludge was reduced while treating SMX and NOR.

Small-Molecule Permeation across Membrane Channels: Chemical Modification to Quantify Transport across OmpF.

Biological channels facilitate the exchange of molecules across membranes, but general tools to quantify transport are missing. Electrophysiology is the method of choice to study the functional properties of channels. However, analyzing the current fluctuation of channels typically does not identify successful transport, that is, distinguishing translocation from binding. To distinguish both processes, we added an additional barrier at the channel exit acting as a molecular counter. To identify permeation, we compare the molecule residence time in the native channel with one that is chemically modified at the exit. We use the well-studied outer membrane channel from E. coli, OmpF. Position 181, which is below the constriction region, was subsequently mutated into cysteine (E181C) in an otherwise cysteine-free system, then functionalized by covalent binding with one of the two blockers MTSES or GLT. We measured the passage of model peptides, mono-, tri-, hepta-arginine and of norfloxacin, as an example for antibiotic permeation.

Effects of individual and combined zinc oxide nanoparticle, norfloxacin, and sulfamethazine contamination on sludge anaerobic digestion.

This work investigated the individual and combined effects of zinc oxide, norfloxacin, and sulfamethazine on sludge anaerobic digestion-associated methane production, protein and carbohydrate metabolism, and microbial diversity. Norfloxacin and sulfamethazine (500 mg/kg) did not inhibit methane production, but inhibited its production rate. Zinc oxide nanoparticles with antibiotics inhibited hydrolysis, fermentation, and methanogenesis over varying digestion periods. Complex pollution had a greater impact on methane production than zinc oxide alone, with acute, synergistic toxicity to methanogenesis over short periods. Complex pollution also had varying effects on bacterial and archaeal communities during digestion. These results aid understanding of the toxicity of emerging contaminants in sludge digestion, with the potential to improve pollution removal and reduce associated risks.

Investigation of the binding sites and orientation of Norfloxacin on bovine serum albumin by surface enhanced Raman scattering and molecular docking.

Norfloxacin (NFX) is an antibacterial agent belonging to the fluoroquinolone family of drugs, known to bind bovine serum albumin (BSA). Surface-enhanced Raman scattering (SERS) and fluorescence spectroscopy in combination with molecular docking were explored to investigate the binding interaction between NFX with Bovine serum albumin (BSA) at a physiological condition. This study focused on identifying the binding site and relevant interaction mechanisms between NFX and BSA. Spectrophotometric titration with molecular docking results demonstrated that the binding site of NFX on BSA was located in sub-domain IIA. The principle binding site was identified within a hydrophobic cavity which is surrounded by the residues Leu 197, Arg 198, Ser 201, Ala 209, Trp 213, Ser 343, Leu 346, Lys 350, Ser 453, Leu 480, Val 481, and the binding force was mainly hydrophobic interaction and hydrogen bond interaction. In addition, the absorptive orientation of the NFX molecule on the colloidal surface underwent a set of changes during the process of NFX binding to BSA.