A Comprehensive Study to Determine the Residual Elimination Pattern of Major Metabolites of Amoxicillin-Sulbactam Hybrid Molecules in Rats by UPLC-MS/MS.

Amoxicillin and sulbactam are widely used in animal food compounding. Amoxicillin-sulbactam hybrid molecules are bicester compounds made by linking amoxicillin and sulbactam with methylene groups and have good application prospects. However, the residual elimination pattern of these hybrid molecules in animals needs to be explored. In the present study, the amoxicillin-sulbactam hybrid molecule (AS group) and a mixture of amoxicillin and sulbactam (mixture group) were administered to rats by gavage, and the levels of the major metabolites of amoxicillin, amoxicilloic acid, amoxicillin diketopiperazine, and sulbactam were determined by UPLC-MS/MS. The residue elimination patterns of the major metabolites in the liver, kidney, urine, and feces of rats in the AS group and the mixture group were compared. The results showed that the total amount of amoxicillin, amoxicilloic acid, amoxicillin diketopiperazine, and the highest concentration of sulbactam in the liver and kidney samples of the AS group and the mixture group appeared at 1 h after drug withdrawal. Between 1 h and 12 h post discontinuation, the total amount of amoxicillin, amoxicilloic acid, and amoxicillin diketopiperazine in the two tissues decreased rapidly, and the elimination half-life of the AS group was significantly higher than that in the mixture group (p < 0.05); the residual amount of sulbactam also decreased rapidly, and the elimination half-life was not significantly different (p > 0.05). In 72 h urine samples, the total excretion rates were 60.61 ± 2.13% and 62.62 ± 1.73% in the AS group and mixture group, respectively. The total excretion rates of fecal samples (at 72 h) for the AS group and mixture group were 9.54 ± 0.26% and 10.60 ± 0.24%, respectively. These results showed that the total quantity of amoxicillin, amoxicilloic acid, and amoxicillin diketopiperazine was eliminated more slowly in the liver and kidney of the AS group than those of the mixture group and that the excretion rate through urine and feces was essentially the same for both groups. The residual elimination pattern of the hybrid molecule in rats determined in this study provides a theoretical basis for the in-depth development and application of hybrid molecules, as well as guidelines for the development of similar drugs.

Involvement of autologous myeloid dendritic cells in the evaluation of immediate hypersensitivity reactions to betalactams.

BACKGROUND: Amoxicillin (AX) and clavulanic acid (CLV) are the betalactam antibiotics (BLs) most used to treat bacterial infections, although they can trigger immediate hypersensitivity reactions (IDHRs). The maturation analysis of monocyte-derived dendritic cells (moDCs) and their capacity to induce proliferative response of lymphocytes are useful to test the sensitisation to a drug, although without optimal sensitivity. Nevertheless, this can be improved using directly isolated DCs such as myeloid DCs (mDCs). METHODS: mDCs and moDCs were obtained from 28 allergic patients (AP), 14 to AX, 14 to CLV and from 10 healthy controls (HC). The expression of CCR7, CD40, CD80, CD83, and CD86 was analysed after stimulation with both BLs. We measured the capacity of these pre-primed DCs to induce drug-specific activation of different lymphocyte subpopulations, CD3+, CD4+, CD8+, CD4+Th1, and CD4+Th2, by flow cytometry. RESULTS: Higher expression of CCR7, CD40, CD80, CD83, and CD86 was observed on mDCs compared to moDCs from AP after stimulating with the culprit BL. Similarly, mDCs induced higher proliferative response, mainly of CD4+Th2 cells, compared to moDCs, reaching up to 67% of positive results with AX, whereas of only 25% with CLV. CONCLUSIONS: mDCs from selective AP efficiently recognise the culprit drug which trigger the IDHR. mDCs also trigger proliferation of lymphocytes, mainly those with a Th2 cytokine pattern, although these responses depend on the nature of the drug, mimicking the patient's reaction.

Embryonic Amoxicillin Exposure Has Limited Impact on Liver Development but Increases Susceptibility to NAFLD in Zebrafish Larvae.

Amoxicillin is commonly used in clinical settings to target bacterial infection and is frequently prescribed during pregnancy. Investigations into its developmental toxicity and effects on disease susceptibility are not comprehensive. Our present study examined the effects of embryonic amoxicillin exposure on liver development and function, especially the effects on susceptibility to non-alcoholic fatty liver disease (NAFLD) using zebrafish as an animal model. We discovered that embryonic amoxicillin exposure did not compromise liver development, nor did it induce liver toxicity. However, co-treatment of amoxicillin and clavulanic acid diminished BESP expression, caused bile stasis and induced liver toxicity. Embryonic amoxicillin exposure resulted in elevated expression of lipid synthesis genes and exacerbated hepatic steatosis in a fructose-induced NAFLD model, indicating embryonic amoxicillin exposure increased susceptibility to NAFLD in zebrafish larvae. In summary, this research broadens our understanding of the risks of amoxicillin usage during pregnancy and provides evidence for the impact of embryonic amoxicillin exposure on disease susceptibility in offspring.

Enlarging the substrate binding pocket of penicillin G acylase from Achromobacter sp. for highly efficient biosynthesis of ß-lactam antibiotics.

Penicillin G acylase (PGA) is a key biocatalyst for the enzymatic production of ß-lactam antibiotics, which can not only catalyze the synthesis of ß-lactam antibiotics but also catalyze the hydrolysis of the products to prepare semi-synthetic antibiotic intermediates. However, the high hydrolysis and low synthesis activities of natural PGAs severely hinder their industrial application. In this study, a combinatorial directed evolution strategy was employed to obtain new PGAs with outstanding performances. The best mutant ßF24G/ßW154G was obtained from the PGA of Achromobacter sp., which exhibited approximately a 129.62-fold and a 52.55-fold increase in specific activity and synthesis/hydrolysis ratio, respectively, compared to the wild-type AsPGA. Thereafter, this mutant was used to synthesize amoxicillin, cefadroxil, and ampicillin; all conversions > 99% were accomplished in 90-135 min with almost no secondary hydrolysis byproducts produced in the reaction. Molecular dynamics simulation and substrate pocket calculation revealed that substitution of the smallest glycine residue at ßF24 and ßW154 expanded the binding pocket, thereby facilitating the entry and release of substrates and products. Therefore, this novel mutant is a promising catalyst for the large-scale production of ß-lactam antibiotics.

Impact of prenatal amoxicillin exposure on hippocampal development deficiency.

BACKGROUND: Amoxicillin has been widely used to treat infectious diseases during pregnancy. Current studies suggest that amoxicillin exposure during pregnancy could lead to developmental disorders in the offspring and increase the incidence of long-term complications such as asthma and kidney damage in adulthood. However, the adverse effects of prenatal amoxicillin exposure (PAmE) including administration stage, doses and courses on fetal hippocampal neurodevelopment and its function in the offspring have not been elucidated. In this study, we intend to investigate the effects of PAmE on fetal hippocampal development and its possible mechanisms. METHOD: Pregnant Kunming mice were given intragastric administration with amoxicillin at different administration stage, doses and courses, and GD (gestational day) 18 offspring hippocampus was collected for morphological and development-related functional assays, and the molecular mechanisms were explored. RESULTS: PAmE induced hippocampal hypoplasia in the offspring with suppressed hippocampal neuronal cell proliferation and impaired neuronal synaptic plasticity comparatively; hippocampal astrocyte and microglia were damaged to varying degrees. The developmental toxicity of PAmE in fetal mices varies by time, dose, and course of treatment. The most severe damage was observed in the late gestation, high dose, and multi-course dosing groups. The significant reduction either in SOX2, an essential gene in regulating neural progenitor cell proliferation, and reduction of genes related to the Wnt/ß-catenin pathway may suggest that the key role of SOX2/Wnt/ß-catenin pathway in impaired hippocampal development in the offspring due to PAmE. CONCLUSION: In this study, PAmE was found to be developmentally toxic to the hippocampus thus to induce developmental damage to various hippocampal cells; Even with current clinically safe doses, potential hippocampal damage to offspring may still present; This study provides a theoretical and experimental basis for guiding the rational usage of drugs during pregnancy and giving effectively assessment of the risk on fetal hippocampal developmental toxicity.

Protective effects of amoxicillin and probiotics on colon disorders in an experimental model of acute diverticulitis disease.

Acute diverticulitis disease is associated with inflammation and infection in the colon diverticula and may lead to severe morbidity. This study aimed to evaluate and compare the protective effects of amoxicillin antibiotic, either alone or in combination with probiotics (Lactobacillus acidophilus and Bifidobacterium lactis), in a rat model of acute diverticulitis disease. Acute diverticulitis was induced, in albino rats, by adding 3% weight/volume of dextran sulfate sodium (DSS) to the rats' drinking water; daily for 7 days, in addition to injecting lipopolysaccharide (LPS) enema (4 mg/kg). The impact of treatments was assessed by measuring the physiological and immunological parameters and evaluating colon macroscopic and microscopic lesions. The results showed that both treatments (especially probiotics with amoxicillin) alleviated the adverse effects of DSS and LPS. This was obvious through the modulation of the rats' body weight and the colon weight-to-length ratio. Also, there was a significant (p < 0.001) decrease in the colon macroscopic lesion score. The pro-inflammatory cytokines [(TNF)-α, (IL)-1ß, (IFN)-γ, and (IL)-18]; in the colon tissue; were significantly (p < 0.001) decreased. Also, both treatments significantly ameliorated the elevation of myeloperoxidase activity and C-reactive protein levels, in addition to improving the histopathological alterations in the colon tissue. In conclusion, amoxicillin and probiotics-amoxicillin were effective in preventing the development of experimentally induced acute diverticulitis, through their anti-inflammatory and immunomodulatory effects. Furthermore, this study has explored the role of probiotics in preventing DSS/LPS-induced acute diverticulitis, so it can be applied as a promising treatment option for acute diverticulitis disease.

Effects of ciprofloxacin, trimethoprim, and amoxicillin on microbial structure and growth as emerging pollutants reaching crop soils.

The presence of emerging pollutants, and specifically antibiotics, in agricultural soils has increased notably in recent decades, causing growing concern as regards potential environmental and health issues. With this in mind, the current study focuses on evaluating the toxicity exerted by three antibiotics (amoxicillin, trimethoprim, and ciprofloxacin) on the growth of soil bacterial communities, when these pollutants are present at different doses, and considered in the short, medium, and long terms (1, 8 and 42 days of incubation). Specifically, the research was carried out in 12 agricultural soils having different physicochemical characteristics and was performed by means of the leucine (3H) incorporation method. In addition, changes in the structure of soil microbial communities at 8 and 42 days were studied in four of these soils, using the phospholipids of fatty acids method for this. The main results indicate that the most toxic antibiotic was amoxicillin, followed by trimethoprim and ciprofloxacin. The results also show that the toxicity of amoxicillin decreases with time, with values of Log IC50 ranging from 0.07 ± 0.05 to 3.43 ± 0.08 for day 1, from 0.95 ± 0.07 to 3.97 ± 0.15 for day 8, and from 2.05 ± 0.03 to 3.18 ± 0.04 for day 42, during the incubation period. Regarding trimethoprim, 3 different behaviors were observed: for some soils the growth of soil bacterial communities was not affected, for a second group of soils trimethoprim toxicity showed dose-response effects that remained persistent over time, and, finally, for a third group of soils the toxicity of trimethoprim increased over time, being greater for longer incubation times (42 days). As regards ciprofloxacin, this antibiotic did not show a toxicity effect on the growth of soil bacterial communities for any of the soils or incubation times studied. Furthermore, the principal component analysis performed with the phospholipids of fatty acids results demonstrated that the microbial community structure of these agricultural soils, which persisted after 42 days of incubation, depended mainly on soil characteristics and, to a lesser extent, on the dose and type of antibiotic (amoxicillin, trimethoprim or ciprofloxacin). In addition, it was found that, in this research, the application of the three antibiotics to soils usually favored the presence of fungi and Gram-positive bacteria.

The acquisition of clinically relevant amoxicillin resistance in Streptococcus pneumoniae requires ordered horizontal gene transfer of four loci.

Understanding how antimicrobial resistance spreads is critical for optimal application of new treatments. In the naturally competent human pathogen Streptococcus pneumoniae, resistance to ß-lactam antibiotics is mediated by recombination events in genes encoding the target proteins, resulting in reduced drug binding affinity. However, for the front-line antibiotic amoxicillin, the exact mechanism of resistance still needs to be elucidated. Through successive rounds of transformation with genomic DNA from a clinically resistant isolate, we followed amoxicillin resistance development. Using whole genome sequencing, we showed that multiple recombination events occurred at different loci during one round of transformation. We found examples of non-contiguous recombination, and demonstrated that this could occur either through multiple D-loop formation from one donor DNA molecule, or by the integration of multiple DNA fragments. We also show that the final minimum inhibitory concentration (MIC) differs depending on recipient genome, explained by differences in the extent of recombination at key loci. Finally, through back transformations of mutant alleles and fluorescently labelled penicillin (bocillin-FL) binding assays, we confirm that pbp1a, pbp2b, pbp2x, and murM are the main resistance determinants for amoxicillin resistance, and that the order of allele uptake is important for successful resistance evolution. We conclude that recombination events are complex, and that this complexity contributes to the highly diverse genotypes of amoxicillin-resistant pneumococcal isolates.

Selective Inhibition of Helicobacter pylori Carbonic Anhydrases by Carvacrol and Thymol Could Impair Biofilm Production and the Release of Outer Membrane Vesicles.

Helicobacter pylori, a Gram-negative neutrophilic pathogen, is the cause of chronic gastritis, peptic ulcers, and gastric cancer in humans. Current therapeutic regimens suffer from an emerging bacterial resistance rate and poor patience compliance. To improve the discovery of compounds targeting bacterial alternative enzymes or essential pathways such as carbonic anhydrases (CAs), we assessed the anti-H. pylori activity of thymol and carvacrol in terms of CA inhibition, isoform selectivity, growth impairment, biofilm production, and release of associated outer membrane vesicles-eDNA. The microbiological results were correlated by the evaluation in vitro of H. pylori CA inhibition, in silico analysis of the structural requirements to display such isoform selectivity, and the assessment of their limited toxicity against three probiotic species with respect to amoxicillin. Carvacrol and thymol could thus be considered as new lead compounds as alternative H. pylori CA inhibitors or to be used in association with current drugs for the management of H. pylori infection and limiting the spread of antibiotic resistance.

Rectal bioavailability of amoxicillin sodium in rabbits: Effects of suppository base and drug dose.

In this paper, rectal absorption and tissue tolerance of amoxicillin sodium (AS) suppositories prepared in a hydrophilic base, polyethylene glycol (PEG) or lipophilic base, Suppocire® NA 15 (SNA 15), were investigated. Following in vitro characterization, including drug distribution in the suppository bases, drug-base interactions and drug release, pharmacokinetics were investigated in rabbits to determine absolute bioavailability (F) at two dose levels (100 mg and 200 mg). Both types of suppositories were found uniform in weight and content. Powder X-ray diffraction (XRD) and differential scanning calorimetry indicated that AS existed as solid dispersion or anhydrous crystalline dispersion in both suppositories at different ratios without changing melting points of the bases. This was supported by Fourier transform infrared (FTIR) spectroscopy and scanning electron microscopy conjugated with energy dispersive X-ray (SEM/EDX). In dissolution medium, melting and spreading of SNA 15 and dissolution of PEG suppositories accounted for their different drug release kinetics and mean dissolution time (MDT). A rapid and complete amoxicillin absorption (F close to 100%) with a double peak pharmacokinetic profile was observed alongside minimal signs of tissue irritation in rabbits treated with SNA 15 suppositories at both dose levels. In contrast, the F of amoxicillin from PEG suppositories was 59%, increasing to 77.3% as AS dose doubled from 100 mg to 200 mg, reflected in the slower release predominately controlled by erosion of the base. An in vitro - in vivo correlation was observed (MDT vs F; p < 0.01). AS was stable in SNA 15 suppositories at least for three months at 20 ± 0.2 °C. This research highlighted the advantages of SNA 15 suppositories over the PEG suppositories in providing rapid and complete rectal absorption of AS and tissue compatibility.

Complexation of amoxicillin by transition metals: Physico-chemical and antibacterial activity evaluation.

Some bacteria have developed resistance to antibiotics that were once commonly used to treat them. Moreover, this resistance has become more and more massive and worrying. During this work, we succeeded in synthesizing "metal-antibiotic" complexes, combining as a ligand for the metals of Cu (II), Zn (II) and Fe (III). These complexes AMX - M (M = Cu, Fe and Zn) were characterized by UV-Vis spectrophotometry, IR spectroscopy, and electrochemical methods. Job's method of continuous variation suggested 1:1 metals to ligand stoichiometry for all amoxicillin complexes. The binding constant/association constant (K) of the AMX with Zn(II), Cu(II), and Fe(III) were found to be 4.46 × 104, 7.17 × 102 and 7.65 × 102 L mol-1, respectively. The IR spectra shows that the ligands coordinated to the metal ions through amino, imino, carboxylate, ß-lactamic and carbonyl groups. The electrochemical results proved that amoxicillin oxidation process can be delayed by transition metal complexation. After, the complex synthesis, the antibacterial activity of ligand and its metal complexes were evaluated against Escherichia. coli bacteria by antibiogram method. The results show that the metal-amoxicillin complexes have better antibacterial activity against Escherichia coli (E. coli) than the free ligand (amoxicillin) due to the AMX protection against oxidation after complexation.

Electrochemical oxidation of amoxicillin, ciprofloxacin and erythromycin in water: Effect of experimental factors on COD removal.

An alternative method of electrochemical oxidation was employed to degrade persistent compounds in the form of antibiotics using strong oxidizing agents such as hydroxyl ions. A 24 factorial design was employed to check the effect of four factors namely pH, current density, electrolysis time and electrolyte concentration set at their high (+) and low (-) levels on the antibiotics (amoxicillin, ciprofloxacin and erythromycin) degradation in water. The response was obtained in the form of COD (chemical oxygen demand) removal. A prediction model was developed to predict the values of COD removal. Later the main effect, contribution and interactions were studied with Design Expert Software 7.0. About 89.5% COD removal was obtained when pH and time were set at their high level and the other two factors at their low level. It was determined that the pH when set at high level (pH 9) had the most effect (24.68) and contribution (43.6) in the degradation process and hence the removal of COD. This technology of electrochemical oxidation can be employed in industries to efficiently remove pharmaceuticals, paints, dyes and other organic compounds.

Mini-Tablets versus Nanoparticles for Controlling the Release of Amoxicillin: In vitro/In vivo Study.

INTRODUCTION: Controlling the drug release from the dosage form at a definite rate is the main challenge for a successful oral controlled-release drug delivery system. In this study, mini-tablets (MTs) and lipid/polymer nanoparticles (LPNs) of lipid polymer and chitosan in different ratios were designed to encapsulate and control the release time of Amoxicillin (AMX). METHODS: Physical characteristics and in vitro release profiles of both MT and LPN formulations were studied. Antimicrobial activity and oral pharmacokinetics of the optimum MT and LPN formulations in comparison to market tablet were studied in rats. RESULTS: All designed formulations of AMX as MTs and LPNs showed accepted characteristics. MT-6 (Compritol/Chitosan 1:1) showed the greatest retardation among all prepared minitablet preparations, releasing about 79.5% of AMX over 8 h. In contrast, LPN-11 (AMX: Cr 1:3/Chitosan 1 mg/mL) had the slowest drug release, revealing the sustained release of 80.9% within 8 h. The MIC of both optimized tablet formula (MT-6) and LPNs formula (LPN-11) was around two-fold lower than the control against H. pylori. The Cmax of MT-6 and LPN11 were non significantly different compared with the marketed AMX product. While the bioavailability experiment proved that the relative bioavailability of the AMX was 1.85 and 1.8 after the oral use of LPN11 and MT-6, respectively, compared to the market tablet. CONCLUSION: The results verified that both controlled-release mini-tablets and lipid/polymer nanoparticles can be used for sustaining the release and hence improve the bioavailability of amoxicillin.

Individual and combined effects of amoxicillin, enrofloxacin, and oxytetracycline on Lemna minor physiology.

We investigated individual and combined effects of environmentally representative concentrations of amoxicillin (AMX; 2 µg l-1), enrofloxacin (ENR; 2 µg l-1), and oxytetracycline (OXY; 1 µg l-1) on the aquatic macrophyte Lemna minor. While the concentrations of AMX and ENR tested were not toxic, OXY decreased plant growth and cell division. OXY induced hydrogen peroxide (H2O2) accumulation and related oxidative stress through its interference with the activities of mitochondria electron transport chain enzymes, although those deleterious effects could be ameliorated by the presence of AMX and/or ENR, which prevented the overaccumulation of ROS by increasing catalase enzyme activity. L. minor plants accumulated significant quantities of AMX, ENR and OXY from the media, although competitive uptakes were observed when plants were submitted to binary or tertiary mixtures of those antibiotics. Our results therefore indicate L. minor as a candidate for phytoremediation of service waters contaminated by AMX, ENR, and/or OXY.

Co-substrate addition accelerated amoxicillin degradation and detoxification by up-regulating degradation related enzymes and promoting cell resistance.

ß-Lactam antibiotics are the most commonly used antibiotics, and are difficult to remove by conventional biological treatments because of their persistent and toxic nature. The addition of co-substrates has been successfully employed to improve the removal of refractory pollutants. So, we hypothesized that the co-substrate strategy would increase antibiotic degradation and benefit microbial survival. In this work, we reported that co-substrate (acetate) addition up-regulated key degrading enzymes and resistance related genes in a model bacteria strain (L. aquatilis) when being treated with 0.055 mM amoxicillin (AMO). ß-Lactamase, amidases, transaminase, and amide C-N hydrolase showed increased activation. As a result, AMO removal reached ∼95 %, a ∼60 % increase over the control. Furthermore, the addition of acetate drove the down-stream TCA cycle, which accelerated the detoxification of the intermediates and reduced the microbial inhibition by the antibiotic products to as low as ∼15 %. Besides, the expression levels of genes encoding the efflux pump, penicillin binding proteins, and ß-Lactamase were up-regulated, and the inhibition of peptidoglycan biosynthesis was down-regulated. The cell density was enhanced by ∼170 % and showed improved DNA replication. In conclusion, the addition of the co-substrate accelerated AMO degradation and detoxification by up-regulating degrading enzymes and promoting cell resistance.

Potential application in amoxicillin removal of Alcaligenes sp. MMA and enzymatic studies through molecular docking.

Antibiotic contamination in environmental matrices is a serious global problem which leads to an increase in the proliferation of antibiotic resistance genes. Amoxicillin is ubiquitous in the environment, but there is hardly any information on the dissipation of amoxicillin by the microbial community. In view of this, the present study focusses on the removal of amoxicillin using amoxicillin-resistant bacteria, Alcaligenes sp. MMA. Bacteria were characterized using antibiotic tests, biochemical and molecular analysis. Alcaligenes sp. MMA was able to remove up to 84% of amoxicillin in 14 days in M9 minimal media, and the degradation products were confirmed using LC-MS/MS, including the benzothiazole, 2-Amino-3-methoxyl benzoic acid, 4-Hydroxy-2-methyl benzoic acid, 5-Amino-2-methylphenol and 3,5-Bis(tert-butyl)-2-hydroxybenzaldehyde, at the end of 14th day which further shows the removal of amoxicillin by the bacterial strain. Differential expression of porins was found in the presence of amoxicillin as a sole source of carbon and energy for the bacterial strain. Molecular interaction using in silico studies were performed which showed the formation of a hydrogen bond between amoxicillin and porins.

Optimization of Culture Conditions for Amoxicillin Degrading Bacteria Screened from Pig Manure.

(1) Objective: The objective of this study was to screen amoxicillin (AMX)-degrading bacterial strains in pig manure and optimize the fermentation conditions for these strains to achieve high fermentation rate, which can provide an effective way for the practical application of bacterial strains as antibiotic-degrading bacterial in treating livestock waste for antibiotic residues. (2) Methods: Antibiotic susceptibility tests and high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) were employed to screen AMX-degrading bacterial strains in pig manure. The culture conditions were optimized for AMX-degrading bacterial strains using Plackeet-Burman design (PBD), the steepest ascent design, and the response surface methods, coupled with the Box-Behnken design (BBD). The effects of culture time, temperature, rotator (mixing) speed, inoculum level, and initial pH value on the growth of AMX-degrading strains were investigated. Experimental data obtained from BBD were utilized to generate a second-order polynomial regression model for evaluating the effects of the tested variables on the optical density at 600 nm (OD600) of culture solutions as the growth indicator for the screened AMX-degrading strains. (3) Results: The initial pH, culture time, and the inoculum level had significant effects on the OD600 value (growth) of the screened AMX-degrading strains. The initial pH value was found to be the most critical factor influencing the growth of bacteria. The optimized culture condition for the bacterial growth determined by the response surface methodology was: the initial pH of 6.9, culture time of 52 h, and inoculum level of 2%. The average OD value of 12 different fermentation conditions in the initial fermentation tests in this study was 1.72 and the optimization resulted in an OD value of 3.00. The verification experiment resulted in an OD value of 2.94, which confirmed the adequacy of the optimization model for the determining the optimal culture condition. (4) Conclusions: The growth of the screened strain of AMX-degrading bacteria could be optimized by changing the fermentation conditions. The optimization could be achieved by using the Box-Behnken response surface method and Plackett-Burman experimental design.

Amoxicillin removal by pre-denitrification membrane bioreactor (A/O-MBR): Performance evaluation, degradation by-products, and antibiotic resistant bacteria.

Membrane bioreactors (MBRs) are one of the treatment technologies with the potential to remove emerging compounds from wastewater. The present work evaluated the efficiency of an MBR pilot system in removing amoxicillin from synthetic wastewater using a continuous flow pre-denitrification MBR (A/O-MBR) pilot unit. The system operated in three phases: (1) synthetic wastewater and hydraulic retention time (HRT) of 40 h; (2) adding amoxicillin 100 µg L-1 to the influent, and (3) varying flowrate to HRT of 20 h. Liquid chromatography coupled to high resolution mass spectrometry analysis confirmed the presence of five amoxicillin degradation by-products in the effluent. The addition of amoxicillin did not affect chemical oxygen demand (COD) or dissolved organic carbon (DOC) removal efficiencies. Respirometry showed that amoxicillin level did not inhibit heterotrophic bacteria metabolism. The change in HRT reduced the DOC removal (from 84% to 66%) but did not influence COD (>94%) or total nitrogen (>72%). The amoxicillin and by-products removal decreased from 80% to 54% with HRT change. Adsorption and biodegradation represented the largest removed fraction of the antibiotic in the A/O-MBR system (68%). Ecotoxicity assays showed P. fluorescens was more resistant and E. coli less resistant to amoxicillin residues at effluent sample matrix.

Mechanistic and ecotoxicological studies of amoxicillin removal through anaerobic degradation systems.

Many studies have been conducted on the evaluation and monitoring of micropollutants and by-products in wastewater treatment plants. Considering the increase in the production and consumption of emerging contaminants, such as drugs, personal care products, and plasticisers, it is necessary to conduct studies that support the elaboration of laws and regulations that promote the environmentally sustainable use of sludge and effluents. In this work, the biological degradation of amoxicillin was studied under two anaerobic conditions: i) using a 6 L reactor operated under semi-continuous flow; and ii) a batch system with 100 mL sealed glass syringes. According to the statistical analysis, amoxicillin was completely removed from the systems, but biogas production inhibition was observed (p < 0.05). Liquid chromatography-high-resolution mass spectrometry analysis identified amoxicillin penicilloic acid, amoxilloic acid, amoxicillin diketopiperazine and phenol hydroxypyrazine as by-products under anaerobic conditions. Ecotoxicity tests on effluent treated under the batch conditions showed that the addition of higher amounts of amoxicillin inhibited the target species Aliivibrio fischeri and Raphidocelis subcaptata, causing functional decreases of 28.5% and 22.2% when the antibiotic concentration was 2500 µg L-1. A. fischeri was the most sensitive organism to effluent treated under semi-continuous flow conditions; a continuous reduction in bioluminescence of up to 88.8% was observed after 39 days of feeding, which was associated with by-products accumulation due to unbalanced conditions during anaerobic digestion. Changes in the physico-chemical characteristics of the effluent caused the accumulation and removal of AMX-DKP IV and modified the toxicity to Lactuca sativa and R. subcapitata.

In vitro equivalence of generic and branded amoxicillin tablet by microbiological assay method.

Background Indonesian Ministry of Health advocate doctors, especially in government-owned healthcare facility, to prescribe generic drugs including amoxicillin. Although BPOM (the National Agency of Drug and Food Control) already guarantees that the generic amoxicillin and the branded one were interchangeable, lack of confidence in generic drugs still remains among patients, pharmacists, and doctors. This issue supported by lack of publication confirmed the therapeutic equivalence of branded and generic drugs. This study aims to evaluate and compare the in vitro microbiological assay of different generic and branded amoxicillin that are available in Indonesian market, especially those used in government-owned healthcare facilities. Methods Microbiological assays for five samples of amoxicillin tablet containing 500 mg amoxicillin available in Indonesia were determined using a method from Indonesia Pharmacopeia. Samples were coded as Products A to E. The assay was carried out by measuring the diameter of the inhibition zones in the plate agar incubated with Escherichia coli and Staphylococcus aureus. The obtained data were evaluated to determine the sample potency and compared with the amoxicillin reference standard. Results Minor and insignificant differences (p > 0.05) were found in the diameters of the inhibition zones. Potency ratio measured both in E. coli and S. aureus were all between 95% and 105%. The lowest of the tested samples were from Product C, which resulted to ratio potencies of 96.3% and 95.5% in E. coli and S. aureus, respectively. Conclusions All five samples were in the range of the acceptance criteria. Therefore, from the view of the microbiological assay, these products are in equivalence in quality and are interchangeable.