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Background: Proton pump inhibitors (PPIs) are widely used for the treatment of gastrointestinal disorders such as peptic ulcer disease and dyspepsia. However, several studies have suggested that PPI use increases the risk of acute kidney injury (AKI). PPIs are often concomitantly used with antibiotics, such as macrolides and penicillins for Helicobacter pylori eradication. Although macrolide antibiotics are considered to have relatively low nephrotoxicity, they are well known to increase the risk of AKI due to drug-drug interactions. In this study, we aimed to investigate the association between PPI use and the development of AKI. We also evaluated the effect of concomitant use of PPIs and macrolide antibiotics on the risk of AKI. Methods: This self-controlled case series study was conducted using electronic medical records at Kyoto University Hospital. We identified patients who were prescribed at least one PPI and macrolide antibiotic between January 2014 and December 2019 and underwent blood examinations at least once a year. An adjusted incident rate ratio (aIRR) of AKI with PPI use or concomitant use macrolide antibiotics with PPIs was estimated using a conditional Poisson regression model controlled for the estimated glomerular filtration rate at the beginning of observation and use of potentially nephrotoxic antibiotics. Results: Of the 3, 685 individuals who received PPIs and macrolide antibiotics, 766 patients with episodes of stage 1 or higher AKI were identified. Any stage of AKI was associated with PPI use (aIRR, 1.80 (95% confidence interval (CI) 1.60 to 2.04)). Stage 2 or higher AKI was observed in 279 cases, with an estimated aIRR of 2.01 (95% CI 1.57 to 2.58, for PPI use). For the period of concomitant use of macrolide antibiotics with PPIs compared with the period of PPIs alone, an aIRR of stage 1 or higher AKI was estimated as 0.82 (95% CI 0.60 to 1.13). Conclusions: Our findings added epidemiological information for the association between PPI use

Objectives: Pseudomonas aeruginosa colonizes the cystic fibrosis (CF) airways causing chronic bacterial lung infections. CF patients are routinely treated with macrolides, however, P. aeruginosa is considered insusceptible as consequence of inadequate susceptibility testing leaving resistance mechanism completely overlooked. Here, we investigated a new mechanism of macrolide resistance caused by ribosomal protein mutations. Methods: Investigating a longitudinal collection of 529 isolates from CF patients and analysing 5758 protein sequences from different sources, mutations in P. aeruginosa's ribosomal proteins connected to macrolide resistance were identified. Using a modified susceptibility testing protocol, isolates harbouring a mutated uL4 ribosomal protein were tested for resistance against macrolide antibiotics and macrolide-induced quorum sensing modulation. Proteome and ribosome profiling were applied to assess the impact of the mutations on the bacterial physiology. Results: Five uL4 mutations were identified in isolates from different CF patients. Most mapped to the conserved loop region of uL4 and resulted in increased macrolide tolerance (>10-fold relative to wt strains). Greater concentrations (>10-fold) of macrolide antibiotic were needed to inhibit the growth, reduce swimming motility, and induce redox sensitivity of the uL4 mutants. 16 proteins involved in ribosome adaptation displayed altered expression possibly to compensate for the uL4 mutations, which changed the ribosome stoichiometry without negatively affecting bacterial physiology. Conclusions: Macrolide antibiotics should, therefore, be considered as active antimicrobial agents against P. aeruginosa and resistance development should be contemplated when patients are treated with prolonged courses of macrolides. Importantly, improved macrolide susceptibility testing is necessary for the detection of resistant bacteria.

Objectives Pseudomonas aeruginosa colonizes the cystic fibrosis (CF) airways causing chronic bacterial lung infections. CF patients are routinely treated with macrolides, however, P. aeruginosa is considered insusceptible as consequence of inadequate susceptibility testing leaving resistance mechanism completely overlooked. Here, we investigated a new mechanism of macrolide resistance caused by ribosomal protein mutations. Methods Investigating a longitudinal collection of 529 isolates from CF patients and analysing 5758 protein sequences from different sources, mutations in P. aeruginosa's ribosomal proteins connected to macrolide resistance were identified. Using a modified susceptibility testing protocol, isolates harbouring a mutated uL4 ribosomal protein were tested for resistance against macrolide antibiotics and macrolide-induced quorum sensing modulation. Proteome and ribosome profiling were applied to assess the impact of the mutations on the bacterial physiology. Results Five uL4 mutations were identified in isolates from different CF patients. Most mapped to the conserved loop region of uL4 and resulted in increased macrolide tolerance (>10-fold relative to wt strains). Greater concentrations (>10-fold) of macrolide antibiotic were needed to inhibit the growth, reduce swimming motility, and induce redox sensitivity of the uL4 mutants. 16 proteins involved in ribosome adaptation displayed altered expression possibly to compensate for the uL4 mutations, which changed the ribosome stoichiometry without negatively affecting bacterial physiology. Conclusions Macrolide antibiotics should, therefore, be considered as active antimicrobial agents against P. aeruginosa and resistance development should be contemplated when patients are treated with prolonged courses of macrolides. Importantly, improved macrolide susceptibility testing is necessary for the detection of resistant bacteria.

The study of the presence of antibiotics in the aquatic environment is a preliminary step to analyse their possible harmful effects on aquatic ecosystems. In order to monitor their occurrence in the aquatic environment, the European Commission established in 2015, 2018, and 2020 three Watch Lists of substances for Union-wide monitoring (Decisions (EU) 2015/495, 2018/840, and 2020/1161), where some antibiotics within the classes of macrolides, fluoroquinolones and penicillins were included. In the Basque coast, northern Spain, three macrolide antibiotics (erythromycin, clarithromycin, azithromycin) and ciprofloxacin were monitored quarterly from 2017 to 2020 (covering a period before and after the COVID19 outbreak), in water samples collected from two Waste Water Treatment Plants (WWTPs), and three control points associated with receiving waters (transitional and coastal water bodies). This work was undertaken for the Basque Water Agency (URA). The three macrolide antibiotics in water showed a frequency of quantification >65 % in the Basque coast, with higher concentrations in the WWTP emission stations than in receiving waters. Their frequency of quantification decreased from 2017 to 2020, as did the consumption of antibiotics in Spanish primary care since 2015. Ciprofloxacin showed higher frequencies of quantification in receiving waters than in wastewaters, but the highest concentrations were observed in the WWTP emission stations. Although consumption of fluoroquinolones (among which is ciprofloxacin) in primary care in the Basque Country has decreased in recent years, this trend was not observed in the waters sampled in the present study. On the other hand, concentrations of clarithromycin, azithromycin, and ciprofloxacin in receiving waters exceeded their respective Predicted No-Effect Concentrations, so they could pose an environmental risk. These substances are widely used in human and animal medicine, so, although only ciprofloxacin is included in the third

In this work, carbon paste electrode (CPE) and carbon paste electrode modified with gold nanoparticles (AuNPs/CPE) were employed with rapid direct anodic square wave voltammetry (SWV) for macrolide antibiotics erythromycin ethylsuccinate (EES), azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) determination. The surfaceof working electrodes was morphologically characterized by scanning electron microscopic (SEM) measurements whereby the presence of randomly distributed AuNPs of 10 nmdiameter size on CPE surface was confirmed by energy dispersive spectrometer (EDS). SWV determination of four macrolide antibiotics using CPE was performed in aqueous Britton-Robinson buffer solutions (pH 2.0 to 11.98) showing that oxidation signals strongly depend upon pH (at pH ≥ 6) and exhibited the most favorable characteristics at pH 8.0 in the case of EES and pH 11.98 in the case of AZI, CLA and ROX. Under optimized conditions, the SWV method using CPE enables determination of all investigated macrolide antibiotics in low μg mL−1 concentration ranges with relative standard deviations (RSDs) lower than 6%and achieved detection limits (LODs) as 0.18, 0.045, 1.43 and 0.30 μgmL−1 for EES, AZI, CLA and ROX, respectively. In the case of AZI and ROX, it was demonstrated that the use of AuNPs/CPE as working electrode could additionally improve the results obtained with the SWV method concerning exhibited sensitivity, reproducibility and linear concentrationrange, due to the electrocatalytic properties of synthesized AuNPs. The optimized experimental parameters with the use of SWV method and CPE or AuNPs/CPE were successfully applied for determination of ROX and AZI in their pharmaceutical preparations Runac® and Hemomycin®, respectively. The reliability of the elaborated procedures and thus the accuracy of the obtained results were validated by comparing them with those obtained by means of HPLC-DAD measurements.

Total synthesis of natural products is an important part of organic chemistry and has an impact on life sciences. Utilization of natural products in combating against human diseases has a long history and total synthesis has become an essential approach to get access to limited supply of rare natural products and its derivatives nowadays. Besides, total synthesis enables exploration and discovery of many unprecedented and novel reactivities and efficient synthetic strategies. This thesis research deals with total synthesis studies on two different types of natural products. The first target molecule is the proposed structure of iriomoteolide-2a, whose C6–C18 fragment has been synthesized in this thesis work. The second type of target molecule is 4β-acetoxyprobotryane-9β,15α-diol and the congeners, which contain a strained trans-[3.3.0]octane core. A novel methodology for the construction of the strained core was explored and total synthesis 4β-acetoxyprobotryane-9β,15α-diol has been accomplished. Iriomoteolide-2a was the first 23-membered macrolide with significant bioactivity and structural novelty. But its bioactivity was limited as increasing dosage did not give a better activity, suggesting the demand for structural modifications. In Chapter 1, a brief background of iriomoteolides family of marine macrolides, including isolation, bioactivity, and reported synthetic studies, is provided. Chapter 2 presents the results of the synthesis of the C6–C18 bis-tetrahydrofuran (bis-THF) fragment of the proposed structure of iriomoteoldie-2a via stepwise double SN2 cyclization reaction. The C9–C12 THF ring was first constructed through an AD–SN2 cascade sequence while the C13–C16 THF ring was then installed via an intramolecular SN2 reaction of a chiral propargylic mesylate. Asymmetric transfer hydrogenation of propargylic ketones was demonstrated as a powerful synthetic tool to secure the stereogenic centers of the C16-propargylic carbon a

Natural products are rich resources for drug discovery and development. However, the complex molecular structures of natural products always complicate the total synthesis, structure–activity relationship investigations and structural optimizations. Therefore, simplifying complex structures without decreasing the desired biological activity is an effective strategy for improving synthetic accessibility and accelerating the drug development process. Amphidinolide C congeners are the cytotoxic macrolides isolated from the genus Amphidinium. Except for amphidinolide C4, they possess the same functionalized 25-membered macrolactone core and differ only on the side chain appended at C28. The 25-membered macrolactone core is decorated with some unusual structural components, such as an s-cis-1,3-diene subunit, an exo-methlyene moiety, and two trans-substituted tetrahydrofuran units. These natural compounds exhibit significant cytotoxic activities and they have attracted much interest from the synthetic community due to their remarkable bioactivities, low natural accessibility, and synthetic challenges for more than 20 years since their isolations. Total synthesis of amphidinolide C1 and F has been achieved by the groups of Carter, Fürstner and Figadère. However, up to now, the action mode of the amphidinolide C congeners is still unknown. Based on the limited information on the bioactivity of amphidinolide C congeners, we envisioned synthesis of the structurally simplified amphidinolide C congeners by depleting four stereogenic centers at C4, C12, C13, and C16. Three fragments were prepared and assembled via the Suzuki-Miyaura cross-coupling reaction under the Johnson conditions at C9-C10, esterification of C24-OH, and ring-closing metathesis at C16-C17. Then, the side chain was assembled onto the macrolactone core via Suzuki–Miyaura cross-coupling reaction of the C-26 alkenyl iodide by using our Aphos-Y as the ligand. In Chapter 1, a brief introduction of the structural

Bacterial resistance has become a major health threat that needs urgent action to maintain the advances of modern medicine. There is an urgent need to discover new bioactive natural compounds such as decalin-forming macrolides which have antibacterial potency and unique mode of action. Genome mining has become an important method to identify novel candidates of natural products through an efficient computational frameworks which utilize algorithms such as antiSMASH, BiG-SCPAE And CORASON. Through analysing ~15% of all available bacterial genomes, we have identified several potential new decalin-forming macrolide biosynthetic gene clusters. The most promising candidate is a BGC within the genome of Streptomyces tsukubaensis which contains all PKS genes of nargenicin and streptoseomycin and maintains the overall modular structure.

Natural products are rich resources for drug discovery and development. However, the complex molecular structures of natural products always complicate the total synthesis, structure–activity relationship investigations and structural optimizations. Therefore, simplifying complex structures without decreasing the desired biological activity is an effective strategy for improving synthetic accessibility and accelerating the drug development process. Amphidinolide C congeners are the cytotoxic macrolides isolated from the genus Amphidinium. Except for amphidinolide C4, they possess the same functionalized 25-membered macrolactone core and differ only on the side chain appended at C28. The 25-membered macrolactone core is decorated with some unusual structural components, such as an s-cis-1,3-diene subunit, an exo-methlyene moiety, and two trans-substituted tetrahydrofuran units. These natural compounds exhibit significant cytotoxic activities and they have attracted much interest from the synthetic community due to their remarkable bioactivities, low natural accessibility, and synthetic challenges for more than 20 years since their isolations. Total synthesis of amphidinolide C1 and F has been achieved by the groups of Carter, Fürstner and Figadère. However, up to now, the action mode of the amphidinolide C congeners is still unknown. Based on the limited information on the bioactivity of amphidinolide C congeners, we envisioned synthesis of the structurally simplified amphidinolide C congeners by depleting four stereogenic centers at C4, C12, C13, and C16. Three fragments were prepared and assembled via the Suzuki-Miyaura cross-coupling reaction under the Johnson conditions at C9-C10, esterification of C24-OH, and ring-closing metathesis at C16-C17. Then, the side chain was assembled onto the macrolactone core via Suzuki–Miyaura cross-coupling reaction of the C-26 alkenyl iodide by using our Aphos-Y as the ligand. In Chapter 1, a brief introduction of the structural

In this work, carbon paste electrode (CPE) and carbon paste electrode modified with gold nanoparticics (AuNPs/CPE) were employed with rapid direct anodic square wave voltammetry (SWV) for macrolide antibiotics erythromycin cthylsuccinate (EIS), azithromycin (A7.1), clarithromycin (OA) and roxithromycin (ROX) determination. The surface of working electrodes was morphologically characterized by scanning electron microscopic (SEM) measurements whereby the presence of randomly distributed AuNPs of 10 nm diameter size on CPE surface was confirmed by energy dispersive spectrometer (EDS). SWV determination of four macrolide antibiotics using CPE was performed in aqueous Britton-Robinson buffer solutions (pH 2.0 to 11.98) showing that oxidation signals strongly depend upon pH (at pH gt = 6) and exhibited the most favorable characteristics at pH 8.0 in the case of EES and pH 11.98 in the case of AZI, CIA and ROX. Under optimized conditions, the SWV method using CPE enables determination of all investigated macrolide antibiotics in low mu g mL(-1) concentration ranges with relative standard deviations (RSDs) lower than 6% and achieved detection limits (1.0Ds) as 0.18, 0.045, 1.43 and 0.30 mu g mL(-1) for EES, AZI, CLA and ROX, respectively. In the case of AZ1 and ROX, it was demonstrated that the use of AuNPs/CPE as working electrode could additionally improve the results obtained with the SWV method concerning exhibited sensitivity, reproducibility and linear concentration range, due to the electrocatalytic properties of synthesized AuNPs. The optimized experimental parameters with the use of SWV method and CPF. or AuNPs/CPE were successfully applied for determination of ROX and AZI in their pharmaceutical preparations Runac (R) and Ilemomycine, respectively. The reliability of the elaborated procedures and thus the accuracy of the obtained results were validated by comparing them with those obtained by means of H PLC-DAD measurements.

In this work, carbon paste electrode (CPE) and carbon paste electrode modified with gold nanoparticics (AuNPs/CPE) were employed with rapid direct anodic square wave voltammetry (SWV) for macrolide antibiotics erythromycin cthylsuccinate (EIS), azithromycin (A7.1), clarithromycin (OA) and roxithromycin (ROX) determination. The surface of working electrodes was morphologically characterized by scanning electron microscopic (SEM) measurements whereby the presence of randomly distributed AuNPs of 10 nm diameter size on CPE surface was confirmed by energy dispersive spectrometer (EDS). SWV determination of four macrolide antibiotics using CPE was performed in aqueous Britton-Robinson buffer solutions (pH 2.0 to 11.98) showing that oxidation signals strongly depend upon pH (at pH gt = 6) and exhibited the most favorable characteristics at pH 8.0 in the case of EES and pH 11.98 in the case of AZI, CIA and ROX. Under optimized conditions, the SWV method using CPE enables determination of all investigated macrolide antibiotics in low mu g mL(-1) concentration ranges with relative standard deviations (RSDs) lower than 6% and achieved detection limits (1.0Ds) as 0.18, 0.045, 1.43 and 0.30 mu g mL(-1) for EES, AZI, CLA and ROX, respectively. In the case of AZ1 and ROX, it was demonstrated that the use of AuNPs/CPE as working electrode could additionally improve the results obtained with the SWV method concerning exhibited sensitivity, reproducibility and linear concentration range, due to the electrocatalytic properties of synthesized AuNPs. The optimized experimental parameters with the use of SWV method and CPF. or AuNPs/CPE were successfully applied for determination of ROX and AZI in their pharmaceutical preparations Runac (R) and Ilemomycine, respectively. The reliability of the elaborated procedures and thus the accuracy of the obtained results were validated by comparing them with those obtained by means of H PLC-DAD measurements.

In this work, carbon paste electrode (CPE) and carbon paste electrode modified with gold nanoparticics (AuNPs/CPE) were employed with rapid direct anodic square wave voltammetry (SWV) for macrolide antibiotics erythromycin cthylsuccinate (EIS), azithromycin (A7.1), clarithromycin (OA) and roxithromycin (ROX) determination. The surface of working electrodes was morphologically characterized by scanning electron microscopic (SEM) measurements whereby the presence of randomly distributed AuNPs of 10 nm diameter size on CPE surface was confirmed by energy dispersive spectrometer (EDS). SWV determination of four macrolide antibiotics using CPE was performed in aqueous Britton-Robinson buffer solutions (pH 2.0 to 11.98) showing that oxidation signals strongly depend upon pH (at pH gt = 6) and exhibited the most favorable characteristics at pH 8.0 in the case of EES and pH 11.98 in the case of AZI, CIA and ROX. Under optimized conditions, the SWV method using CPE enables determination of all investigated macrolide antibiotics in low mu g mL(-1) concentration ranges with relative standard deviations (RSDs) lower than 6% and achieved detection limits (1.0Ds) as 0.18, 0.045, 1.43 and 0.30 mu g mL(-1) for EES, AZI, CLA and ROX, respectively. In the case of AZ1 and ROX, it was demonstrated that the use of AuNPs/CPE as working electrode could additionally improve the results obtained with the SWV method concerning exhibited sensitivity, reproducibility and linear concentration range, due to the electrocatalytic properties of synthesized AuNPs. The optimized experimental parameters with the use of SWV method and CPF. or AuNPs/CPE were successfully applied for determination of ROX and AZI in their pharmaceutical preparations Runac (R) and Ilemomycine, respectively. The reliability of the elaborated procedures and thus the accuracy of the obtained results were validated by comparing them with those obtained by means of H PLC-DAD measurements.

In this work, carbon paste electrode (CPE) and carbon paste electrode modified with gold nanoparticics (AuNPs/CPE) were employed with rapid direct anodic square wave voltammetry (SWV) for macrolide antibiotics erythromycin cthylsuccinate (EIS), azithromycin (A7.1), clarithromycin (OA) and roxithromycin (ROX) determination. The surface of working electrodes was morphologically characterized by scanning electron microscopic (SEM) measurements whereby the presence of randomly distributed AuNPs of 10 nm diameter size on CPE surface was confirmed by energy dispersive spectrometer (EDS). SWV determination of four macrolide antibiotics using CPE was performed in aqueous Britton-Robinson buffer solutions (pH 2.0 to 11.98) showing that oxidation signals strongly depend upon pH (at pH gt = 6) and exhibited the most favorable characteristics at pH 8.0 in the case of EES and pH 11.98 in the case of AZI, CIA and ROX. Under optimized conditions, the SWV method using CPE enables determination of all investigated macrolide antibiotics in low mu g mL(-1) concentration ranges with relative standard deviations (RSDs) lower than 6% and achieved detection limits (1.0Ds) as 0.18, 0.045, 1.43 and 0.30 mu g mL(-1) for EES, AZI, CLA and ROX, respectively. In the case of AZ1 and ROX, it was demonstrated that the use of AuNPs/CPE as working electrode could additionally improve the results obtained with the SWV method concerning exhibited sensitivity, reproducibility and linear concentration range, due to the electrocatalytic properties of synthesized AuNPs. The optimized experimental parameters with the use of SWV method and CPF. or AuNPs/CPE were successfully applied for determination of ROX and AZI in their pharmaceutical preparations Runac (R) and Ilemomycine, respectively. The reliability of the elaborated procedures and thus the accuracy of the obtained results were validated by comparing them with those obtained by means of H PLC-DAD measurements.

Natural products are rich resources for drug discovery and development. However, the complex molecular structures of natural products always complicate the total synthesis, structure–activity relationship investigations and structural optimizations. Therefore, simplifying complex structures without decreasing the desired biological activity is an effective strategy for improving synthetic accessibility and accelerating the drug development process. Amphidinolide C congeners are the cytotoxic macrolides isolated from the genus Amphidinium. Except for amphidinolide C4, they possess the same functionalized 25-membered macrolactone core and differ only on the side chain appended at C28. The 25-membered macrolactone core is decorated with some unusual structural components, such as an s-cis-1,3-diene subunit, an exo-methlyene moiety, and two trans-substituted tetrahydrofuran units. These natural compounds exhibit significant cytotoxic activities and they have attracted much interest from the synthetic community due to their remarkable bioactivities, low natural accessibility, and synthetic challenges for more than 20 years since their isolations. Total synthesis of amphidinolide C1 and F has been achieved by the groups of Carter, Fürstner and Figadère. However, up to now, the action mode of the amphidinolide C congeners is still unknown. Based on the limited information on the bioactivity of amphidinolide C congeners, we envisioned synthesis of the structurally simplified amphidinolide C congeners by depleting four stereogenic centers at C4, C12, C13, and C16. Three fragments were prepared and assembled via the Suzuki-Miyaura cross-coupling reaction under the Johnson conditions at C9-C10, esterification of C24-OH, and ring-closing metathesis at C16-C17. Then, the side chain was assembled onto the macrolactone core via Suzuki–Miyaura cross-coupling reaction of the C-26 alkenyl iodide by using our Aphos-Y as the ligand. In Chapter 1, a brief introduction of the structural

Bacterial resistance has become a major health threat that needs urgent action to maintain the advances of modern medicine. There is an urgent need to discover new bioactive natural compounds such as decalin-forming macrolides which have antibacterial potency and unique mode of action. Genome mining has become an important method to identify novel candidates of natural products through an efficient computational frameworks which utilize algorithms such as antiSMASH, BiG-SCPAE And CORASON. Through analysing ~15% of all available bacterial genomes, we have identified several potential new decalin-forming macrolide biosynthetic gene clusters. The most promising candidate is a BGC within the genome of Streptomyces tsukubaensis which contains all PKS genes of nargenicin and streptoseomycin and maintains the overall modular structure.

Cytochromes P450 (P450s) are nature's catalysts of choice for performing demanding and physiologically vital oxidation reactions. Biochemical characterization of these enzymes over the past decades has provided detailed mechanistic insight and highlighted the diversity of substrates P450s accommodate and the spectrum of oxidative transformations they catalyze. Previously, we discovered that the bacterial P450 MycCI from the mycinamicin biosynthetic pathway in Micromonospora griseorubida possesses an unusually broad substrate scope, whereas the homologous P450 from tylosin-producing Streptomyces fradiae (TylHI) exhibits a high degree of specificity for its native substrate. Here, using biochemical, structural, and computational approaches, we aimed to understand the molecular basis for the disparate reactivity profiles of these two P450s. Turnover and equilibrium binding experiments with substrate analogs revealed that TylHI strictly prefers 16-membered ring macrolides bearing the deoxyamino sugar mycaminose. To help rationalize these results, we solved the X-ray crystal structure of TylHI in complex with its native substrate at 1.99-Å resolution and assayed several site-directed mutants. We also conducted molecular dynamics simulations of TylHI and MycCI and biochemically characterized a third P450 homolog from the chalcomycin biosynthetic pathway in Streptomyces bikiniensis These studies provided a basis for constructing P450 chimeras to gain further insight into the features dictating the differences in reaction profile among these structurally and functionally related enzymes, ultimately unveiling the central roles of key loop regions in influencing substrate binding and turnover. Our work highlights the complex nature of P450/substrate interactions and raises interesting questions regarding the evolution of functional diversity among biosynthetic enzymes.

Copyright © 2019 by the American Thoracic Society Rationale: The macrolide antibiotic azithromycin reduces exacerbations in adults with persistent symptomatic asthma. However, owing to the pleotropic properties of macrolides, unintended bacteriological consequences such as augmented pathogen colonization or dissemination of antibiotic-resistant organisms can occur, calling into question the long-term safety of azithromycin maintenance therapy. Objectives: To assess the effects of azithromycin on the airway microbiota, pathogen abundance, and carriage of antibiotic resistance genes. Methods: 16S rRNA sequencing and quantitative PCR were performed to assess the effect of azithromycin on sputum microbiology from participants of the AMAZES (Asthma and Macrolides: The Azithromycin Efficacy and Safety) trial: a 48-week, double-blind, placebo-controlled trial of thrice-weekly 500 mg oral azithromycin in adults with persistent uncontrolled asthma. Pooled-template shotgun metagenomic sequencing, quantitative PCR, and isolate whole-genome sequencing were performed to assess antibiotic resistance. Measurements and Main Results: Paired sputum samples were available from 61 patients (n = 34 placebo, n = 27 azithromycin). Azithromycin did not affect bacterial load (P = 0.37) but did significantly decrease Faith’s phylogenetic diversity (P = 0.026) and Haemophilus influenzae load (P, 0.0001). Azithromycin did not significantly affect levels of Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, or Moraxella catarrhalis. Of the 89 antibiotic resistance genes detected, five macrolide resistance genes and two tetracycline resistance genes were increased significantly. Conclusions: In patients with persistent uncontrolled asthma, azithromycin reduced airway H. influenzae load compared with placebo but did not change total bacterial load. Macrolide resistance increased, reflecting previous studies. These results highlight the need for studies assessing the efficacy

The renewable silver-amalgam film electrode (Hg(Ag)FE) was applied for voltammetric characterization and determination of semi-synthetic macrolide antibiotics azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) in the Britton-Robinson buffer as supporting electrolyte ranging the pH from 4.0 to 11.9. All three macrolides showed reduction signals in fairly negative potential range. During direct cathodic square wave voltammetric (SWV) investigations conducted over the potential range from −0.75 V to −2.00 V vs SCE, either one or two reduction peaks were obtained in the potential range from −1.5 to −1.9 V. The shapes and intensities of the signals depend on the applied pH values in wider pH ranges. For analytical purposes concerning the development of direct cathodic SWV and adsorptive stripping SWV (SW-AdSV) methods the neutral and slightly alkaline media were suitable as pH 7.2, pH 7.4 and pH 7.0 for AZI, CLA and ROX, respectively. Based on the cyclic voltammograms recorded at these pH values, adsorption-controlled electrode kinetics process can be proposed for all three macrolides. Furthermore, the water suppressed 1H NMR measurements in the pH range between 6.0 and 10.5 indicated that the macrolide molecules at the optimal analytical conditions are predominantly in protonated form via their tertiary amino groups which supported in all three cases their adsorption on the appropriately polarized Hg(Ag)FE electrode. The optimized direct cathodic SWV methods showed good linearity in concentration ranges 4.81–23.3 μg mL−1, 1.96–28.6 μg mL−1 and 1.48–25.9 μg mL−1 for AZI, CLA and ROX, respectively. The development of the SW-AdSV methods resulted in the linear responses at lower concentration ranges as 1.0–2.46 μg mL−1, 0.05–0.99 μg mL−1 and 0.10–0.99 μg mL−1, for AZI, CLA and ROX, respectively. The relative standard deviation for all developed methods was not higher than 1.0% except the SWV method for AZI with 4.7%. In the case of all three investigated macr

Terrestrial and marine microorganisms provide a valuable source of structurally diverse and biologically active secondary metabolites. As introduced in Chapter 1, maltepolide A–F are a family of polyketide-derived 20-membered macrolides isolated from the fermentation broth of myxobacterium Sorangium cellulosum So ce1485. Most of maltepolides show moderate cytostatic activity on L929 mouse fibroblast cell lines with IC50 values of 4.6–39 μM. Maltepolide A and E cause unique morphological changes in the dividing transformed PtK2 cells with the phenotype closely resembling the effect of monastrol which is known as an inhibitor of the kinesin Eg5. Maltepolides possess either a THF moiety or a hydroxy vinyl epoxide within the macrocyclic core and the side chain appended at C19 contains either a vinyl epoxide or a corresponding epoxide ring-opening moiety by H2O/MeOH. It has been demonstrated that maltepolide E is the common precursor of the entire family. Total synthesis of the proposed structure of maltepolide C and synthesis of the related fragments have been reported but the structure of maltepolide C is confirmed incorrect. The main theme of this thesis work aims at establishment of a viable synthetic strategy for assembling maltepolide E possessing two sets of labile vinyl epoxide moieties, one at C11–C14 within the core and the other at C20–C23 on the side chain. In order to simplify our synthetic planning, this thesis work focuses on synthesis of the truncated maltepolide E without the C20–C24 side chain via 1,3-diene–ene ring-closing metathesis (RCM). After brief introduction of the reported examples of 1,3-diene–ene RCM in total synthesis in Chapter 1, our strategic consideration and the retrosynthetic analysis of the maltepolide E core are presented. Synthesis of the TBS-protected C1–C8 acid fragment and the C9–C19 alcohol fragment are compiled in Chapter 2. Of particular note are the DDQ-mediated over-oxidation of a dienyl alco

The renewable silver-amalgam film electrode (Hg(Ag)FE) was applied for voltammetric characterization and determination of semi-synthetic macrolide antibiotics azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) in the Britton-Robinson buffer as supporting electrolyte ranging the pH from 4.0 to 11.9. All three macrolides showed reduction signals in fairly negative potential range. During direct cathodic square wave voltammetric (SWV) investigations conducted over the potential range from -0.75 V to -2.00V vs SCE, either one or two reduction peaks were obtained in the potential range from -1.5 to -1.9 V. The shapes and intensities of the signals depend on the applied pH values in wider pH ranges. For analytical purposes concerning the development of direct cathodic SWV and adsorptive stripping SWV (SW-AdSV) methods the neutral and slightly alkaline media were suitable as pH 7.2, pH 7.4 and pH 7.0 for AZI, CIA and ROX, respectively. Based on the cyclic voltammograms recorded at these pH values, adsorption-controlled electrode kinetics process can be proposed for all three macrolides. Furthermore, the water suppressed H-1 NMR measurements in the pH range between 6.0 and 10.5 indicated that the macrolide molecules at the optimal analytical conditions are predominantly in protonated form via their tertiary amino groups which supported in all three cases their adsorption on the appropriately polarized Hg(Ag)FE electrode. The optimized direct cathodic SWV methods showed good linearity in concentration ranges 4.81-23.3 mu gmL(-1), 1.96-28.6 mu gmL(-1) and 1.48-25.9 mu gmL(-1) for AZI, CIA and ROX, respectively. The development of the SW-AdSV methods resulted in the linear responses at lower concentration ranges as 1.0-2.46 mu gmL(-1), 0.05-0.99 mu gmL(-1) and 0.10-0.99 mu gmL(-1), for AZI, CIA and ROX, respectively. The relative standard deviation for all developed methods was not higher than 1.0% except the SWV method for AZI with 4.7%. In the case of all three

The renewable silver-amalgam film electrode (Hg(Ag)FE) was applied for voltammetric characterization and determination of semi-synthetic macrolide antibiotics azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) in the Britton-Robinson buffer as supporting electrolyte ranging the pH from 4.0 to 11.9. All three macrolides showed reduction signals in fairly negative potential range. During direct cathodic square wave voltammetric (SWV) investigations conducted over the potential range from -0.75 V to -2.00V vs SCE, either one or two reduction peaks were obtained in the potential range from -1.5 to -1.9 V. The shapes and intensities of the signals depend on the applied pH values in wider pH ranges. For analytical purposes concerning the development of direct cathodic SWV and adsorptive stripping SWV (SW-AdSV) methods the neutral and slightly alkaline media were suitable as pH 7.2, pH 7.4 and pH 7.0 for AZI, CIA and ROX, respectively. Based on the cyclic voltammograms recorded at these pH values, adsorption-controlled electrode kinetics process can be proposed for all three macrolides. Furthermore, the water suppressed H-1 NMR measurements in the pH range between 6.0 and 10.5 indicated that the macrolide molecules at the optimal analytical conditions are predominantly in protonated form via their tertiary amino groups which supported in all three cases their adsorption on the appropriately polarized Hg(Ag)FE electrode. The optimized direct cathodic SWV methods showed good linearity in concentration ranges 4.81-23.3 mu gmL(-1), 1.96-28.6 mu gmL(-1) and 1.48-25.9 mu gmL(-1) for AZI, CIA and ROX, respectively. The development of the SW-AdSV methods resulted in the linear responses at lower concentration ranges as 1.0-2.46 mu gmL(-1), 0.05-0.99 mu gmL(-1) and 0.10-0.99 mu gmL(-1), for AZI, CIA and ROX, respectively. The relative standard deviation for all developed methods was not higher than 1.0% except the SWV method for AZI with 4.7%. In the case of all three

The renewable silver-amalgam film electrode (Hg(Ag)FE) was applied for voltammetric characterization and determination of semi-synthetic macrolide antibiotics azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) in the Britton-Robinson buffer as supporting electrolyte ranging the pH from 4.0 to 11.9. All three macrolides showed reduction signals in fairly negative potential range. During direct cathodic square wave voltammetric (SWV) investigations conducted over the potential range from -0.75 V to -2.00V vs SCE, either one or two reduction peaks were obtained in the potential range from -1.5 to -1.9 V. The shapes and intensities of the signals depend on the applied pH values in wider pH ranges. For analytical purposes concerning the development of direct cathodic SWV and adsorptive stripping SWV (SW-AdSV) methods the neutral and slightly alkaline media were suitable as pH 7.2, pH 7.4 and pH 7.0 for AZI, CIA and ROX, respectively. Based on the cyclic voltammograms recorded at these pH values, adsorption-controlled electrode kinetics process can be proposed for all three macrolides. Furthermore, the water suppressed H-1 NMR measurements in the pH range between 6.0 and 10.5 indicated that the macrolide molecules at the optimal analytical conditions are predominantly in protonated form via their tertiary amino groups which supported in all three cases their adsorption on the appropriately polarized Hg(Ag)FE electrode. The optimized direct cathodic SWV methods showed good linearity in concentration ranges 4.81-23.3 mu gmL(-1), 1.96-28.6 mu gmL(-1) and 1.48-25.9 mu gmL(-1) for AZI, CIA and ROX, respectively. The development of the SW-AdSV methods resulted in the linear responses at lower concentration ranges as 1.0-2.46 mu gmL(-1), 0.05-0.99 mu gmL(-1) and 0.10-0.99 mu gmL(-1), for AZI, CIA and ROX, respectively. The relative standard deviation for all developed methods was not higher than 1.0% except the SWV method for AZI with 4.7%. In the case of all three

The renewable silver-amalgam film electrode (Hg(Ag)FE) was applied for voltammetric characterization and determination of semi-synthetic macrolide antibiotics azithromycin (AZI), clarithromycin (CLA) and roxithromycin (ROX) in the Britton-Robinson buffer as supporting electrolyte ranging the pH from 4.0 to 11.9. All three macrolides showed reduction signals in fairly negative potential range. During direct cathodic square wave voltammetric (SWV) investigations conducted over the potential range from -0.75 V to -2.00V vs SCE, either one or two reduction peaks were obtained in the potential range from -1.5 to -1.9 V. The shapes and intensities of the signals depend on the applied pH values in wider pH ranges. For analytical purposes concerning the development of direct cathodic SWV and adsorptive stripping SWV (SW-AdSV) methods the neutral and slightly alkaline media were suitable as pH 7.2, pH 7.4 and pH 7.0 for AZI, CIA and ROX, respectively. Based on the cyclic voltammograms recorded at these pH values, adsorption-controlled electrode kinetics process can be proposed for all three macrolides. Furthermore, the water suppressed H-1 NMR measurements in the pH range between 6.0 and 10.5 indicated that the macrolide molecules at the optimal analytical conditions are predominantly in protonated form via their tertiary amino groups which supported in all three cases their adsorption on the appropriately polarized Hg(Ag)FE electrode. The optimized direct cathodic SWV methods showed good linearity in concentration ranges 4.81-23.3 mu gmL(-1), 1.96-28.6 mu gmL(-1) and 1.48-25.9 mu gmL(-1) for AZI, CIA and ROX, respectively. The development of the SW-AdSV methods resulted in the linear responses at lower concentration ranges as 1.0-2.46 mu gmL(-1), 0.05-0.99 mu gmL(-1) and 0.10-0.99 mu gmL(-1), for AZI, CIA and ROX, respectively. The relative standard deviation for all developed methods was not higher than 1.0% except the SWV method for AZI with 4.7%. In the case of all three

Background: Long-term use of macrolide antibiotics is effective to prevent exacerbations in chronic obstructive pulmonary disease (COPD). As risks and side effects of long-term intervention outweigh the benefits in the general COPD population, the optimal dose, duration of treatment, and target population are yet to be defined. Hospitalization for an acute exacerbation (AE) of COPD may offer a targeted risk group and an obvious risk period for studying macrolide interventions. Methods/design: Patients with COPD, hospitalized for an AE, who have a smoking history of ≥10 pack-years and had ≥1 exacerbation in the previous year will be enrolled in a multicenter, randomized, double-blind, placebo-controlled trial (NCT02135354). On top of a standardized treatment of systemic corticosteroids and antibiotics, subjects will be randomized to receive either azithromycin or placebo during 3 months, at an uploading dose of 500 mg once a day for 3 days, followed by a maintenance dose of 250 mg once every 2 days. The primary endpoint is the time-to-treatment failure during the treatment phase (ie, from the moment of randomization until the end of intervention). Treatment failure is a novel composite endpoint defined as either death, the admission to intensive care or the requirement of additional systemic steroids or new antibiotics for respiratory reasons, or the diagnosis of a new AE after discharge. Discussion: We investigate whether azithromycin initiated at the onset of a severe exacerbation, with a limited duration and at a low dose, might be effective and safe in the highest risk period during and immediately after the acute event. If proven effective and safe, this targeted approach may improve the treatment of severe AEs and redirect the preventive use of azithromycin in COPD to a temporary intervention in the subgroup with the highest unmet needs., SCOPUS: ar.j, info:eu-repo/semantics/published

Lanthanides have provided many exciting opportunities in the metal-catalyzed reactions and reductive chemistry in organic chemistry area, with samarium diiodide (SmI2) being the most popular reagent. This thesis research deals with the acrylates and crotonates possessing an electron-withdrawing achiral or chiral aryloxy group for the SmI2-mediated reductive coupling with aliphatic aldehydes. The products, α,γ-trans- and β,γ-cis-disubstituted y-butyrolactones, obtained in high stereoselectivity and yields, are the versatile precursors to trisubstituted tetrahydrofurans and other stereochemically defined building blocks. Applications in diverted total synthesis of natural macrolides and designed analogues are demonstrated. A brief overview is given in Chapter 1 about target-oriented synthesis (TOS), diversity-oriented synthesis (DOS), and diverted total synthesis (DTS), and selective examples of the SmI2-mediated reductive coupling reactions used in total synthesis. It is followed by the structural analysis of apratoxin A, palmyrolide A, and laingolides consisting of a unique 1,3-anti-Me/t-Bu subunit, and the progress in total synthesis of apratoxin A and palmyrolide A. Chapter 2 summarizes in more details the representative types of reductive carbon-carbon bond-forming reactions mediated by SmI2 and the two proposed single-electron transfer pathways for reductive coupling of acrylates/crotonates with carbonyl compounds. Our new results on SmI2-mediated reductive coupling reactions of α-substituted acrylates in achiral and chiral forms are then complied for efficient formation of trans-α,γ-disubstituted γ-butyrolactones in high stereoselectivity and high chemical yields. Particularly, racemic or chiral form of trans-5-tert-butyl-3-methyldihydrofuran-2-one serves as the synthetic precursor to 1,3-anti-Me/t-Bu subunit. Usefulness of the SmI2-mediated reductive coupling of the chiral crotonate consisting o

The Swedish Medical Product Agency (MPA) has listed erythromycin as a suggested human teratogen, causing cardiovascular malformations. It is further suggested that this may be a class effect of macrolide antibiotics. The proposed teratogenic mechanism is blockade of the human ether-a-go-go-related (hERG)/I-Kr current in the embryonic heart causing bradycardia and arrhythmia resulting in altered cardiac blood flow and/or embryonic hypoxia. To test this hypothesis, we examined the effect of three macrolide antibiotics on the function of the rat embryonic heart. Gestational day 13 rat embryos in vitro were exposed to erythromycin (25-500 mu M), clarithromycin (25-500 mu M), or azithromycin (100 mu M to 1mM) for 3 hr. The effect on the embryonic heart was monitored every hour. The results showed that erythromycin and clarithromycin caused a concentration-dependent bradycardia. Twenty-five micromolar was a no-effect concentration for erythromycin and was close to a no-effect concentration for clarithromycin. Azithromycin only caused significant bradycardia at 1mM. Additional studies were performed with the embryos cultured at 40 degrees C instead of 38 degrees C, to mimic fever. The increased temperature increased the number of arrhythmias but did not worsen the drug-induced bradycardia. The results support the concept that erythromycin and clarithromycin can adversely affect the embryonic heart but only at concentrations well outside expected embryonic exposure in the human

Background: Chronic obstructive pulmonary disease (COPD) is characterised by progressive development of airflow limitation that is poorly reversible. Because of a poor understanding of COPD pathogenesis, treatment is mostly symptomatic and new therapeutic strategies are limited. There is a direct relationship between the severity of the disease and the intensity of the inflammatory response. Besides smoking, one of the hypotheses for the persistent airway inflammation is the presence of recurrent infections. Macrolide antibiotics have bacteriostatic as well as anti-inflammatory properties in patients with cystic fibrosis and other inflammatory pulmonary diseases. There is consistent evidence that macrolide therapy reduces infectious exacerbations, decreases the requirement for additional antibiotics and improves nutritional measures. Because of these positive effects we hypothesised that maintenance macrolide therapy may also have beneficial effects in patients with COPD who have recurrent exacerbations. The effects on development of bacterial resistance to macrolides due to this long-term treatment are unknown. Until now, studies investigating macrolide therapy in COPD are limited. The objective of this study is to assess whether maintenance treatment with macrolide antibiotics in COPD patients with three or more exacerbations in the previous year decreases the exacerbation rate in the year of treatment and to establish microbial resistance due to the long-term treatment.Methods/design: The study is set up as a prospective randomised double-blind placebo-controlled single-centre trial. A total of 92 patients with COPD who have had at least three exacerbations of COPD in the previous year will be included. Subjects will be randomised to receive either azithromycin 500 mg three times a week or placebo. Our primary endpoint is the reduction in the number of exacerbations of COPD in the year of treatment.Discussion: We investigate whether long-term therapy with macroli

Schapen, die worden behandeld met tilmicosine tegen rotkreupel en daarvan te veel krijgen, kunnen doodgaan. Bij dit geneesmiddel is er al gauw sprake van een overdosis.

Cilj ove disertacije bio je da se ispitivanjem elektrohemijske aktivnosti makrolidnih antibiotika roksitromicina i midekamicina i poredjenjem dobijenih rezultata sa rezultatima ranije ispitanih azitromicina, klaritromicina i eritromicina izvede opšti zaključak o elektrohemijskom ponašanju ispitanih makrolidnih antibiotika, njihovim mogućim elektrohemijskim transformacijama kao i o mogućnosti njihovog kvalitativnog i kvantitativnog odredjivanja. Posebno je razmatrana mogućnost primene novih voltametrijskih metoda za odredjivanja makrolidnih antibiotika kao sastojaka komercijalnih i bioloških uzoraka. Na osnovu detaljnog ispitivanja elektrohemijskog ponašanja roksitromicina metodama ciklične voltametrije i diferencijalne pulsne voltametrije i midekamicina metodom ciklične voltametrije na elektrodi od zlata, simultano praćeno HPLC i LC/MS metodama kvalitativno i kvantitativno analitički su određene farmaceutske supstance roksitromicin i midekamicin, a roksitromicin i kao sastojak komercijalnog roksitromicina (Runac® tablete) i biološke tečnosti (humani urin). Poredjenjem sa ranije ispitivanim makrolidnim antibioticima pod istim uslovima, utvrdjeno je da su azitromicin, roksitromicin i midekamicin kvalitativno i kvantitativno analitički određeni. Eritromicin i klaritromicin su određeni kvalitativno a transformacije kojima podležu, za razliku od azitromicina, roksitromicina i midekamicina su praćene FTIR i HPLC metodama., The aim of the present PhD thesis was to examinate the electrochemical activity of macrolide antibiotics roxithromycin and midecamyin and to compare with the previously investigated azithromycin, clarithromycin and erithromycin in order to have general conclusion concerning electrochemical behavior of macrolide antibiotics. That means, their possible electrochemical transformations as well as the possibility of their quantitative and qualitative determination. Separetely was analysed the possibility of the new voltammetric methods application for the determination of macrolide antibiotics and as a content of commercial an biological samples. Taking into account the detailed investigation of the electrochemical behavior of roxithromycin by cyclic voltammetry and differential pulse voltammetry and a midecamyin by cyclic voltammetry on gold electrode by simultaneous detection of antibiotics by HPLC and LC/MS methods. Their standards were quantitatively and qualitatively were determined and roxithromycin was determined in the same manner and as a content of commercial tablets Runac® and as content of humane urine. The comparison with the previously investigated macrolide antibiotics under the identical experimental conditions, shown that azithromycin, roxithromycin and midecamyin are succesfully quantitatively and qualitatively determined. Clarithromycin and erithromycin undergo to the electrochemical transformations during the qualitative determination and these transformations were simultaneosly followed by HPLC and FTIR methods.

Using the cyclic voltammetry, it has been shown that hydrogen evolution at a gold electrode is necessary in the electrochemical activation of azithromycin dihydrate and erythromycin A. After four hours of the potential holding at -1.2 V vs. SCE, the pH of the electrolyte has been changed from 8.40 to 8.96; from 8.40 to 8.77 in the presence of erythromycin A, and from 8.40 to 9.18 in the presence of azithromycin, indicating the reaction of the hydrogen species with antibiotics. This effect has been confirmed by using the phenolphthalein indicator and by analysing colours of the solutions by UV-Vis, as well as by FTIR spectroscopy. Under the identical experimental conditions at the gold electrode, in contrast to azithromycin dihydrate and erythromycin A, roxithromycin and midecamycin electroactivity promotion has been obtained during the first forward sweep starting from the area of a double layer region., Metodom ciklične voltametrije je pokazano da je izdvajanje vodonika na elektrodi od zlata neophodno za elektrohemijsku aktivaciju azitromicin-dihidrata i eritromicina A. Četiri sata držanja potencijala na vrednosti 1,2 V vs. SCE je rezultovalo u promeni pH elektrolita sa 8,40 na 8,96; sa 8,40 na 8,77 u prisustvu eritromicina A, i sa 8,40 na 9,18 u prisustvu azitromicina, ukazujući na reakciju čestica izdvojenog vodonika sa antibioticima. Efekat je potvrđen upotrebom fenolftalein indikatora i analizom boje elektrolita UV-Vis spektroskopijom, a molekula antibiotika FTIR spektroskopijom. Pod istim eksperimentalnim uslovima na elektrodi od zlata, u suprotnosti sa ponašanjem azitromicin-dihidrata i eritromicina A, elektroaktivnost roksitromicina i midekamicina je uočena počevši od oblasti formiranja dvojnog sloja, tj. bez ikakvog učešća čestica izdvojenog vodonika.

Using the cyclic voltammetry, it has been shown that hydrogen evolution at a gold electrode is necessary in the electrochemical activation of azithromycin dihydrate and erythromycin A. After four hours of the potential holding at -1.2 V vs. SCE, the pH of the electrolyte has been changed from 8.40 to 8.96; from 8.40 to 8.77 in the presence of erythromycin A, and from 8.40 to 9.18 in the presence of azithromycin, indicating the reaction of the hydrogen species with antibiotics. This effect has been confirmed by using the phenolphthalein indicator and by analysing colours of the solutions by UV-Vis, as well as by FTIR spectroscopy. Under the identical experimental conditions at the gold electrode, in contrast to azithromycin dihydrate and erythromycin A, roxithromycin and midecamycin electroactivity promotion has been obtained during the first forward sweep starting from the area of a double layer region., Metodom ciklične voltametrije je pokazano da je izdvajanje vodonika na elektrodi od zlata neophodno za elektrohemijsku aktivaciju azitromicin-dihidrata i eritromicina A. Četiri sata držanja potencijala na vrednosti 1,2 V vs. SCE je rezultovalo u promeni pH elektrolita sa 8,40 na 8,96; sa 8,40 na 8,77 u prisustvu eritromicina A, i sa 8,40 na 9,18 u prisustvu azitromicina, ukazujući na reakciju čestica izdvojenog vodonika sa antibioticima. Efekat je potvrđen upotrebom fenolftalein indikatora i analizom boje elektrolita UV-Vis spektroskopijom, a molekula antibiotika FTIR spektroskopijom. Pod istim eksperimentalnim uslovima na elektrodi od zlata, u suprotnosti sa ponašanjem azitromicin-dihidrata i eritromicina A, elektroaktivnost roksitromicina i midekamicina je uočena počevši od oblasti formiranja dvojnog sloja, tj. bez ikakvog učešća čestica izdvojenog vodonika.

Using the cyclic voltammetry, it has been shown that hydrogen evolution at a gold electrode is necessary in the electrochemical activation of azithromycin dihydrate and erythromycin A. After four hours of the potential holding at -1.2 V vs. SCE, the pH of the electrolyte has been changed from 8.40 to 8.96; from 8.40 to 8.77 in the presence of erythromycin A, and from 8.40 to 9.18 in the presence of azithromycin, indicating the reaction of the hydrogen species with antibiotics. This effect has been confirmed by using the phenolphthalein indicator and by analysing colours of the solutions by UV-Vis, as well as by FTIR spectroscopy. Under the identical experimental conditions at the gold electrode, in contrast to azithromycin dihydrate and erythromycin A, roxithromycin and midecamycin electroactivity promotion has been obtained during the first forward sweep starting from the area of a double layer region., Metodom ciklične voltametrije je pokazano da je izdvajanje vodonika na elektrodi od zlata neophodno za elektrohemijsku aktivaciju azitromicin-dihidrata i eritromicina A. Četiri sata držanja potencijala na vrednosti 1,2 V vs. SCE je rezultovalo u promeni pH elektrolita sa 8,40 na 8,96; sa 8,40 na 8,77 u prisustvu eritromicina A, i sa 8,40 na 9,18 u prisustvu azitromicina, ukazujući na reakciju čestica izdvojenog vodonika sa antibioticima. Efekat je potvrđen upotrebom fenolftalein indikatora i analizom boje elektrolita UV-Vis spektroskopijom, a molekula antibiotika FTIR spektroskopijom. Pod istim eksperimentalnim uslovima na elektrodi od zlata, u suprotnosti sa ponašanjem azitromicin-dihidrata i eritromicina A, elektroaktivnost roksitromicina i midekamicina je uočena počevši od oblasti formiranja dvojnog sloja, tj. bez ikakvog učešća čestica izdvojenog vodonika.

Using the cyclic voltammetry, it has been shown that hydrogen evolution at a gold electrode is necessary in the electrochemical activation of azithromycin dihydrate and erythromycin A. After four hours of the potential holding at -1.2 V vs. SCE, the pH of the electrolyte has been changed from 8.40 to 8.96; from 8.40 to 8.77 in the presence of erythromycin A, and from 8.40 to 9.18 in the presence of azithromycin, indicating the reaction of the hydrogen species with antibiotics. This effect has been confirmed by using the phenolphthalein indicator and by analysing colours of the solutions by UV-Vis, as well as by FTIR spectroscopy. Under the identical experimental conditions at the gold electrode, in contrast to azithromycin dihydrate and erythromycin A, roxithromycin and midecamycin electroactivity promotion has been obtained during the first forward sweep starting from the area of a double layer region., Metodom ciklične voltametrije je pokazano da je izdvajanje vodonika na elektrodi od zlata neophodno za elektrohemijsku aktivaciju azitromicin-dihidrata i eritromicina A. Četiri sata držanja potencijala na vrednosti 1,2 V vs. SCE je rezultovalo u promeni pH elektrolita sa 8,40 na 8,96; sa 8,40 na 8,77 u prisustvu eritromicina A, i sa 8,40 na 9,18 u prisustvu azitromicina, ukazujući na reakciju čestica izdvojenog vodonika sa antibioticima. Efekat je potvrđen upotrebom fenolftalein indikatora i analizom boje elektrolita UV-Vis spektroskopijom, a molekula antibiotika FTIR spektroskopijom. Pod istim eksperimentalnim uslovima na elektrodi od zlata, u suprotnosti sa ponašanjem azitromicin-dihidrata i eritromicina A, elektroaktivnost roksitromicina i midekamicina je uočena počevši od oblasti formiranja dvojnog sloja, tj. bez ikakvog učešća čestica izdvojenog vodonika.

Inhalation of sulfur mustard (SM) causes airway inflammation and injury. There is increasing evidence of the effectiveness of macrolide antibiotics in treating chronic airway inflammatory diseases. In this study, the anti-cytotoxic and anti-inflammatory effects of a representative macrolide antibiotic, roxithromycin, were tested in vitro using SM-exposed normal human small airway epithelial (SAE) cells and bronchial/tracheal epithelial (BTE) cells. Cells were exposed to varying concentrations of SM with or without roxithromycin. Cytotoxicity was measured with the MTS assay and Calcein AM/ethidium homodimer (EthD-1) fluorescence staining. Cytokine protein expression was analyzed by enzyme-linked immunosorbent assay (ELISA) and cytokine mRNA expression by real-time RT-PCR. Inducible nitric oxide synthase (iNOS) protein expression was examined by a new immunocytochemical method using quantum dots as the fluorophore. Our results show that roxithromycin decreased SM cytotoxicity in both SAE and BTE cells. Also, roxithromycin inhibited the SM-stimulated overproduction of the proinflammatory cytokines interleukin (IL)-1 Beta, IL-6, IL-8 and TNF at both the mRNA level and the protein level. In addition, roxithromycin inhibited the SM-induced overexpression of iNOS. In conclusion, the present study demonstrates that roxithromycin has anti-cytotoxic and anti-inflammatory activities in human airway epithelial cells that likely depend on the ability of roxithromycin to down-regulate the production of proinflammatory mediators and cytokines, and suggests that macrolide antibiotics may serve as potential vesicant respiratory therapeutics through mechanisms independent of their antibacterial activity., See also ADM002075. Presented at the Army Science Conference (25th) held in Orlando, FL on 27-30 Nov 2006. Document includes briefing charts (24 slides, title same as report). The original document contains color images.

The chromatographic behavior of roxithromycin, midecamycin, erythromycin, azithromycin, and erythromycin ethylsuccinate has been studied by salting-out thin-layer chromatography (SOTLC) with cellulose as a adsorbent and aqueous ammonium sulfate solutions of different concentration as mobile phases. Hydrophobicity was determined from the linear relationships between solute R-M values and the ammonium sulfate content of the mobile phase. Hydrophobicity values R-M(0) and C-0 were calculated on the basis of the results obtained. Lipophilicity determined in this way was correlated with calculated log P values.

The chromatographic behavior of roxithromycin, midecamycin, erythromycin, azithromycin, and erythromycin ethylsuccinate has been studied by salting-out thin-layer chromatography (SOTLC) with cellulose as a adsorbent and aqueous ammonium sulfate solutions of different concentration as mobile phases. Hydrophobicity was determined from the linear relationships between solute R-M values and the ammonium sulfate content of the mobile phase. Hydrophobicity values R-M(0) and C-0 were calculated on the basis of the results obtained. Lipophilicity determined in this way was correlated with calculated log P values.

The chromatographic behavior of roxithromycin, midecamycin, erythromycin, azithromycin, and erythromycin ethylsuccinate has been studied by salting-out thin-layer chromatography (SOTLC) with cellulose as a adsorbent and aqueous ammonium sulfate solutions of different concentration as mobile phases. Hydrophobicity was determined from the linear relationships between solute R-M values and the ammonium sulfate content of the mobile phase. Hydrophobicity values R-M(0) and C-0 were calculated on the basis of the results obtained. Lipophilicity determined in this way was correlated with calculated log P values.

The projects described in this dissertation use cutting edge technology to quantitate various drugs in plasma. Highly sensitive assay methods for the quantification of clarithromycin, carbamazepine and its major metabolite, carbamazepine-l0, Il-epoxide in human plasma were developed and validated. The application of these methods to analyse samples generated during pharmacokinetic studies is also discussed where up to 230 samples were analysed per day. The methods were sensitive enough to quantitate the analytes for at least 5 half-lives of the drug after an oral dose to human volunteers. A sensitive method for the determination of clarithromycin, a macrolide antibiotic, USing high-performance liquid chromatographic separation with tandem mass spectrometry, and the method development thereof, are described in this dissertation. Samples were prepared using liquid-liquid extraction and were separated on a CI8 column with a mobile phase consisting of acetonitrile, methanol and acetic acid. Detection was performed by a mass speetrometer in the multiple reaction monitoring (MRM) mode, using TurboIonSpray ionisation. The mean recovery of clarithromycin was 87.3%, with a lower limit of quantification of 2.95 ng/ml when using 300f!1 plasma. This high-throughput method was used to quantify 230 samples per day, and is sufficiently sensitive to be employed in pharmacokinetic studies. Carbamazepine, an anticonvulsant, and its major metabolite, carbamazepine-10, l l-epóxide, were quantified using tandem liquid chromatography-mass spectrometry, with electrospray ionisation. Both these analytes were extracted from 500f!1 of human plasma, using a liquid-liquid extraction method that showed recoveries greater than 95% for both these analytes. Concentrations as low as 0.722 ng/ml for carbamazepine and 5.15 ng/ml for carbamazepine-10, 11- epoxide could be quantified by using this method. These are the first LC-MSIMS assay methods described for the quantification of carbamazepine and i

Summary: The bryostatins are a family of macrolide lactones isolated from the marine bryozoan Bugu/a neritina. Since its detection in 1968, bryostatin 1 has demonstrated remarkable anticancer, immunopotentiating, biomodulatory and radioprotective effects which result mainly from its ability to activate protein kinase C, a family of isozymes involved in cellular signal transduction. It is currently being tested in several phase I and phase II clinical trials as a potential anticancer drug for leukemia, melanoma and nephrotoma. A series of experiments was undertaken to elucidate the biosynthetic origins of bryostatin, using a fortified crude cell-free enzyme preparation and radiolabelled precursors. A regional characterization of Bugula neritina from Sicily, Italy and Daytona Beach, Florida is also described., 1997, Includes bibliography., Thesis (M.S.)--Florida Atlantic University, 1997., Charles E. Schmidt College of Science

Diplomová práce je zaměřena na stanovení makrolidových antibiotik v pitných vodách metodou vysokoúčinné kapalinové chromatografie. Z hlediska výběru antibiotik se jedná o erythromycin a clarithromycin, které se v současné době řadí mezi často předepisované léky. Pro zakoncentrování a přečištění vybraných analyzovaných složek z reálných vzorků pitných vod byla vybrána metoda extrakce tuhou fází (SPE) na kolonkách Oasis HLB. Optimalizace a konečná analýza vzorků byla prováděna na vysokoúčinném kapalinovém chromatografu s hmotnostně spektrometrickou detekcí (HPLC-MS). Vybraná metoda byla použita pro stanovení vybraných makrolidů v reálných vzorcích pitných vod odebíraných ze dvou zdrojů, z jímacího zařízení Litovel a Černovír Olomouc., The aim of the diploma thesis is the determination of macrolide antibiotics in drinking water by using of high performance liquid chromatography. Erythromycin and clarithromycin were selected such as representative macrolides due to frequently prescribed pharmaceuticals in this time. Solid phase extraction (SPE) by using of Oasis HLB cartridges was applied for pre concentration and purification of chosen analytes in real samples of drinking water. Optimalization of method and analysis were performed by using of high performance liquid chromatography with mass spectrometry detection (HPLC-MS). The suitable method was selected for determination of macrolides in real samples taken from two sources of drinking water, the interception of water in Litovel and Černovír Olomouc.

Diplomová práce se zabývá stanovením makrolidových antibiotik v odpadních vodách. Konkrétně se jedná o erythromycin, clarithromycin a roxithromycin, které jsou v současné době poměrně často předepisovanými léčivy. Pro izolaci a přečištění vybraných analytů z vodné matrice byla použita metoda extrakce tuhou fází (SPE); vhodný postup byl nalezen na kolonkách Oasis HLB. Pro vlastní analýzu vybraných léčiv byla optimalizována metoda na vysokoúčinném kapalinovém chromatografu s hmotnostně-spektrometrickou detekcí (HPLC–MS). Optimalizovaná metoda byla využita pro stanovení reálných vzorků vod odebraných na přítoku a odtoku městské čistírny odpadních vod Brno–Modřice., This diploma thesis deals with the determination of macrolide antibiotics in wastewater, especially with erythromycin, clarithromycin and roxithromycin. In this time are these pharmaceuticals prescribed quite frequently. Solid phase extraction (SPE) was used for the isolation and the purification of selected analytes from an aqueous matrix; as the suitable procedure was found the using Oasis HLB cartridges. High performance liquid chromatography with mass spectrometry detection (HPLC-MS) was optimized for its analysis of selected pharmaceuticals. The optimized method was used for the determination of pharmaceuticals in real water samples, which was taken at the inflow and the outflow of the urban wastewater treatment plant in Brno-Modřice.

Diplomová práce je zaměřena na stanovení makrolidových antibiotik v pitných vodách metodou vysokoúčinné kapalinové chromatografie. Z hlediska výběru antibiotik se jedná o erythromycin a clarithromycin, které se v současné době řadí mezi často předepisované léky. Pro zakoncentrování a přečištění vybraných analyzovaných složek z reálných vzorků pitných vod byla vybrána metoda extrakce tuhou fází (SPE) na kolonkách Oasis HLB. Optimalizace a konečná analýza vzorků byla prováděna na vysokoúčinném kapalinovém chromatografu s hmotnostně spektrometrickou detekcí (HPLC-MS). Vybraná metoda byla použita pro stanovení vybraných makrolidů v reálných vzorcích pitných vod odebíraných ze dvou zdrojů, z jímacího zařízení Litovel a Černovír Olomouc., The aim of the diploma thesis is the determination of macrolide antibiotics in drinking water by using of high performance liquid chromatography. Erythromycin and clarithromycin were selected such as representative macrolides due to frequently prescribed pharmaceuticals in this time. Solid phase extraction (SPE) by using of Oasis HLB cartridges was applied for pre concentration and purification of chosen analytes in real samples of drinking water. Optimalization of method and analysis were performed by using of high performance liquid chromatography with mass spectrometry detection (HPLC-MS). The suitable method was selected for determination of macrolides in real samples taken from two sources of drinking water, the interception of water in Litovel and Černovír Olomouc.

Diplomová práce se zabývá stanovením makrolidových antibiotik v odpadních vodách. Konkrétně se jedná o erythromycin, clarithromycin a roxithromycin, které jsou v současné době poměrně často předepisovanými léčivy. Pro izolaci a přečištění vybraných analytů z vodné matrice byla použita metoda extrakce tuhou fází (SPE); vhodný postup byl nalezen na kolonkách Oasis HLB. Pro vlastní analýzu vybraných léčiv byla optimalizována metoda na vysokoúčinném kapalinovém chromatografu s hmotnostně-spektrometrickou detekcí (HPLC–MS). Optimalizovaná metoda byla využita pro stanovení reálných vzorků vod odebraných na přítoku a odtoku městské čistírny odpadních vod Brno–Modřice., This diploma thesis deals with the determination of macrolide antibiotics in wastewater, especially with erythromycin, clarithromycin and roxithromycin. In this time are these pharmaceuticals prescribed quite frequently. Solid phase extraction (SPE) was used for the isolation and the purification of selected analytes from an aqueous matrix; as the suitable procedure was found the using Oasis HLB cartridges. High performance liquid chromatography with mass spectrometry detection (HPLC-MS) was optimized for its analysis of selected pharmaceuticals. The optimized method was used for the determination of pharmaceuticals in real water samples, which was taken at the inflow and the outflow of the urban wastewater treatment plant in Brno-Modřice.

Diplomová práce je zaměřena na stanovení makrolidových antibiotik v odpadních vodách. Kontaminace životního prostředí léčivy představuje v současnosti velký problém. Ke zjištění úrovně této kontaminace je nezbytný vývoj vhodných analytických technik. Ze skupiny makrolidových antibiotik byly vybrány čtyři zástupci, a to erythromycin, klarithromycin, azithromycin a roxithromycin, které patří v České republice k nejvíce aplikovaným. Pro jejich izolaci z odpadní vody byla zvolena extrakce tuhou fází a jako finální metoda pro identifikaci a kvantifikaci vysokoúčinná kapalinová chromatografie s hmotnostně spektrometrickou detekcí. Za optimalizovaných podmínek bylo provedeno desetidenní sledování přítoku a odtoku na ČOV Brno-Modřice, dvoudenní a jednodenní sledování přítoku a odtoku na ČOV Mikulov respektive ČOV Veterinární a farmaceutické univerzity Brno., Diploma thesis is focused on determination of macrolide antibiotics in wastewater. Environmental contamination with drugs currently represents a major problem. To determine the level of contamination is necessary to develop the appropriate analytical techniques. From the group of macrolide antibiotics were selected four representatives: erythromycin, clarithromycin, azithromycin and roxithromycin due to their frequent use in Czech Republic. For their isolation from wastewater was selected solid phase extraction and for analysis high performance liquid chromatography with mass spectrometric detection was chosen. Under optimized conditions, the ten days analysis of wastewater from the WWTP Brno-Modřice, two days analysis of wastewater from the WWTP Mikulov and one day analysis of wastewater from WWTP of University of Veterinary and Pharmaceutical Sciences Brno was performed.

Diplomová práce je zaměřena na stanovení makrolidových antibiotik v pitných vodách metodou vysokoúčinné kapalinové chromatografie. Z hlediska výběru antibiotik se jedná o erythromycin a clarithromycin, které se v současné době řadí mezi často předepisované léky. Pro zakoncentrování a přečištění vybraných analyzovaných složek z reálných vzorků pitných vod byla vybrána metoda extrakce tuhou fází (SPE) na kolonkách Oasis HLB. Optimalizace a konečná analýza vzorků byla prováděna na vysokoúčinném kapalinovém chromatografu s hmotnostně spektrometrickou detekcí (HPLC-MS). Vybraná metoda byla použita pro stanovení vybraných makrolidů v reálných vzorcích pitných vod odebíraných ze dvou zdrojů, z jímacího zařízení Litovel a Černovír Olomouc., The aim of the diploma thesis is the determination of macrolide antibiotics in drinking water by using of high performance liquid chromatography. Erythromycin and clarithromycin were selected such as representative macrolides due to frequently prescribed pharmaceuticals in this time. Solid phase extraction (SPE) by using of Oasis HLB cartridges was applied for pre concentration and purification of chosen analytes in real samples of drinking water. Optimalization of method and analysis were performed by using of high performance liquid chromatography with mass spectrometry detection (HPLC-MS). The suitable method was selected for determination of macrolides in real samples taken from two sources of drinking water, the interception of water in Litovel and Černovír Olomouc.