by Wong Yin Kwan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (leaves 105-115). / Abstract also in Chinese. / Title page --- p.i / Acknowledgements --- p.ii / Table of contents --- p.iii / Abbreviations --- p.v / Abstract --- p.vi / 摘要 --- p.xi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter Chapter 2 --- Assay Development for Ciprofloxacin / Chapter 2.1 --- Introduction --- p.23 / Chapter 2.2 --- HPLC Assay --- p.24 / Chapter 2.3 --- Microbiological Assay --- p.29 / Chapter 2.4 --- Discussion --- p.30 / Chapter Chapter 3 --- The Effects of Oral Ferrous Sulfate on Pharmacokinetics and Pharmacodynamics of Intravenous Ciprofloxacin / Chapter 3.1 --- Introduction --- p.43 / Chapter 3.2 --- Materials and Methods --- p.45 / Chapter 3.3 --- Results --- p.49 / Chapter 3.4 --- Discussion --- p.51 / Chapter Chapter 4 --- The Effects of Oral Ferrous Sulfate on Pharmacokinetics and Pharmacodynamics of Oral Ciprofloxacin / Chapter 4.1 --- Introduction --- p.61 / Chapter 4.2 --- Materials and Methods --- p.62 / Chapter 4.3 --- Results --- p.66 / Chapter 4.4 --- Discussion --- p.68 / Chapter Chapter 5 --- Influence of Mineral Rich Traditional Chinese Medicines on the Pharmacokinetics of Oral Ciprofloxacin / Chapter 5.1 --- Introduction --- p.77 / Chapter 5.2 --- Materials and Methods --- p.81 / Chapter 5.3 --- Results --- p.85 / Chapter 5.4 --- Discussion --- p.86 / Chapter Chapter 6 --- General Conclusion --- p.98 / References --- p.105

Orientador: Anselmo Gomes de Oliveira / Banca: Maria Virginia Costa Scarpa / Banca: Maria Vitória Lopes Badra Bentley / Resumo: Neste trabalho estudou-se a encapsulação do cloridrato de ciprofloxacino (CIPRO) em lipossomas revestidos de ácido poli-láctico-co-glicólico (PLGA) ou o copolímero termosensível Pluronic® F127 (PLU). A eficiência de encapsulação foi obtida a partir das frações contendo lipossomas carregados de CIPRO separadas por cromatografia de exclusão em gel de Sephadex G-50, e mostrou-se melhor para as amostras contendo PLGA e maior concentração de CIPRO (5 mg/mL). A determinação do diâmetro médio dos lipossomas foi realizada por espalhamento dinâmico de luz (Light Scattering) e demonstrou redução no tamanho das estruturas quando PLGA ou PLU estavam presentes nas preparações. A incorporação de CIPRO aos lipossomas provocou aumento do tamanho das estruturas quando comparadas com as preparações isentas de fármaco. Verificou-se que o aumento na concentração de fármaco provocou a diminuição do diâmetro médio dos lipossomas. Experimentos de liberação in vitro mostraram que a liberação do CIPRO a partir dos lipossomas foi mais lenta em relação ao CIPRO não encapsulado. A liberação do CIPRO a partir de lipossomas revestidos com PLGA mostrou que a liberação foi mais lenta em relação aos lipossomas não revestidos. Os resultados demonstram que as preparações de CIPRO em lipossomas revestidos com PLGA ou PLU podem representar sistemas de liberação de fármacos antibióticos com grande potencial de utilização. O estudo de biodisponibilidade ocular demonstrou que lipossomas revestidos com PLGA e PLU mantiveram a MIC90 de CIPRO para os principais patógenos oculares por mais tempo que o CIPRO em solução no humor aquoso, quando administrados por via subconjuntival. Estes resultados demonstram que a associação de PLGA e PLU com lipossomas pode ser utilizada como um eficiente sistema de liberação ocular de fármacos. / Abstract: In the present work we studied the encapsulation of ciprofloxacin hydrochloride (CIPRO) in liposomes coated either by the poly-lactic-co-glycolic acid (PLGA) or thermosensitive copolymer Pluronic® F127 (PLU). Unilamellar liposomes containing 40 or 50 mM of hydrogenated soy phosphatidylcholine (FSH) were prepared by reverse phase evaporation (REV) method, followed by sonication. Encapsulation efficiency was obtained using fractions containing liposomes loaded with CIPRO, separated by exclusion chromatography on Sephadex G-50 gel. Best encapsulation efficiency was obtained with samples containing PLGA with CIPRO at a higher concentration (5 mg/mL). Liposome medium diameter was determined by dynamic light scattering, and showed a size reduction when either PLGA or PLU was present in the preparations. The incorporation of CIPRO into the liposomes caused a size increasement of the structures when compared to preparations lacking the drug. Nevertheless, increasing the drug concentration caused a decrease of the liposome medium diameter. Experiments of in vitro release showed that the liberation of CIPRO from the liposomes was slower when compared to not encapsulated CIPRO. The release of CIPRO from liposomes coated by PLGA showed that the liberation was slower when compared to non-coated liposomes. The results show that liposome preparations containing CIPRO and covered either by PLGA or PLU represent antibiotic drug delivery systems with great possibilities. The bioavailability study shows that liposomes covered by both PLGA and PLU maintained the MIC90 of CIPRO against the main ocular pathogens for longer time than CIPRO solution in aqueous humor, when subconjunctivally injected. These results demonstrate that the PLGA and PLU association with liposomes can be used as an efficient ocular drug delivery system. / Mestre

The emergence of antimicrobial resistance is a major global threat to modern medicine. The rapid dissemination of resistant pathogens and the associated loss of efficacy of many important drugs needs to be met with the development of new antibiotics and alternative treatment options. A better understanding of the evolution of resistance could help in developing strategies to slow down the spread of antimicrobial drug resistance. In this thesis we investigated the evolution of resistance to two important antibiotics, rifampicin and ciprofloxacin, paying special attention to the resistance patterns occurring with high frequency in clinical isolates. Rifampicin is a first-line drug in tuberculosis treatment and resistance to this valuable drug limits treatment options. Our work on rifampicin resistance helps to explain the extreme bias seen in the frequency of specific resistance mutations in resistant clinical isolates of M. tuberculosis. We identified an important interplay between the level of resistance, relative fitness and selection of fitness-compensatory mutations among the most common resistant isolates. Fluoroquinlones are widely used to treat infections with Gram-negatives and the frequency of resistance to these important drugs is increasing. Resistance to fluoroquinolones is the result of a multi-step evolutionary process. Our studies on the development of resistance to the fluoroquinolone drug ciprofloxacin provide insights into the evolutionary trajectories and reveal the order in which susceptible wild-type E. coli acquire multiple mutations leading to high level of resistance. We found that the evolution of ciprofloxacin resistance is strongly influenced by the mutation supply rate and by the relative fitness of competing strains at each successive step in the evolution. Our data show that different classes of resistance mutations arise in a particular, predictable order during drug selection. We also uncovered strong evidence for the existence of a novel class of mutations affecting transcription and translation, which contribute to the evolution of resistance to ciprofloxacin.

Made available in DSpace on 2014-06-11T19:24:11Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-08-04Bitstream added on 2014-06-13T19:48:18Z : No. of bitstreams: 1 oliveira_lc_me_arafcf.pdf: 1008043 bytes, checksum: 986417155d3a5f5fe5fe070125ff8109 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Neste trabalho estudou-se a encapsulação do cloridrato de ciprofloxacino (CIPRO) em lipossomas revestidos de ácido poli-láctico-co-glicólico (PLGA) ou o copolímero termosensível Pluronic® F127 (PLU). A eficiência de encapsulação foi obtida a partir das frações contendo lipossomas carregados de CIPRO separadas por cromatografia de exclusão em gel de Sephadex G-50, e mostrou-se melhor para as amostras contendo PLGA e maior concentração de CIPRO (5 mg/mL). A determinação do diâmetro médio dos lipossomas foi realizada por espalhamento dinâmico de luz (Light Scattering) e demonstrou redução no tamanho das estruturas quando PLGA ou PLU estavam presentes nas preparações. A incorporação de CIPRO aos lipossomas provocou aumento do tamanho das estruturas quando comparadas com as preparações isentas de fármaco. Verificou-se que o aumento na concentração de fármaco provocou a diminuição do diâmetro médio dos lipossomas. Experimentos de liberação in vitro mostraram que a liberação do CIPRO a partir dos lipossomas foi mais lenta em relação ao CIPRO não encapsulado. A liberação do CIPRO a partir de lipossomas revestidos com PLGA mostrou que a liberação foi mais lenta em relação aos lipossomas não revestidos. Os resultados demonstram que as preparações de CIPRO em lipossomas revestidos com PLGA ou PLU podem representar sistemas de liberação de fármacos antibióticos com grande potencial de utilização. O estudo de biodisponibilidade ocular demonstrou que lipossomas revestidos com PLGA e PLU mantiveram a MIC90 de CIPRO para os principais patógenos oculares por mais tempo que o CIPRO em solução no humor aquoso, quando administrados por via subconjuntival. Estes resultados demonstram que a associação de PLGA e PLU com lipossomas pode ser utilizada como um eficiente sistema de liberação ocular de fármacos. / In the present work we studied the encapsulation of ciprofloxacin hydrochloride (CIPRO) in liposomes coated either by the poly-lactic-co-glycolic acid (PLGA) or thermosensitive copolymer Pluronic® F127 (PLU). Unilamellar liposomes containing 40 or 50 mM of hydrogenated soy phosphatidylcholine (FSH) were prepared by reverse phase evaporation (REV) method, followed by sonication. Encapsulation efficiency was obtained using fractions containing liposomes loaded with CIPRO, separated by exclusion chromatography on Sephadex G-50 gel. Best encapsulation efficiency was obtained with samples containing PLGA with CIPRO at a higher concentration (5 mg/mL). Liposome medium diameter was determined by dynamic light scattering, and showed a size reduction when either PLGA or PLU was present in the preparations. The incorporation of CIPRO into the liposomes caused a size increasement of the structures when compared to preparations lacking the drug. Nevertheless, increasing the drug concentration caused a decrease of the liposome medium diameter. Experiments of in vitro release showed that the liberation of CIPRO from the liposomes was slower when compared to not encapsulated CIPRO. The release of CIPRO from liposomes coated by PLGA showed that the liberation was slower when compared to non-coated liposomes. The results show that liposome preparations containing CIPRO and covered either by PLGA or PLU represent antibiotic drug delivery systems with great possibilities. The bioavailability study shows that liposomes covered by both PLGA and PLU maintained the MIC90 of CIPRO against the main ocular pathogens for longer time than CIPRO solution in aqueous humor, when subconjunctivally injected. These results demonstrate that the PLGA and PLU association with liposomes can be used as an efficient ocular drug delivery system.

A 27-year-old Peruvian woman living with human immunodeficiency virus (HIV) in clinical stage B3 and not on antiretroviral therapy presented with a ten-day history of fever, chills, night sweats and a two-day history of skin lesions. On physical examination, several erythematous-purplish lesions were found on the face and legs. Meningococcal infection was suspected and ceftriaxone was started. Blood culture grew nontyphoidal Salmonella enterica. A biopsy of the skin lesions showed leukocytoclastic vasculitis (LCV); therefore, corticosteroids were added. After two weeks of antibiotic and corticosteroid treatment, the lesions had resolved, but they recurred two days after treatment with prednisone was stopped. Corticosteroids and combination antiretroviral therapy were started simultaneously and the lesions resolved without recurrence. HIV infection has been associated with higher rates of skin lesions in salmonellosis. LCV has been described both in the setting of HIV infection and salmonellosis. However, our review of the literature found no previous cases of LCV in concurrent HIV and salmonellosis. / Revisión por pares / Revisión por pares

京都大学 / 0048 / 新制・論文博士 / 博士(医学) / 乙第12918号 / 論医博第2093号 / 新制||医||1009(附属図書館) / 32128 / (主査)教授 羽賀 博典, 教授 中川 一路, 教授 上杉 志成 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

The kinetics and mechanism of accumulation for liposomal ciprofloxacin (Lipo-CPFX) into the rat alveolar macrophage NR8383 cells were studied in vitro, in comparison to unformulated ciprofloxacin (CPFX). Upon incubation with CPFX or Lipo-CPFX, cellular drug accumulation was determined from the cell lysates or efflux was from the extracellular media by fluorescence-HPLC. The accumulation for Lipo-CPFX reached the asymptotic values at ≥ 2 hours, which was a result of uptake and efflux. The uptake appeared to be due to liposomes, mediated via cellular energy-independent mechanism like lipid fusion. In contrast, the efflux appeared to be due to ciprofloxacin, partly cellular energy-dependent, and involve probenecid-sensitive multidrug resistance proteins (MRPs). Overall, Lipo-CPFX enabled greater drug accumulation into the NR8383 cells than CPFX. This logically suggests a greater potential to treat respiratory infections especially caused by bacteria resistant to phagocytic killing.

No description available.

MSc (Med), Clinical Microbiology and Infectious Diseases, Faculty of Health Sciences, University of the Witwaterstrand / Emergence and spread of resistance to ciprofloxacin among Neisseria gonorrhoeae strains has reduced the options of effective treatment for gonococcal infections and has become a concern worldwide. Up until 2008, ciprofloxacin was recommended first-line therapy for treatment of presumptive N. gonorrhoeae infections in South Africa. At the time this MSc project was conceived, ciprofloxacin was still used as first-line therapy for presumptive gonococcal infections. A real-time polymerase chain reaction (PCR) assay was used to detect ciprofloxacin-resistant N. gonorrhoeae in DNA extracted from non-invasive urine samples collected as part of the national microbiological surveillance (NMS) programme during 2006-2007. The molecular epidemiology of ciprofloxacinresistant Neisseria gonorrhoeae was investigated by sequencing the quinolone resistance determining regions (QRDR) of the gyrA and parC genes of N. gonorrhoeae and performing N. gonorrhoeae multi-antigen sequence typing (NGMAST). As part of the NMS program for sexually transmitted infections (STIs) urine and urethral swabs were collected from men presenting with urethral discharge at primary health care clinics in Johannesburg (Gauteng), Cape Town (Western Cape) and Kimberley (Northern Cape). Urine samples and cultured N. gonorrhoeae isolates from 2006-2007 were stored at -700C and available for this study. Gonococci, previously isolated from urethral swabs, were subcultured directly onto New York City media. Isolate identity was re-confirmed by typical colony morphology and biochemical tests. Urine samples from Johannesburg were tested in order to develop the real-time PCR protocol. Subsequently, paired urethral swab DNA and N. gonorrhoeae cultures were tested from NMS patients recruited in Kimberley and Cape Town. Where possible, the PCR assay results were compared with paired antibiotic susceptibility data for ciprofloxacin. Quinolone resistance determining regions (QRDR) for gyrA and parC were screened for known point mutations associated with resistance to ciprofloxacin. Detection of mutations by the real-time PCR assay generally agreed with the phenotype of either decreased susceptibility or resistance to ciprofloxacin. All ciprofloxacin resistant gonococcal isolates had the same gyrA and parC mutations, which initially suggested that quinolone resistant N. gonorrhoeae (QRNG) in Kimberley, Cape Town and Johannesburg, may be attributed to the spread of a single clone. The use of a more discriminatory typing scheme, Neisseria gonorrhoeae Multi-Antigen Sequence Typing (NG-MAST) genotyping, revealed that ciprofloxacin resistant gonococcal isolates in Johannesburg and Cape Town were heterogeneous, with sequence type (ST) 217 being most prevalent in both cities (5/16, Johannesburg; 7/11, Cape Town). In contrast, all eight QRNG isolates from Kimberley were typed as ST 533. The use of molecular methods allowed ciprofloxacin antimicrobial susceptibility determination by PCR in non-invasive specimens. This is useful in situations where bacterial cultures are unavailable or die before antimicrobial susceptibility testing can be performed. Molecular assays to detect ciprofloxacin resistance may guide physicians as to the most ideal antimicrobial combinations for individual patient treatment. As a result of emerging widespread resistance gonococci to ciprofloxacin, in 2008, the Department of Health recommended that ciprofloxacin be removed as a first line therapy in the South African national sexually transmitted infections treatment guidelines for treatment of urethritis, cervicitis and their complications. Although ciprofloxacin is no longer used as a first-line therapy to treat gonorrhoea within our country, it may still be used in cases of severe penicillin allergy or as part of multi-drug therapy for gonococcal infections in the future. The ability to detect ciprofloxacin resistance by real-time PCR will be a useful technique in such situations.

Diante da presença de microrganismos resistentes à terapêutica endodôntica, novas formulações e alternativas medicamentosas têm sido investigadas. Desse modo, esse estudo teve como objetivo avaliar a citotoxicidade de diferentes medicações indicadas como medicação intracanal. Culturas de células de fibroblastos foram estimuladas de acordo com os seguintes grupos experimentais: Grupo I: controle; Grupo II: cloridrato de ciprofloxacina; Grupo III: cloridrato de clindamicina e Grupo IV: metronidazol. Para tal, as concentrações utilizadas para cada antibiótico foram de 5, 50, 150 e 300 mg/L, nos tempos experimentais de 24, 48, 72 e 96 horas. A citotoxicidade dessas substâncias foi avaliada usando a técnica de análise do MTT e leitura em espectrofotômetro de ELISA. Os resultados obtidos foram analisados pelo software BioEstat 4.0, por meio do teste de Kruskal-Wallis, complementado pelo teste de Dunn, com nível de significância de 5%. A viabilidade celular foi analisada diante da relação entre as diferentes concentrações e uma mesma concentração nos diferentes tempos experimentais. Diante dos resultados obtidos, foi possível concluir que: todos os antibióticos testados apresentaram citotoxicidade dosedependente. As concentrações de 5 e 50 mg/L, de todos os antibióticos, permitiram a viabilidade dos fibroblastos em todos os tempos avaliados. Independente da medicação avaliada, a viabilidade celular em 24 horas foi mais elevada em comparação aos demais tempos experimentais. A comparação entre os antibióticos nas mesmas concentrações, em diferentes tempos experimentais demonstrou que o gel de metronidazol apresentou menor citotoxicidade em 72 e 96 horas no confronto com os dois outros antibióticos, enquanto que o cloridrato de clindamicina apresentou maior viabilidade celular em relação ao cloridrato de ciprofloxacina em 72 horas. / New medicaments have been assessed due to the presence of resistantmicroorganism to therapeutic procedures. Thus, this study aimed to evaluate the citotoxicity of differents drugs indicated as intracanal medicament. The human gingival fibroblasts were estimulated according to the following experimental groups: Group I: control; Group II: Ciprofloxacin Hydrochloride; Group III: Clindamicin Hydrochloride and Group IV: Metronidazole. The concentration used for each drugs were 5, 50, 150, and 300 mg/L, in the experimental time of 24, 48, 72, and 96 hours. The citotoxicicty were evaluated with MTT analysis and ELISA spectrophotometer reading. The results were evaluated by BioEstat 4.0 software, according to Kruskal- Wallis test supplemented by Dunn test with significance level of 5%. Cell viability analysis has been assessed in different concentrations in each experimental period. According to the results, it is possible to conclude that: all the tested drugs showed dose-dependent citotoxicity. The concentrations of 5 and 50mg/L of all groups allowed fibroblast viability in all evaluated periods. The cell viability in 24 hours was higher than the others experimental periods. The comparing between the antibiotics at the same concentrations, in different times showed the gel metronidazole obtained lowest citotoxicity at 72 and 96 hours when compared with the others antibiotics, meanwhile, the Clindamicin Hydrochloride showed the highest cell viability compared to Ciprofloxacin Hydrochloride, at 72 hours.