Inkorporace antibiotik do biodegradabilních nanovlákenných vrstev pro nové medicínské aplikace
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V této diplomové práci byl vyvinut a otestován nanovlákenný materiál s inkorporovaným antibiotikem, který by mohl být slibným řešením nebezpečných pooperačních komplikací v gastrointestinální chirurgii spojených s výskytem bakterií. Na bezjehlovém elektrostatickém zařízení NanospiderTM byly zvlákněny polymerní roztoky poly-e-kaprolaktonu (PCL) s přídavkem antibiotika Gentamicin sulfát (GS) o koncentraci 0; 1; 3; 5; 7,5 a 10 hm. %. Vyrobené vrstvy byly charakterizovány pomocí skenovací elektronové mikroskopie a byla měřena jejich smáčivost. Obsah antibiotika ve vrstvách byl testován pomocí elementární analýzy, infračervené spektroskopie (FTIR), UV-spektrofotometrie a vysoce-účinné kapalinové chromatografie (HPLC). Byla sledována absorpce destilované vody a simulované střevní tekutiny vyrobených materiálu, hodnotila se cytotoxicita extraktů materiálů, a i samotného antibiotika GS v médium, na buněčné linii 3T3 myší fibroblasty. Vrstvy byly vyhodnoceny jako cytokompatibilní. Materiály byly také testovány in vitro, kdy po 2, 7, 14 a 21 dnech byla hodnocena adheze a proliferace 3T3 buněk pomocí metabolického testu CCK-8 a fluorescenční a skenovací elektronové mikroskopie. Byly provedeny antibakteriální testy, které potvrdily přítomnost, uvolňování a účinnost antibiotika GS z nanovlákenných vrstev. Během celého experimentu byl také sledován vliv sterilizace ethylenoxidem na nanovlákenné vrstvy s inkorporovaným antibiotikem. Sterilizace nezpůsobila žádné významné změny nanovlákenných materiálů s inkorporovaným antibiotikem a sterilní vrstvy měly antibakteriální účinky.
In this diploma thesis, a nanofibrous material with an incorporated antibiotic was developed and tested. The introduced strucuture could serve as promising prevention of the life-threathning postoperative in gastrointestinal surgery associated with the occurrence of bacteria. Polymer solutions of poly-e-caprolactone (PCL) with antibiotic Gentamicin sulfate (GS), in concentrations 0; 1; 3; 5; 7,5 and 10 wt. % were electrospun via needleless NanospiderTM device. The produced layers were characterized by scanning electron microscopy and their wettability was measured. The amount of antibiotic in the layers was tested by elementar analysis, infrared spectroscopy (FTIR), UV-spectrophotometry and high-performance liquid chromatography (HPLC). The absorption of water and simulated intestinal liquid of the materials was monitored, the cytotoxicity of the material extracts was evaluated, as well as the cytotoxicity of the GS antibiotic in the medium, on the 3T3 mouse fibroblast cell line. The layers were evaluated as cytocompatible. The materials were also tested in vitro, where after 2, 7, 14 and 21 days, the cell adhesion and proliferation was assessed using the CCK-8 metabolic assay and fluorescence and scanning electron microscopy. Antibacterial tests were performed to confirm the presence release and efficiency of the antibiotic GS from the nanofiber layers. The effect of ethylene oxide sterilization on nanofibrous layers with incorporated antibiotic was also studied throughout the experiment part. Sterilization did not cause any significant changes in the nanofibrous materials with an incorporated antibiotic, and the sterile layers had antibacterial properties.
In this diploma thesis, a nanofibrous material with an incorporated antibiotic was developed and tested. The introduced strucuture could serve as promising prevention of the life-threathning postoperative in gastrointestinal surgery associated with the occurrence of bacteria. Polymer solutions of poly-e-caprolactone (PCL) with antibiotic Gentamicin sulfate (GS), in concentrations 0; 1; 3; 5; 7,5 and 10 wt. % were electrospun via needleless NanospiderTM device. The produced layers were characterized by scanning electron microscopy and their wettability was measured. The amount of antibiotic in the layers was tested by elementar analysis, infrared spectroscopy (FTIR), UV-spectrophotometry and high-performance liquid chromatography (HPLC). The absorption of water and simulated intestinal liquid of the materials was monitored, the cytotoxicity of the material extracts was evaluated, as well as the cytotoxicity of the GS antibiotic in the medium, on the 3T3 mouse fibroblast cell line. The layers were evaluated as cytocompatible. The materials were also tested in vitro, where after 2, 7, 14 and 21 days, the cell adhesion and proliferation was assessed using the CCK-8 metabolic assay and fluorescence and scanning electron microscopy. Antibacterial tests were performed to confirm the presence release and efficiency of the antibiotic GS from the nanofiber layers. The effect of ethylene oxide sterilization on nanofibrous layers with incorporated antibiotic was also studied throughout the experiment part. Sterilization did not cause any significant changes in the nanofibrous materials with an incorporated antibiotic, and the sterile layers had antibacterial properties.
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Nanovlákenné vrstvy, poly-e-kaprolakton, elektrostatické zvlákňování, inkorporace, antibiotikum, Gentamicin sulfát, antibakteriální, Nanofibrous layers, poly-e-caprolacton, electrospinning, incorporation, antibiotics, Gentamicin sulfate, antibacterial