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Koža predstavlja prvi stik z okolico, zato je tudi nenehno izpostavljena tveganju za nastanek poškodb in ran. Kadar proces celjenja traja več kot 8 tednov, govorimo o kroničnih ranah. Nanovlakna s svojo mrežasto strukturo posnemajo strukturne elemente in lastnosti zunajceličnega ogrodja, zato predstavljajo velik potencial kot sodobne obloge za celjenje ran. Le-te morajo biti sterilne oz. zadoščati kriterijem za ustrezno mikrobiološko kakovost farmacevtskih izdelkov, kar predstavlja velik izziv pri izdelavi le-teh. V okviru magistrske naloge smo vrednotili mikrobiološko kakovost nanovlaken, izdelanih z metodo elektrostatskega sukanja. Nanovlakna smo pripravili iz polivinilalkohola in polietilenoksida ter z vrstično elektronsko mikroskopijo preučevali njihovo morfologijo. Z aseptičnim delom in metodami sterilizacije (s suho toploto, avtoklaviranjem in z ultravijoličnim sevanjem) smo optimizirali celoten proces priprave nanovlaken s ciljem razviti sterilen končni izdelek. Mikrobiološko kakovost smo preverjali s testi po Evropski farmakopeji 9.0. Uspešno smo izdelali polimerna nanovlakna iz obeh polimerov s povprečnim premerom okoli 200 nm. Ugotovili smo, da avtoklaviranje polimernih raztopin bistveno vpliva na povprečni premer izdelanih nanovlaken, medtem ko končna sterilizacija s suho toploto ni vplivala na morfologijo polivinilalkoholnih nanovlaken. Test sterilnosti smo izvedli za polivinilalkoholna nanovlakna, sterilizirana s suho toploto, in dokazali njihovo sterilnost. Izvedli smo tudi numerični test za nesterilne izdelke. Na petrijevkah z vzorci polietilenoksidnih nanovlaken iz sterilizirane raztopine polietilenoksida nismo opazili rasti nobene kolonije, medtem ko smo pri polivinilalkoholnih nanovlaknih iz sterilizirane polivinilalkoholne raztopine prešteli 1 (50 CFU/g izdelka) oz. 2 koloniji (100 CFU/g). Tako pripravljena nanovlakna so ustrezne mikrobiološke kakovosti za dermalno uporabo (kriterij 102 CFU/g izdelka). V raziskavi smo uspeli izdelati sterilna nanovlakna po dveh različnih postopkih, in sicer s končno sterilizacijo izdelanih polivinilalkoholnih nanovlaken (suha toplota) in z aseptično pripravo, kjer je bil polimer izbora polietilenoksid. Takšna nanovlakna imajo tako z vidika zgradbe in morfologije kot ustrezne mikrobiološke kakovosti velik potencial za uporabo kot sodobne obloge za celjenje ran. The skin is the first contact with the outer world, thus it is constantly exposed to the risk of injuries and wounds. The unsuccessful wound healing, lasting for more than 8 weeks, is characteristic of chronic wounds. The structure of nanofibers mimics the morphological properties of extracellular matrix, which plays a crucial role in wound healing process. Therefore, nanofibers represent modern dressing for wound healing application. The dressings should be sterile and thus meet the criteria for adequate microbiological quality of pharmaceutical products, which represents a big challenge during their preparation. The aim of the master thesis was to evaluate microbiological quality of nanofibers produced by electrospinning. Nanofibers were produced from polyvinyl alcohol and polyethylene oxide and characterized by scanning electron microscopy. We have optimizied our working protocol using different sterilization methods (dry heat, steam and ultraviolet sterilization) and aseptic procedure with the aim to produce sterile nanofibers. Microbiological quality of nanofibers was evaluated according to the tests specified in European Pharmacopoeia 9.0. Polyvinyl alcohol and polyethylene oxide nanofibers, both with avarage diameter of approximately 200 nm, were successfully developed. The steam sterilization of polymer solutions significantly influenced the average diameter of nanofibers from polyvinyl alcohol or polyethylene oxide, whereas the high temperatures during dry heat sterilization did not affect the morphology of polyvinyl alcohol nanofibers. The sterility test performed on polyvinyl alcohol nanofibers sterilized by dry heat confirmed their sterility. We have also performed the numeric test for nonsterile products. Petri dishes with polyethylene oxide nanofibers prepared from sterilized polymer solution did not show growth of any colonies after the incubation period, while samples with polyvinyl alcohol nanofibers prepared from sterilized polymer solution showed growth of 1 (50 CFU/g of the product) and 2 colonies (100 CFU/g of the product), thus the nanofibers meet the criteria of microbiological quality of nonsterile pharmaceutical products for dermal use (the upper limit being 100 CFU/g of the product). We have managed to produce sterile nanofibers using two different approaches. Polyvinyl alcohol nanofibers were prepared and then sterilized by dry heat, while sterile polyethylene oxide nanofibers were produced by the aseptic procedure. These nanofibers represent an immense potential as modern dressings for wound healing application, due to their composition, morphology and also adequate microbiological quality. |
