Development of new antimicrobial bioactive composites for medical application in wound treatment

Danas, razvoj novih kompozitnih biomaterijala za medicinsku primenu u terapiji rana bazira se na poznavanju različitih faktora povezanih sa procesom zarastanja i podrazumeva rešavanje specifičnih problema koji prate hronične rane. Cilj ove doktorske disertacije bio je razvoj novih kompozitnih hidrogelova na bazi alginata i čestica aktivnog uglja (AU) koji će u potencijalnoj primeni u tretmanu rana ispoljiti više povoljnih efekata. Naime, cilj je bio razvoj kompozita iz koga se, u dodiru sa biološkim fluidima, čestice AU kontrolisano i efikasno otpuštaju tokom dužeg perioda. Pri tome, čestice AU mogu biti nosači različitih aktivnih agenasa, dok bi alginatni hidrogel efikasno upijao vlagu. Kao model aktivne supstance izabrani su povidon-jod (PVP-I) i lignin model jedinjenja (LMJ) kao agensi koji ispoljavaju antimikrobno dejstvo. Rezultati istraživanje su pokazali da su metodom ekstruzije uspešno dobijene kompozitne Ca-alginatne čestice (CaA/AU) i to iz smeša alginata koncentracija u opsegu od 0,5 - 2 mas. % i AU u opsegu od 1-20 mas. %. Ispitivanjem kinetike otpuštanja AU iz suvih CaA/AU čestica u fiziološkom rastvoru utvrđeno je da je sastav optimalne smeše 0,5 mas. % alginata i 20 mas. % AU. Kinetika otpuštanja AU je uspešno opisana modelom unutrašnje difuzije osim u slučaju čestica dobijenih iz smeše optimalnog sastava gde je otpuštanje uzrokovano i erozijom čestica što je opisano Korsmejer-Pepasovim modelom Ekstruzijom smeše optimalnog sastava u rastvor za geliranje koji je sadržao jone cinka uspešno su dobjene i Zn0,5/V kompozitne čestice. Čestice na bazi Ca- i Zn-alginata dobijene iz smeše optimalnog sastava (Ca- i Zn0,5/V čestice) su u ii fiziološkom rastvoru otpustile oko 50 - 60 % ukupnog sadržaja AU tokom perioda od 5 dana koji je relevantan za tretman hroničnih rana. Ove čestice su zatim uspešno impregnirane prvom model supstancom (PVP-I), a u ispitivanjima desorpcije utvrđeno je da je PVP-I čvrsto vezan za čestice AU i da je stepen desorpcije u okolni alginatni matriks, kao i u fiziološki rastvor zanemarljiv. Na čestice AU je takođe uspešno adsorbovana i druga model supstanca (LMJ) i pokazano je da ne dolazi ni u ovom slučaju ne dolazi do desorpcije u fiziološkom rastvoru. Kompozitne Ca- i Zn-alginatne čestice impregnirane sa PVP-I su pokazale izrazitu antimikrobnu aktivnost u suspenzijama svih ispitivanih, standardnih i kliničkih sojeva mikroorganizama. Antimikrobni efekat adsorbovanog joda se može pripisati direktnom kontaktu sa ćelijskom membranom mikroorganizama što je u slučaju ispitivanih Gram pozitivnih i Gram negativnih bakterija bilo dovoljno za baktericidni efekat, za razliku od gljivice C. albicans koja se pokazala najotpornijom. Sa druge strane čestice AU sa adsorbovanim LMJ nisu pokazale antimikrobne efekte u ispitivanim suspenzijama što zahteva dalja istraživanja mehanizma adsorpcije ovih jedinjenja, kao i mehanizma njihovog antimikrobnog dejstva. Kompozitni Ca- i Zn-alginatni hidrogelovi sa sadržanim česticama AU na kojima je adsorbovan PVP-I su pokazali potencijal za primenu u oblogama za rane zbog efikasnosti, dostupnosti i širokog spektra dejstva joda, kao i potencijalnog sinergističkog dejstva sa otpuštenim katjonima koji imaju pozitivan efekat na zarastanje oštećenog tkiva.Today, the development of new composite biomaterials for wound treatment application is based on knowledge of different factors which affects the process of wound healing in order to identify solutions for specific problems related to chronic wounds. The aim of this thesis was to develop new composite hydrogels based on alginate and activated charcoal particles (AC) which would show beneficial effects in potential wound treatment application. Specifically, the aim was to develop composites which would during contact with biological fluids, release AC particles in a controlled and efficient manner during a prolonged period of time. Simultaneously, AC particles could be carriers of different active agents while alginate hydrogel would efficiently absorb the fluids. Povidone-iodine (PVP-I) and lignin model compounds (LMC) were chosen as model substances of active agents with antimicrobial activity. The results showed that the composite Ca-alginate beads (CaA/AC) were successfully obtained by extrusion method using mixtures of alginate in concentration range of 0.5-2 % w/w and AC in a concentration range of 1-20 % w/w. The diffusion model was successfully applied on the release kinetics experimental data of all groups of A/AC beads, except in the case of the beads obtained from the mixture with the optimal composition. In this case, the release of AC particles was a result of diffusion as well as erosion and the Korsmeyer-Peppas model was successfully applied. Zn0.5/H composite beads were successfully obtained by extrusion of optimal mixture in the gelling bath that contained Zn ions. The beads based on Ca- and Znalginate obtained from the optimal mixture (Ca- and Zn0.5/H beads) released around 50-60 % of iv the total AC content during a 5 day period, which is a period relevant for chronic wound treatment. These beads were successfully impregnated with the first model substance (PVP-I). The desorption studies established that PVP-I was firmly adsorbed onto AC particles and that the level of desorption into alginate matrix as well as saline solution was below the detection level. The second model substance (LMC) was successfully absorbed and the level of desorption in saline solution was below the detection level as well. Both composite Ca- and Zn-alginate beads impregnated with PVP-I showed high antimicrobial activity in the suspensions of all examined ATCC and clinical strains as well. The antimicrobial effect of adsorbed iodine can be explained by direct contact with the cell membrane of microorganisms which was enough to establish bactericidal effect in case of Gram positive and Gram negative bacteria while the yeast Candida albicans was the most resistant. On the contrary, AC particles with adsorbed LMC did not exhibit antimicrobial effect in any of the examined suspensions which demands a further investigation of adsorption mechanism and antimicrobial activity of LMC as well. Composite Ca- and Zn-alginate hydrogels with incorporated AC particles with adsorbed PVP-I showed potential for wound dressing application because of its high efficiency, availability and broth antimicrobial spectrum of iodine as well as its potential synergistic effect with released cations that exhibit wound healing effect