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Porast rezistencije bakterija na antibiotike koji su trenutno na tržištu potaknuo je potrebu za otkrivanjem novih. Makrolidni antibiotik azitromicin koristi se za liječenje bolesti uzrokovanih Gram-pozitivnim i Gram-negativnim bakterijama. Makrozoni su konjugati azitromicina i tiosemikarbazona pripremljeni za liječenje infekcija uzrokovanih rezistentnim bakterijama, s potencijalom zaobilaženja razvijenih mehanizama rezistencije. Kako bi se dodatno povećala biološka aktivnost i farmakološka svojstva, napravljeni su kompleksi tiosemikarbazona s metalnim ionima. U ovom radu prvo je pripravljen tiosemikarbazon izveden iz 4- aminobenzojeve kiseline i salicilaldehida. Nakon toga, dobiveni tiosemikarbazon je vezan na 4''-aminopropilni derivat azitromicina dajući pripadajući makrozon. Istražena je mogućnost kompleksiranja s ionima bakra(II) i nikla(II). Sintetizirani spojevi karakterizirani su spektroskopijom NMR. The increase in bacterial resistance to currently marketed antibiotics has prompted the need to discover new ones. The macrolide antibiotic azithromycin is used to treat diseases caused by Gram-positive and Gram-negative bacteria. Macrozones are conjugates of azithromycin and thiosemicarbazones prepared to treat infections caused by resistant bacteria, with potential of bypassing the developed resistance mechanisms. In order to further increase biological activity and pharmacological properties thiosemicarbazone complexes with metal ions have been made. In this work, a thiosemicarbazone derived from 4-aminobenzoic acid and salicylaldehyde was prepared first. Subsequently, the resulting thiosemicarbazone was bound to the 4''-aminopropyl derivative of azithromycin. The possibility of complexation with copper (II) and nickel (II) ions was explored. The synthesized compounds were characterized by NMR spectroscopy.

Značajan porast bakterijske rezistencije utječe na sve veću potrebu za istraživanjem i razvojem novih antibiotika. U sklopu ovog rada pripravljene su tri nove klase makrozona, konjugata makrolidnog antibiotika azitromicina i tiosemikarbazona. Tiosemikarbazoni su korišteni za kemijsku modifikaciju azitromicina na tri različita položaja. Cilj rada je priprava novih derivata makrolida sa širim spektrom biološke aktivnosti. Pripravljenim spojevima određena je biološka aktivnost in vitro na susceptibilne i rezistentne Gram-pozitivne i Gram-negativne bakterije te su istražene interakcije s biološkim receptorima. Za proučavanje interakcija primijenjena je spektroskopija NMR, cirkularni dikroizam, fluorescencijska spektroskopija i molekulsko modeliranje. Makrozoni su pokazali zadovoljavajuću biološku aktivnost, posebice na neke rezistentne bakterijske sojeve na koje azitromicin nije aktivan. Rezultati dobiveni spektroskopskim metodama i molekulskim modeliranjem ukazuju na odnos strukture i aktivnosti novih derivata azitromicina, što može poslužiti za dizajn i otkriće biaktivnijih derivata makrolida. Owing to a significant increase in bacterial resistance, a high need for discovery and development of new antibiotics has emerged. As part of this work, three novel classes of macrozones, conjugates of macrolide antibiotic azithromycin and thiosemicarbazones, were prepared. Thiosemicarbazones were used to chemically modify azithromycin at three different positions. The aim of this thesis was to prepare compounds with a wider range of biological activity including resistant bacterial strains. In vitro biological activity of the prepared compounds was determined against susceptible and resistant Gram-positive and Gram-negative bacteria, and interactions with biological receptors were investigated. Macrozones showed satisfactory biological activity, especially against some resistant bacterial strains to which azithromycin is inactive. NMR spectroscopy, circular dichroism, fluorescence spectroscopy, and molecular modeling were further used to study interactions of selected compounds with biological targets. The results obtained by spectroscopic methods and molecular modeling indicate a relationship between structure and activity of new azithromycin derivatives, which can be used for design and discovery of more active macrolide derivatives.

Pojava sve veće otpornosti bakterija na postojeće antibiotike predstavlja ozbiljan zdravstveni problem pa se stoga u području farmaceutske i medicinske kemije ulažu veliki napori u razvoj novih antibiotika. Makrozoni predstavljaju nove konjugate azitromicina i derivata tiosemikarbazida koji pokazuju izvrsnu antibakterijsku aktivnost in vitro na susceptibilne i rezistentne bakterijske sojeve. Proces razvoja novih antibiotika sastoji se od mnogo koraka, pri čemu je izuzetno važna identifikacija farmaceutskih onečišćenja i novosintetiziranih spojeva – potencijalnih antibiotika. U tu svrhu vrlo često se koriste vezani analitički sustavi pomoću kojih se postiže učinkovitije i brže pročišćavanje te identifikacija spojeva u odnosu na klasične tehnike off-line. U ovom diplomskom radu provedena je izolacija i strukturna karakterizacija novosintetiziranih 3-trifluormetoksifenil- i 3-(3-fenilpropil)-makrozona te njihovih dijastereomera, kao i nekih onečišćenja iz reakcijskih smjesa pomoću sustava LC-SPE/krioNMR. Za strukturnu karakterizaciju izoliranih komponenata korištene su jedno- i dvodimenzijske tehnike NMR te spektrometrija masa. Na temelju analize prikupljenih podataka predložene su strukture izoliranih spojeva. Increasing bacterial resistance to existing antibiotics is a serious health problem, and great efforts are being made to discover and develop new antibiotics to overcome this problem. Macrozones are novel conjugates of azithromycin and thiosemicarbazides that show very good in vitro activity against susceptible and resistant bacterial strains. Development of new antibiotics is a multistep process and one of the most important steps is the identification of pharmaceutical impurities and newly synthesized compounds – potential antibiotics. For this purpose, hyphenated systems are very oftenly used to achieve more efficient and faster purification and identification of compounds compared to off-line techniques. In this study, isolation and structural characterization of newly synthesized 3-trifluoromethoxyphenyl- and 3-(3-phenylpropyl)-macrozones and their diastereoisomers, as well as some impurities from reaction mixtures, have been performed using hyphenated system LC-SPE/cryo-NMR. Oneand two-dimensional NMR techniques and mass spectrometry were used for structural characterization of isolated components. Based on the analysis of collected data, the structures of isolated compounds were proposed.

Makrolidni antibiotici, kao što je azitromicin, imaju široku primjenu u liječenju infekcija dišnih puteva uzrokovanih Gram-pozitivnim i Gram-negativnim bakterijama. Tiosemikarbazoni, kao i neki njihovi metalni kompleksi, također pokazuju antibiotsku, protuupalnu i antitumorsku aktivnost. Racionalnim pristupom u spajanju ovih dviju klasa spojeva moguće je dobiti novu klasu makrolidnih konjugata „makrozona“, s poboljšanom biološkom aktivnošću, ujedno aktivnim i na rezistentne bakterijske sojeve. Stoga su sintetizirani novi biološki aktivni konjugati makrolidnih antibiotika dobivenih vezanjem različitih tiosemikarbazonskih derivata na 9a-N i 4''-O γ-aminopropil-azitromicin uz izolaciju putem preparativnog LC-MS-a i njihovu karakterizaciju pomoću 1D i 2D NMR-a. Pripravljenim spojevima ispitano je antimikrobno djelovanje na niz osjetljivih i rezistentnih sojeva Gram-pozitivnih (Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis, Streptococcus pyogenes) i Gram-negativnih bakterija (Escherichia coli, Moraxella catarrhalis) ključnim za područje djelovanja makrolidnih antibiotika. Macrolide antibiotics, such as azithromycin, have wide application in the treatment of respiratory infections caused by Gram-positive and Gram-negative bacteria. Thiosemicarbazones, as well as some of their metal complexes, also show antibiotic, antiinflammatory and antitumor activity. With a rational approach in combining these two classes of compounds, it is possible to obtain a new class of macrolide conjugates "macrozones", with improved biological activity, while also active in resistant bacterial strains. Thus, new biologically active conjugates of macrolide antibiotics obtained by binding various thiosemicarbazone derivatives to 9a-N and 4''-O γ-aminopropyl-azithromycin were synthesized with isolation by preparative LC-MS and their characterization by 1D and 2D NMR. The prepared compounds were tested for antimicrobial activity on a number of sensitive and resistant strains of Gram-positive (Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis, Streptococcus pyogenes) and Gram-negative bacteria (Escherichia coli and Moraxella catarrhalis) which are crucial for the area of application of macrolide antibiotics

Makrozoni su novi konjugati azitromicina i derivata tiosemikarbazona koji pokazuju izvrsno antibakterijsko djelovanje in vitro na odabrane Gram-pozitivne i Gram-negativne bakterije. Zbog sve većeg razvoja bakterijske rezistencije na dosadašnje antibiotike cilj mnogih istraživanja u medicinskoj i farmaceutskoj kemiji je razvoj novih spojeva s poboljšanom biološkom aktivnošću. Nikal je element u tragovima bitan za bakterije, biljke životinje i ljude i kao kofaktor ima važnu ulogu u mnogim enzimima. U nizu slučajeva su kompleksi nikla s tiosemikarbazonima pokazali veću biološku aktivnost od nekompleksiranih liganada. U ovom je radu uspješno pripravljen kompleks nikla(II) s makrozonskim 4'' derivatom azitromicina, koji je dobiven konjugacijom 4''-aminopropilnog derivata azitromicina i kompleksa nikla(II) s tiosemikarbazonom. Pripravljeni spoj okarakteriziran je spektroskopijom NMR. Također je određena biološka aktivnost spoja in vitro na susceptibilne i rezistentne sojeve bakterija. Macrozones are new conjugates of azithromycin and thiosemicarbazones that possess excelent in vitro antibacterial activity against selected Gram-positive and Gram-negative bacteria. Due to the increasing number of macrolide-resistant bacteria, the aim of many studies in medicinal and pharmaceutical chemistry is development of new compounds with improved biological activity. Nickel is a trace element essential for bacteria, plants, animals and humans. In multiple cases, nickel complexes with thiosemicarbazones showed increased biological activity than uncomplexed ones. In this study succesful synthesis and structural characterisation of nickel(II) coordination compounds with macrozone 4'' derivative of azithromycin have been performed using NMR spectroscopy. In vitro biological activity of the compound against susceptible and resistant bacterial strains was also determined and discussed.

CILJ ISTRAŽIVANJA: Glavni cilj ovog istraživanja bio je procijeniti stavove i znanja liječnika obiteljske medicine Splitsko-dalmatinske županije o propisivanju antibiotske terapije COVID-19 oboljelim pacijentima, te upoznatost sa najrecentnijim smjernicama za liječenje koronavirusne bolesti i nadomjesnom terapijom vitaminom D. ISPITANICI I METODE: Provedeno je istraživanje u svrhu ispitivanja stavova i znanja liječnika obiteljske medicine Splitsko-dalmatinske županije o upotrebi antibiotske terapije kod COVID-19 pozitivnih osoba. Podaci su prikupljeni pomoću anketnog upitnika kojem se pristupalo putem Google forms internetske aplikacije tijekom svibnja 2022. Upitnik se sastojao od 23 pitanja. Za analizu podataka korišten je statistički software STATISTICA 12, Tibco, Kalifornija. REZULTATI: Sa zadnjim smjernicama za liječenje oboljelih od koronavirusne bolesti (verzija 5 od 8.veljače 2022. godine) je upoznato 100 (87%) liječnika obiteljske medicine. Od 115 liječnika, njih 88 (77,2%) se u potpunosti slaže sa preporukom da je osnova liječenja COVID-19 infekcije ublažavanje simptoma, sprječavanje komplikacija i progresije bolesti te simptomatsko i suportivno liječenje. Najveći broj LOM smatra da su propisali antibiotsku terapiju za 10-30% oboljelih (41,67%). Na pitanje o izboru i trajanju antibiotske terapije kod COVID-19 pozitivnih pacijenata 80 (70,80%) LOM bi se odlučilo na makrolide (azitromicin) u trajanju od 3 dana, njih 20 (17,70%) na makrolide (azitromicin) u trajanju od 5 do 6 dana, a njih 13 (11,50%) bi se odlučilo na neku drugu skupinu antibiotika. Ispitivanjem nije utvrđena povezanost između odabira i trajanja antibiotske terapije s obzirom na dob (P=0,182) i specijalistički status liječnika (P=0,650). Ispitivanjem je utvrđena statistički značajna razlika u zastupljenosti stava liječnika kako nitko iz skupine pacijenata sa nekompliciranom infekcijom koji može imati vrućicu, opću slabost, glavobolju, mialgiju, hunjavicu, grlobolju i/ili kašalj ne treba dobiti antibiotik (P, OBJECTIVES: The main goal of this research was to assess the attitudes and knowledge of family medicine doctors in the Split-Dalmatia County regarding the prescription of antibiotic therapy to patients with COVID-19, as well as their familiarity with the most recent guidelines for the treatment of coronavirus disease and vitamin D replacement therapy. PATIENTS AND METHODS: A study was conducted to examine the attitudes and knowledge of family medicine doctors in the Split-Dalmatia County regarding the use of antibiotic therapy in COVID-19 positive patients. The data was collected by solving a questionnaire via the Google forms internet application during May 2022. The questionnaire consisted of 23 questions. The statistical software STATISTICA 12, Tibco, California, was used for data analysis. RESULTS: The majority of family medicine doctors, 100 (87%), are familiar with the latest guidelines for the treatment of patients with the coronavirus disease (version 5 of February 8, 2022). Out of 115 doctors, 88 of them (77.2%) fully agree with the recommendation that the basis of the treatment of the COVID-19 infection is the alleviation of symptoms, complications and progression of the disease, as well as symptomatic and supportive treatment. The largest number of LOMs believe that they prescribed antibiotic therapy for 10-30% of patients (41.67%). When asked about the choice and duration of antibiotic therapy in COVID-19 positive patients, 80 (70.80%) LOM would decide on a macrolide (azithromycin) for 3 days, 20 of them (17.70%) on a macrolide (azithromycin) in lasting from 5 to 6 days, and 13 of them (11.50%) would have opted for another group of antibiotics. The examination did not establish a connection between the selection and duration of antibiotic therapy with regard to age (P=0.182) and specialist status of the doctor (P=0.650). The examination revealed a statistically significant difference in the representation of the physician's opinion that no one from the group of patients with an uncomplicated infection who may have fever, general weakness, headache, myalgia, wheezing, sore throat and/or cough should receive antibiotics (P

Makrolidni antibiotici koriste se u posljednjih 60 godina najčešće za liječenje infekcija gornjih i donjih dišnih puteva. Djelovanje makrolidnih antibiotika temelji se na inhibiciji sinteze bakterijskih proteina vezanjem antibiotika na 50S podjedinicu bakterijskog ribosoma. Povećanje bakterijske rezistencije na postojeće makrolidne antibiotike dovelo je do potrebe za otkrićem i razvojem novih antibiotika. Za potpuno razumijevanje bioaktivnosti makrolidnog antibiotika, potrebno je odrediti konformacije u slobodnom i vezanom stanju te istražiti interakcije s bakterijskim ribosomom. U ovom radu okarakteriziran je 9a-aminopropil derivat azitromicina, prekursor u sintezi makrozona, pomoću jedno- i dvodimenzijskih tehnika NMR. Makrozoni su konjugati azitromicina i tiosemikarbazona. Pomoću tehnike NOESY određena je konformacija prekursora u slobodnom stanju. Konformacija prekursora u vezanom stanju određena je pomoću tehnike trNOESY, dok su interakcije prekursora s ribosomom izoliranim iz bakterije Escherichia coli istražene pomoću tehnike STD. Macrolide antibiotics have been used over the past 60 years, to treat upper and lower respiratory tract infections. Macrolides inhibit bacterial protein synthesis by binding to the 50S subunit of the bacterial ribosome. Increasing bacterial resistance to existing macrolide antibiotics has speed up the process of discovery of new antibiotics. To fully understand bioactivity of macrolide antibiotics, it is necessary to determine structure and conformation in the free and bound states and to investigate the interactions with the bacterial ribosome. In this paper, the structure of 9a-aminopropyl derivative of azithromycin, a precursor in the synthesis of the corresponding macrozone, was characterized by using one- and two-dimensional NMR spectroscopy. Macrozones are conjugates of azithromycin and thiosemicarbazones. The conformation of the precursor in the free and bound states was determined by NOESY and transferred NOESY spectral analysis, while the interactions with the ribosome isolated from Escherichia coli were investigated using the STD technique.

Cilj istraživanja Jedan od ciljeva ovog istraživanja bio je ispitati intrinzični antimikrobni učinak izvornih kitozanskih hidrogelova koji su korišteni kao polučvrste podloge za uklapanje liposomskih disperzija azitromicina. Kako bi se utvrdilo utječe li kitozan na antibakterijsku aktivnost liposomski uklopljenog antibiotika te doprinosi li fosfolipidni sastav liposoma antimikrobnoj aktivnosti, ispitan je antibakterijski učinak kitozanskih hidrogelova s liposomski uklopljenim azitromicinom, a dobiveni rezultati uspoređivani su s antibakterijskom aktivnosti: liposoma s azitromicinom, otopine azitromicina, kitozanskih hidrogelova s azitromicinom te izvornih kitozanskih hidrogelova. Materijali i metode Eksperimentalni dio rada obuhvaćao je izradu kitozanskih hidrogelova iz kitozana različitih molekularnih masa, pripravu nekoliko tipova liposoma s uklopljenim azitromicinom, te izradu liposomskih hidrogelova umješavanjem liposoma u kitozanske hidrogelove. Ispitivanja intrinzičnog antimikrobnog učinka izvornih kitozanskih hidrogelova provedena su tako da su hidrogelovi inokulirani određenim test mikroorganizmima (Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Candida albicans i Aspergillus brasiliensis) i čuvani u propisanim uvjetima, a redukcija broja mikroorganizama praćena je kroz određene vremenske intervale tijekom 28 dana. Ispitivanje utjecaja kitozana na antibakterijsku aktivnost različitih liposoma s azitromicinom provedeno je primjenom in vitro agar-difuzijskog testa na bakterijskom soju Staphylococcus aureus ATCC 6538. Rezultati Neovisno o tipu kitozana iz kojeg su pripravljeni, svi izvorni kitozanski hidrogelovi su pokazali intrinzičnu antimikrobnu aktivnost na ispitivane mikroorganizme, čime je potvrđeno da nije potrebno dodatno konzerviranje topikalnih formulacija koje sadrže kitozanske hidrogelove kao polučvrste podloge. Nadalje, čvrstoća/elastičnost liposomskih dvoslojeva utjecala je na antibakterijsku aktivnost ispitivanih liposoma s azitromicinom i liposomskih hidrogelova. Pritom je najjači učinak postignut s liposomima karakteriziranim izrazito čvrstim (rigidnim) dvoslojevima (konvencionalnim liposomima). Svi kitozanski hidrogelovi potencirali su antibakterijski učinak liposomski uklopljenog azitromicina, što je potvrđeno većim zonama inhibicije u odnosu na one postignute otopinom azitromicina. Zaključak Rezultati svih provedenih ispitivanja ukazuju na mogućnost primjene kitozanskih hidrogelova kao podloga za topikalne lijekove i nanoterapeutike namijenjene liječenju infekcija kože. Međutim, neophodno je provesti daljnja in vivo ispitivanja kako bi se dokazala njihova terapijska učinkovitost. Objectives One of objectives of this study was to examine the intrinsic antimicrobial activity of the chitosan hydrogels, which were used as vehicles (bases) for incorporation of the azithromycin liposomes. To determine whether chitosan hydrogels could potentiate antibacterial activity of the incorporated azithromycin liposomes and to examine the effect of bilayer fluidity on the antibacterial activity of encapsulated azithromycin, the studies were performed with chitosan hydrogels containing incorporated azithromycin liposomes and the results were compared with azithromycin liposomes (liquid dispersions), solution of azithromycin, hydrogels containing azithromycin solution and the plain chitosan hydrogels. Material and Methods The experimental part of this study included preparation of the plain chitosan hydrogels from the different molecular weight chitosans as well as preparation and characterization of the several types of liposomes encapsulating azithromycin, which were incorporated into the chitosan hydrogels (liposomal hydrogels). Intrinsic antimicrobial activities of the plain chitosan hydrogels were examined by the reduction of number of the test microorganisms (Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Candida albicans and Aspergillus brasiliensis), which were inoculated into the hydrogels, stored for 28 days and tested at determined time intervals. In vitro antibacterial activity of the different azithromycin containing liposomal hydrogels toward Staphylococcus aureus ATCC 6538 was performed by agar well diffusion method. Results All the plain chitosan hydrogels have shown intrinsic antimicrobial activities, confirming that there is no need for the use of preservatives in the topical formulations based on the chitosan hydrogels. Furthermore, the elasticities/rigidities of the different azithromycin liposomes have influenced their antibacterial activities as well as antibacterial activities of the liposomal hydrogels. Conventional liposomes, characterized by rigid bilayers, have demonstrated the most pronounced antibacterial activity. All the chitosan hydrogels contributed the antibacterial activity of the incorporated azithromycin liposomes, what was confirmed by a larger inhibition zones formed, compared to those obtained by hydrogels containing azithromycin solution and the zones formed by the free drug. Conclusion Considering all the results, it can be assumed that the chitosan hydrogels could be used as vehicles for topical formulations and liposomes aimed for the treatment of skin infections. However, further in vivo studies should be carried out to confirm the efficacy of the innovative formulations.

U sklopu ovoga diplomskoga rada provedeno je ispitivanje kako promjena glikozilacijskog uzorka α1-kiselog glikoproteina utječe na konstantu vezanja dipiridamola, imatiniba i azitromicina. Tehnika korištena u ispitivanju bila je fluorescencijska spektroskopija. Rezultati ispitivanja ukazuju na to da desijalinizacija α1-kiselog glikoproteina uzrokuje promjene u konstantama vezanja izabranih lijekova i α1-kiselog glikoproteina. Desijalinizacija navedenog proteina kod imatiniba uzrokuje blago povećanje konstante vezanja za razliku od dipiridamola gdje je učinak suprotan, odnosno nakon desijalinizacije α1-kiselog glikoproteina dipiridamol se slabije veže za takav oblik te je manja i konstanta vezanja. Zbog male konstante vezanje i premalog raspona koncentracija, mjerenja provedena za azitromicin nisu relevantna. Within the scope of this diploma thesis, the effect of the binding constant of dipyridamole, imatinib and azithromycin on the α1-acid glycoprotein due to a change in the pattern of it's glycosylation was examined by fluorescence spectroscopy. Result indicates effect of desialylation of α1-acid glycoprotein on binding constants of selected drugs. Desialylation of α1-acid glycoprotein will slightly increase the binding constant for imatinib. On the other hand, desialylation of the same protein will reduce the binding constant for dipyridamole. Due to small binding constant and narrow range of used concentrations, measurements of azithromycin binding was not succesfull.

Makrolidni antibiotici (npr. azitromicin) imaju široku primjenu u liječenju infekcija uzrokovanih Gram-pozitivnim, Gram-negativnim bakterijama i nekim unutarstaničnim organizmima, te su ujedno pokazali antiupalno i antikancerogeno djelovanje. Djeluju na principu inhibicije sinteze bakterijskog proteina vežući se na 50S podjedinicu bakterijskog ribosoma. S obzirom na sve bržu prilagodbu bakterija i njihovu rezistenciju na makrolidne antibiotike, javlja se potreba za novim spojevima koji će pokazati veću biološku aktivnost. Osim makrolida, dobru antibakterijsku aktivnost su pokazali i tiosemikarbazoni. Makrozoni predstavljaju nove konjugate derivata azitromicina i tiosemikarbazona. U ovom radu uspješno je pripravljeno pet makrozona reakcijom 4"-aminopropilnog derivata azitromicina i odgovarajućih tiosemikarbazona. Derivat azitromicina pročišćen je kromatografijom na stupcu, a makrozoni tehnikom LC-SPE i semipreparativnom tekućinskom kromatografijom. Pripravljeni i pročišćeni makrozoni okarakterizirani su: spektrometrijom mase, spektroskopijom NMR i tekućinskom kromatografijom visoke djelotvornosti. Idući korak istraživanja uključit će in vitro testove biološke aktivnosti pripravljenih makrozona na susceptibilne i rezistentne sojeve bakterija. Macrolide antibiotics (e.g. azithromycin) are widely used in the treatment of infectious deseases caused by Gram-positive and Gram-negative bacteria and some intracellular organisms. They have also shown anti-inflammatory and anticancer activities. Macrolides exert their activity by binding to the 50S subunit of the bacterial ribosome thus inhibiting bacterial protein synthesis. Owing to the fact that bacteria develop resistance to existing antibiotics, new chemical entities are urgently needed to fight resistance mechanisms. Thiosemicarbazones have also shown good antibacterial activity. Hence, we believe that conjugation of azithromycin and thiosemicarbazones could yield a novel class of bioactive compounds, the macrozones. Here we report synthesis and characterization of five macrozones by reaction of 4“-aminopropyl azithromycin and the corresponding thiosemicarbazones. All prepared compounds were purified by column and semipreparative liquid chromatography and LC-SPE system. The next step of the study will include in vitro biological evaluation against a panel of susceptible and resistant bacteria strains.

Makrolidni antibiotici, kao što je azitromicin, imaju široku primjenu u liječenju infekcija dišnih puteva. Djeluju kao inhibitori sinteze proteina vežući se na bakterijsku ribosomsku podjedinicu 50S. Zbog povećane rezistencije bakterija javlja se potreba za razvojem novih, poboljšanih antibiotika. Dobru antibakterijsku aktivnost in vitro na odabrane Gram-pozitivne i Gram-negativne bakterije pokazuju makrozoni, novi konjugati azitromicina i tiosemikarbazida. Konjugacijom 9a-(γ-aminopropil) derivata azitromicina i novosintetiziranih tiosemikarbazida e1-e5 pripravljeno je pet novih spojeva, f1-f5. Sintetizirani spojevi pročišćeni su kolonskom kromatografijom, a struktura im je određena na temelju snimljenih jedno- i dvodimenzijskih spektara NMR te spektara MS. Budućim istraživanjima ispitat će se biološka aktivnost pripravljenih spojeva. Macrolide antibiotics, such as azithromycin, have been widely prescribed for the treatment of respiratory tract infections. They act as inhibitors of protein synthesis by binding to the bacterial ribosomal subunit 50S. Drug resistant microbial pathogens today represent a serious global problem that requires the discovery of new and more effective antimicrobial agents. Macrozones are novel conjugates of azithromycin and thiosemicarbazides that shows very good antibacterial activity. Five new compounds f1-f5 were prepared by conjugation of the 9a-(γ-aminopropil) azithromycin derivative and the newly synthesized thiosemicarbazides e1-e5. The synthesized compounds were purified by column chromatography. The structures of novel macrozones were proposed on the basis of recorded one- and two-dimensional NMR spectra and the MS spectra, respectively.

Makrolidni antibiotici, poput azitromicina, koriste se u liječenju infekcija uzrokovanih Gram-pozitivnim i Gram-negativnim bakterijama. Djeluju kao inhibitori sinteze bakterijskih proteina tako što se vežu na 50S podjedinicu bakterijskog ribosoma u blizini centra peptidil-transferaze. Učestalo korištenje makrolidnih antibiotika dovelo je do pojave i razvoja bakterijske rezistencije. Iz tog razloga javlja se potreba za otkrićem i razvojem novih antibiotika. Za razumijevanje cjelokupnog biološkog profila novog potencijalnog lijeka važno je odrediti konformaciju molekule u slobodnom i vezanom stanju te istražiti interakcije s biološkim metama. U ovom radu uspješno je pripravljen 3-aminopropil dekladinozil derivat azitromicina (S6), prekursor u sintezi makrozona, novih bioaktivnih konjugata azitromicina i tiosemikarbazona. Pripravljeni spoj okarakteriziran je spektroskopijom NMR i spektrometrijom masa. Upotrebom tehnike ROESY određena je konformacija prekursora u slobodnom stanju, a pomoću tehnika tr-NOESY i STD istražene su interakcije prekursora s ribosomom izoliranim iz bakterije Escherichia coli te je određena konformacija u vezanom stanju. Macrolide antibiotics, such as azithromycin, are used to treat infections caused by Gram-positive and Gram-negative bacteria. They act as inhibitors of bacterial protein synthesis by binding to the 50S subunit of the bacterial ribosome at or near the peptidyl transferase center. Frequent use of macrolide antibiotics has led to the emergence of bacterial resistance. For this reason, there is a need to discover and develop new antibiotics. To understand the overall biological profile of a new potential drug, it is important to determine the conformation of molecules in the free and bound state and to investigate interactions with biological targets. In this work, 3-aminopropyl decladinosyl derivative of azithromycin (S6), a precursor in the synthesis of macrozones, a new bioactive conjugate of azithromycin and thiosemicarbazone, was successfully prepared. The compound S6 was characterized by NMR spectroscopy and mass spectrometry. ROESY technique was applied to estimate free conformation in solution. The interactions of S6 with the ribosome isolated from Escherichia coli were investigated by using tr-NOESY and STD techniques. The analysis of the recorded spectra enabled determination of conformation in bound state.

Makrozoni su novi konjugati azitromicina i derivata tiosemikarbazida koji pokazuju zadovoljavajuće antibakterijsko djelovanje in vitro na odabrane Gram-pozitivne i Gram-negativne bakterije. Zbog sve učestalije pojave bakterija rezistentnih na makrolide, nastavljaju se istraživanja s ciljem razvoja novih oblika makrolidnih antibiotika koji će biti djelotvorni na otporne bakterijske sojeve. Izolacija i određivanje strukture komponenata reakcijske smjese 9a-(2-feniletil)makrozona u ovome radu postignuta je primjenom vezanog sustava LC-SPE/NMR. Istraženi su uvjeti za uspješnu izolaciju i zadržavanje analita na SPE-nosačima. Makrozon je pokazao dobro zadržavanje na nepolarnoj nepokretnoj fazi SPE-nosača baziranoj na silika-materijalu, dok se veći broj polarnih onečišćenja zadržao bolje na divinilbenzenskim polimernim SPE-nosačima. Uz to, većina polarnih onečišćenja uspješno je ekstrahirana koristeći pritom veći dodatni protok vodene pokretne faze. Na temelju snimljenih jedno- i dvodimenzijskih spektara NMR, te analizom spektara MS, predložene su strukture izoliranih komponenata. Macrozones are new conjugates of azithromycin analogues, 9a-(γ-aminopropyl) derivatives, and thiosemicarbazides which show good in vitro activity against selected Gram-positive and Gram-negative bacterial strains. Due to the increasing number of macrolide-resistant bacteria, a search for new antibiotics continues, aimed at discovery of more effective antimicrobial agents. In this work, isolation and determination of the 9a-(2-phenylethyl)macrozone reaction mixture components was achieved using the LC-SPE/NMR system. Several parameters for successful separation and extraction of analytes were optimised. It was found that silica-bonded phase gave satisfactory results for the main compound, macrozone, however, for more polar compounds divinylbenzene polymer phase was better. Additionally, extraction efficiency for more polar impurities was obtained by using higher make-up flow. Structures of the isolated components were proposed on the basis of the recorded one- and two-dimensional NMR spectra and the MS spectra, respectively.

Različite vrste liposoma, konvencionalni, deformabilni, propilenglikol i kationski liposomi s uklopljenim azitromicinom, pripremljeni su film metodom. Ekstruzijom kroz polikarbonske membrane, formulacije liposoma su homogenizirane (indeks polidisperznosti < 0,43), a srednji promjeri značajno smanjeni (< 250 nm). Fosfolipidni sastav liposoma imao je utjecaja na površinski naboj liposoma. Konvencionalni, deformabilni i propilenglikol liposomi imali su zeta potencijale od -48 do -44 mV, dok je zeta potencijal kationskih liposoma iznosio + 62 mV. In vitro ispitivanja biokompatibilnosti liposoma s uklopljenim azitromicinom provedena na HaCaT stanicama procijenjena su pomoću MTT testa, mjerenjem metaboličke aktivnosti keratinocita 24 sata nakon inkubacije liposoma s HaCaT stanicama. Uklapanjem u liposome citotoksičnost azitromicina je smanjena. Fosfolipidni sastav i prisustvo suotapala u liposomima su imali utjecaja na biokompatibilnost. Propilenglikol je pokazao povoljan učinak na stanice keratinocita, dok su natrijev deoksikolat i DODAB pokazali potencijalnu citotoksičnost, ali tek pri izuzetno visokim koncentracijama lipida. Imajući u vidu koncentracije azitromicina u liposomima s kojima je postignut odgovarajući antibakterijski učinak, svi ispitivani tipovi liposoma su bili biokompatibilni s HaCaT stanicama. Conventional, deformable, propylene glycol and cationic liposomes encapsulating azithromycin were prepared by the film hydration method. Extrusion of the liposomal dispersions through the polycarbonate membranes resulted with liposomes of more homogenous size distributions with a polydispersity indexes < 0,43 nm and a significantly reduced mean diameters (< 250 nm). The (phospo)lipid composition affected the surface charge of the liposomes. Zeta potential values for the conventional, deformable and propylene glycol liposomes ranged from -48 mV to -44 mV, while the cationic liposomes were shown to have a positive zeta potential of +62 mV. In vitro biocompatibilities of the different types of liposomes containing azithromycin with the human HaCaT cells were evaluated 24 h after the incubation of the cells with the liposomes using the cellular metabolic activity (MTT assay). The encapsulation of azithromycin in liposomes reduced the cytotoxicity of the drug, whereas the (phospo)lipid composition and the presence of the surfactant and cosolvent affected the biocompatibility. Propylene glycol was well tolerated by the keratinocytes in comparison to sodium deoxycholate and DODAB, where potential cytotoxicities were observed. All types of the liposomes were biocompatible with the keratinocytes even at the very high concentrations of azithromycin tested.

Cilj ovog rada bio je provesti in vitro ispitivanje oslobađanja azitromicina iz konvencionalnih i deformabilnih liposoma uklopljenih u kitozanski gel, inovativne formulacije namijenjene lokalnoj primjeni na kožu. Liposomi su pripremljeni metodom hidratacije suhog fosfolipidnog sloja uz ekstruziju kroz polikarbonatne membrane od 400 i 200 nm, dok je gel izrađen iz kitozana velike molekulske mase. Deformabilni liposomi bili su manjeg srednjeg promjera, ali nešto većeg indeksa polidisperznosti od konvencionalnih liposoma, što je posljedica veće elastičnosti fosfolipidnih dvoslojeva. Oba tipa liposoma imali su negativni zeta potencijal kao preduvjet fizičke stabilnosti liposomskih disperzij. Liposomi, odijeljeni od neuklopljene frakcije azitromicina, umiješani su u 1.5% kitozanski gel u koncentraciji 30% (w/w). In vitro ispitivanja oslobađanja azitromicina iz liposoma uklopljenih u kitozanski gel potvrdila su produljeno oslobađanje antibiotika u usporedbi s kontrolnim gelom (otopina azitromicina u gelu). Usporedba profila oslobađanja antibiotika iz različitih tipova liposoma-u-gelu pokazala je sporije oslobađanje iz konvencionalnih liposoma zbog manje propusnosti membrane za ukloljeni azitromicin u odnosu na deformabilne liposome. The aim of this study was to evaluate in vitro release of azithromycin from conventional and deformable liposomes incorporated into chitosan gel. Liposomes were prepared by film hydration method and extrusion through polycarbonate membranes of 400 and 200 nm, while the gel was prepared using high molecular weight chitosan. Deformable liposomes were of smaller size, but somewhat higher polydispersity index than conventional liposomes. Both types of liposomes had negative zeta potentials as a precondition for physical stability of liposomal dispersions. Liposomes, separated from unentrapped azithromycin, were mixed into 1.5% chitosan gel at a concentration of 30% (w/w, liposomes/gel). In vitro assessment of azithromycin release from the different liposomes incorporated into chitosan gel have confirmed prolonged release of the drug from the both types of liposomes compared to control gel (azithromycin solution incorporated in gel). Comparison of the release profiles of azithromycin from the different types of liposomal gels showed slower release from conventional liposomes.

CILJ RADA Azitromicin je makrolidni antibiotik koji se koristi za liječenje bakterijskih infekcija. Danas se nalazi na tržištu u obliku tableta, sirupa, kapsula i injekcija. Jedno od analitičkih ispitivanja koja se provode u kontroli kvalitete gotovih proizvoda azitromicina je analiza onečišćenja (srodnih spojeva i razgradnih produkata). Cilj ovog rada je opisati i objasniti kromatografske metode za analizu onečišćenja u proizvodima azitromicina koje se koriste u kontroli njihove kvalitete. Prikazane su sličnosti i razlike navedenih metoda te su uspoređene s farmakopejskim metodama za analizu onečišćenja azitromicina. MATERIJALI I METODE Onečišćenja mogu nastati tijekom proizvodnje ili skladištenja farmaceutskog proizvoda i njihov sadržaj se mora kontrolirati. Onečišćenja azitromicina se analiziraju tehnikom tekućinske kromatografije visoke djelotvornosti (HPLC) u svim vrstama dozirnih oblika. U analitici azitromicinskih proizvoda UV detektori se koriste za detekciju onečišćenja u tabletama, kapsulama i injekcijama azitromicina, dok se elektrokemijski detektor (EC) koristi za detekciju onečišćenja u sirupima azitromicina. Opisani su eksperimentalni uvjeti HPLC metoda: vrste stacionarnih faza, vrste i brzine protoka mobilnih faza, gradijente eluacije, načini detekcije, pripreme otopina standarda i ispitivanje prikladnosti sustava. U tablicama su prikazani kromatografski parametri za identifikaciju pikova onečišćenja u azitromicinskim proizvodima. RASPRAVA Metode za analizu onečišćenja u tabletama, kapsulama i injekcijama azitromicina koriste istu kolonu, iste instrumentalne uvjete, iste kalibracijske i identifikacijske otopine Postoji razlika u zahtjevima za provjeru prikladnosti sustava i na nju treba obratiti pažnju ukoliko se paralelno na istom instrumentu analiziraju uzorci tableta, kapsula ili injekcija. Ukoliko se provjere svi uvjeti za ispitivanje prikladnosti sustava nema prepreke za analizu ovih vrsta uzoraka paralelno na istom sustavu. Slično se može zaključiti za uzorke sirupa azitromicina. Sve doze i formulacije sirupa se mogu analizirati paralelno na istom sustavu s elektrokemijskim detektorom ukoliko se obrati pažnja na ispunjavanje svih uvjeta prikladnosti sustava. ZAKLJUČAK U redovnoj analitici farmaceutskih proizvoda u laboratorijima kontrole kvalitete od velike je važnosti da se analize različitih doza ili formulacija s istom djelatnom tvari mogu raditi zajedno na istom kromatografskom sustavu. Ukoliko usporedimo metodu za analizu onečišćenja u azitromicin injekcijama koja se koristi u Plivi s onom u farmakopeji, vidljivo je da je farmakopejska metoda složenija i da bi samo za analizu onečišćenja injekcija vjerojatno trebalo više radnih dana i više analitičara. Tako bi se smanjila produktivnost laboratorija i broj analiza koje se izvrše u jedinici vremena, što u konačnici može dovesti do skupljeg lijeka za pacijenta. AIM OF WORK Azithromycin is a macrolide antibiotic used to treat bacterial infections. Today, it is marketed in the form of tablets, syrups, capsules and injections. One of the analytical tests carried out in the quality control of finished products of azithromycin is the analysis of impurities (related compounds and degradation products). The aim of this paper is to describe and explain chromatographic methods for analysis of impurities in azithromycin products which are used in their quality control. The similarities and differences of the methods are shown and compared with the pharmacopoeial methods for the analysis of azithromycin degradation products. MATERIALS AND METHODS Amount of degradation products may arise during the manufacture or storage of a pharmaceutical product and their content must be controlled. Azithromycin degradation products are analyzed by High Performance Liquid Chromatography (HPLC) in all types of dosage forms. In the analysis of azithromycin finished products UV detectors are used for detecting impurities in tablets, capsules and azithromycin injections, while the electrochemical detector (EC) is used to detect impurities in azithromycin syrups. The experimental conditions of the HPLC methods are described: stationary phase types, types and rates of mobile phase flow, gradient elution, detection methods, standard solution preparation and system suitability test. The tables show the chromatographic parameters used for the identification of degradation product peaks in azithromycin products. DISCUSSION Methods for analysis of degradation products in tablets, capsules and azithromycin injections use the same column, the same instrumental conditions, the same calibration and identification solutions. There is a difference in system suitability check requirements and attention should be paid if the samples of tablets, capsules or injections are analyzed together. If all the conditions for the system suitability test are checked, there are no obstacles to the analysis of these types of samples in parallel in the same system. Similar can be concluded for samples of azithromycin syrups. All dosages and formulations of the syrups can be analyzed together on the same system with an electrochemical detector if attention is paid to the fulfillment of all system suitability conditions. CONCLUSION In a regular analytics of pharmaceutical products in quality control labs it is of great importance that analyzes of different doses or formulations with the same active substance can be performed together on the same chromatographic system. If we compare the method for the analysis of azithromycin injections used in Pliva with those in Pharmacopoeia, it is apparent that the pharmacopoeia method is more complex and that it would probably take more days and more analysts to analyze degradation products in samples of azithromycin injections. This would reduce the laboratory's productivity and the number of analyzes performed in the unit of time, which ultimately can lead to a more expensive drug for the patient.

Učinkovita topikalna terapija infekcija kože te prevencija stvaranja biofilmova i razvoja rezistencije mikroorganizama od velikog su medicinskog značaja. Cilj ovog rada bio je razviti biorazgradljiv, djelotvoran i siguran terapijski sustav za topikalnu antimikrobnu terapiju, zasnovan na fosfolipidnim vezikulama (liposomima) uklopljenima u kitozansku podlogu. U tu svrhu pripravljeno je i evaluirano nekoliko tipova liposoma s uklopljenim azitromicinom (AZT) različitog naboja na površini i čvrstoće/elastičnosti fosfolipidnih dvoslojeva: konvencionalni liposomi (CL), deformabilni liposomi (DL), propilenglikol liposomi (PGL) i kationski liposomi (CATL). Čvrstoća/elastičnost fosfolipidnih dvoslojeva AZT-liposoma utjecala je na fizičku stabilnost, profil oslobađanja AZT-a i lokalizaciju antibiotika u koži. Najsporije oslobađanje AZT-a, te odlaganje na površini kože, postignuto je CATL-om, karakteriziranim čvrstim dvoslojevima. Elastični liposomi (DL i PGL) pokazali su brže oslobađanje te nakupljanje AZT-a u unutrašnjosti kože. Svi AZT-liposomi pokazali su antibakterijski učinak na MRSA kliničke izolate, pri čemu su minimalne biofilm inhibitorne koncentracije bile i do 32 puta manje od onih utvrđenih za AZT-otopinu. Najjači anti-MRSA učinak imali su CATL, potom slijede DL, PGL i CL. Svi AZT-liposomi bili su biokompatibilni s keratinocitima i fibroblastima, čak i pri značajno većim koncentracijama AZT-a od pripadajućih minimalnih inhibitornih koncentracija i minimalnih biofilm inhibitornih koncentracija. Uklapanjem AZT-liposoma u kitozansku podlogu produljeno je oslobađanje AZT-a i postignuta prikladna viskoznost za topikalnu primjenu na kožu. Anti-MRSA učinak svih formulacija liposomi-u-podlozi bio je znatno veći od onog postignutog s kontrolom. Teksturna analiza ispitivanih formulacija liposomi-u-podlozi pokazala je prikladnu čvrstoću, kohezivnost i adhezivnost formulacija, posebice PGL-u-podlozi, a koje su direktno povezane s njihovom primjenom i zadržavanjem na koži. Uzimajući u obzir rezultate svih provedenih ispitivanja, formulacija CATL-u-podlozi bila bi prikladna za liječenje MRSA-om uzrokovanih infekcija kod kojih je potrebna lokalizacija antibiotika u površinskim slojevima kože, dok bi formulacija PGL-u-podlozi omogućila bolju dostavu antibiotika u dublje slojeve kože. Međutim, potrebno je provesti in vivo animalna ispitivanja terapijske učinkovitosti kako bi se te pretpostavke i potvrdile. Effective topical therapy of complicated skin infections, prevention of biofilm formation and antimicrobial resistance is of great medical importance. The aim of this research was to develop a biodegradable, efficient and safe delivery system for topical antimicrobial therapy, based on the drug-loaded phospholipid vesicles (liposomes) incorporated into the chitosan-based vehicle. For that purpose several types of azithromycin-loaded (AZT) liposomes with different surface charges and rigidity/elasticity of (phospho)lipid bilayers have been prepared and evaluated: conventional liposomes (CLs), deformable liposomes (DLs), propylene glycol-embedding liposomes (PGLs) and cationic liposomes (CATLs). The rigidity/elasticity of AZT-loaded liposomes influenced their physical stability, AZT release profile and its localization inside the skin. The slowest release and deposition of AZT on the skin surface was achieved by CATLs, characterized with rigid bilayers, whereas elastic liposomes (DLs and PGLs) demostrated faster release and accumulation of the drug within the skin. All AZT-loaded liposomes showed antibacterial activity against MRSA clinical isolates, with minimal biofilm inhibitory concentrations up to 32-fold lower than those determined for the AZT-solution. The strongest anti-MRSA effect was demonstrated by CATLs, followed by DLs, PGLs and CLs. All AZT-loaded liposomes were biocompatible with keratinocytes and fibroblasts, even at AZT concentrations significantly higher than the corresponding minimal inhibitory concentrations and minimal biofilm inhibitory concentrations. Incorporation of AZT-loaded liposomes into the chitosan vehicle resulted in the prolonged AZT release and viscosity appropriate for skin application. AZT-liposomes incorporated into chitosan vehicle, especially CATL-in-vehicle, demonstrated significantly enhanced anti-MRSA effect, as compared to the chitosan vehicle embedding AZT-solution. Texture analysis of AZT-liposomes incoroporated into chitosan vehicle showed a suitable hardness, cohesiveness and adhesiveness, in particular PGL-in-vehicle, which are directly related to their skin application and retention. Considering the results of all the experiments performed, CATL-in-vehicle would be suitable for the treatment of MRSA-infections requiring localization of the antibiotic in the upper layers of the skin, while PGL-in-vehicle facilitates delivery of AZT into deeper skin layers. However, further in vivo animal studies are necessary to confirm those assumptions.

Svrha ovog rada bila je ispitati biokompatibilnost/citotoksičnost različitih tipova liposoma s azitromicinom na MJ90hTERT fibroblastima. Konvencionalni, deformabilni, propilenglikol i kationski liposomi s azitromicinom međusobno su se razlikovali po (fosfo)lipidnom sastavu i prisustvu suotapala (propilenglikol). Liposomi su pripravljeni tzv. film metodom, a primjenom ekstruzije postignuta je prikladna veličina liposoma za topikalnu primjenu. Sastav liposoma utjecao je na fizikalna svojstva liposoma, posebice naboj na površini i uspješnost uklapanja azitromicina. Provedena in vitro ispitivanja citotoksičnosti pokazala su biokompatibilnost svih ispitivanih liposoma s fibroblastima u koncentracijama azitromicina s kojima se postiže adekvatan antimikrobni učinak liposomskih formulacija. Usporedba različitih tipova liposoma pokazala je najbolju kompatibilnost propilenglikol liposoma, a najmanju kationskih liposoma koja je posljedica prisutnosti kationskog lipida (DODAB-a) u fosfolipidnom dvosloju. The aim of this study was to investigate the biocompatibility/cytotoxicity of the different types of azithromycin-loaded liposomes on the MJ90hTERT fibroblasts. Conventional, deformable, propylene glycol and cationic liposomes encapsulating azithromycin, and differing in (phospho)lipid composition and the presence of co-solvent (propylene glycol), were prepared using the film hydration method. The suitable size of the liposomes for topical application was achieved by their extrusion through the membranes of 400 and 200 nm. The composition of liposomes has proven to influence the physical properties of liposomes, particularly the surface charge and the entrapment efficiency of azithromycin. All the azithromycin-loaded liposomes were biocompatible with the fibroblasts at azithromycin concentrations showing appropriate antibacterial effect. The comparison of the different types of liposomes demonstrated the highest level of compatibility with the MJ90hTERT cells for propylene glycol liposomes and the lowest for cationic liposomes, due to the presence of the cationic lipid (DODAB) in the (phospho)lipid bilayers.

Azitromicin (AZM) je makrolidni antibiotik širokog spektra djelovanja čije je prisustvo detektirano u vodnom okolišu. Pripada skupini „novih onečišćujućih tvari“ te se vrlo teško uklanja konvencionalnim metodama obrade otpadnih voda. Iako su farmaceutici u okolišu općenito prisutni u niskim koncentracijama, njihovo dugoročno ispuštanje može ostaviti neizbrisive posljedice na sve okolišne sastavnice, ali i na ljudsko zdravlje. Kako bi se spriječio njihov ulazak u okoliš, potreban je razvoj i primjena novih i učinkovitih metoda koje će u kombinaciji s konvencionalnim metodama obrade uspjeti učinkovito ukloniti navedene tvari. Jedna od metoda koja se pokazala učinkovitom pri uklanjanju farmaceutika jesu napredni oksidacijski procesi. Cilj ovog rada bilo je odrediti toksičnost vodene otopine azitromicina početne masene koncentracije od 10 mg/L nakon provođenja dvije vrste naprednih oksidacijskih procesa, direktne UV-fotolize i fotokatalize s TiO2 uz primjenu UV-A i UV-C zračenja. Procjena toksičnosti provedena je korištenjem bioluminiscentne metode za određivanje akutne toksičnosti s bakterijama Vibrio fischeri. Osim navedenog, pomoću tekućinske tekućinske kromatografije visoke djelotvornosti s tandemskom spektrometrijom masa detektirani su i identificirani nastali razgradni produkti ispitivanog spoja nakon tretiranja naprednim oksidacijskim postupcima. Određivanje toksičnosti je u ovom slučaju od ključne važnosti budući da nastali razgradni produkti mogu biti toksičniji od polaznih tvari. Ispitivanjima se pokazalo kako nitijedna od ispitivanih otopina azitromicina nakon primjene naprednih oksidacijskih procesa nije toksična, bez obzira na prisutnost nastalih razgradnih produkata. Nastajanje razgradnih produkata zabilježeno je tijekom provođenja procesa fotokatalize s TiO2 pri valnoj duljini zračenja od 254 nm. Azithromycin (AZ) is a wide-spectrum macrolide antibiotic which presence is detected in the aquatic environment. It belongs to the group of „new emerging contamintants“ and it is very difficult removed by conventional wastewater treatment methods. Even though pharmaceuticals are generally present in the environment in low concentrations, their long-term discharge can leave irreversible consequences on all environmental constituents and also on human health. In order to prevent their entry into the environment, development and application of new and effective methods will be required and which, in combination with conventional treatment methods, will be effective in removing these substances. One of the methods that has proved effective in removal of pharmaceuticals are advanced oxidation processes. The aim of this paper was to determine the toxicity of aqueous azithromycin solution with initial mass concentration of 10 mg/L after performing two types of advanced oxidation processes, direct UV photolysis and photocatalysis with TiO2 by using UV-A and UV-C radiation. The toxicity determination was performed by using a bioluminescent method for the determination of acute toxicity with Vibrio fischeri bacteria. Also, by high performance liquid chromatography-tandem mass spectrometry, degradation products were detected and identified after the treatment with advanced oxidation processes. In this case, toxicity determination is of crucial importance since the established degradation products may be more toxic than the starting substances. Studies have shown that none of the tested solutions of azithromycin after the application of advanced oxidation processes is toxic, regardless of the presence of formed degradation products. The formation of degradation products was recorded during the photolytic process with TiO2 at a wavelength of radiation of 254 nm.

Azitromicin je vrlo brzo nakon dolaska u široku kliničku uporabu našao istaknuto mjesto u liječenju različitih bakterijskih infekcija, osobito u respiratornom sustavu. S vremenom je rastao broj istraživanja koja su utvrdila važne imunomodulatorne učinke makrolida, što je znatno proširilo indikacijsko područje azitromicina, makrolida koji u odnosu prema drugim makrolidima ima brojne komparativne prednosti. Za razliku od kratkotrajnog propisivanja azitromicina kao antimikrobnog agensa, njegovo se imunomodulatorno djelovanje očituje tek tijekom dugotrajne, višemjesečne kontinuirane primjene. Ovaj tekst kratak je osvrt na primjenu azitromicina u kroničnim respiratornim bolestima u kojima se djelovanje tog lijeka temelji upravo na njegovim imunomodulatornim učincima., Soon after it was introduced in everyday clinical practice, azithromycin became widely used for the treatment of a number of bacterial infections, especially those affecting the respiratory tract. Over time, a growing body of research found that macrolides exert significant immunomodulatory effects, thus significantly expanding indications for azithromycin, a macrolide antibiotic with numerous advantages over the other antibiotics from its class. As opposed to short-term use of azithromycin as an antimicrobial agent, its immunomodulatory effect is presented only after long-term use over a course of several months. This paper briefly addresses the use of azithromycin in patients with chronic respiratory diseases where the effects of the drug are based on its immunomodulatory effects.

Uz već dokazane indikacije, azitromicin se može primijeniti u velikom broju neuobičajenih bolesti kod kojih do izražaja dolaze dva mehanizma djelovanja: klasična antibakterijska aktivnost i imunomodulatorno djelovanje. Ekstenzivna pretraga baza podataka i analiza literature s naglaskom na kliničkim ispitivanjima azitromicina izvan uobičajenih indikacija upućuju na djelotvornost ovog lijeka – ne samo u liječenju cijelog spektra bakterijskih bolesti nego i parazitoza te stanja bez dokazane infektivne etiologije. Vodimo li računa o takvim indikacijama, povećava se mogućnost uspješnog liječenja pacijenata kod kojih drugi modaliteti liječenja nisu pokazali zadovoljavajući učinak., Alongside already established medical indications, azithromycin can be used in a plethora of uncommon diseases with two salient mechanisms of action: classical antibacterial activity and immunomodulatory action. Extensive database search and literature analysis, with an emphasis on clinical trials where azithromycin was used beyond the classical indications, reveal the efficacy of this drug – not only in the treatment of the entire spectrum of bacterial diseases, but also for certain parasitoses and conditions without proven infectious etiology. Taking into account such indications increases the possibility of successfully managing patients when other treatment modalities fail to show a satisfactory effect.

Unatoč rastućoj rezistenciji na makrolide među najčešćim respiratornim patogenima, osobito u zemljama s dugotrajnom primjenom makrolida, azitromicin ima važnu ulogu u liječenju čestih dječjih infekcija. Respiratorne infekcije, ponajprije upala pluća iz opće populacije, posebice među djecom starijom od 5 godina, kao i terapijska kombinacija za liječenje pneumonije u mlađe djece ako postoji sumnja na miješanu infekciju, odnosno teži / komplicirani klinički tijek bolesti, ostaju važno indikacijsko područje za azitromicin. Također, dokazi o ulozi atipičnih bakterija (Mycoplasma pneumoniae i Chlamydia pneumoniae) u etiologiji akutnoga tonzilofaringitisa i astme, kao i kontinuirana prisutnost hripavca u populaciji ističu jasne indikacije za primjenu makrolida / azalida u pedijatrijskoj medicini. Liječenje streptokoknog tonzilofaringitisa u Hrvatskoj, u kojoj rezistencija Streptococcusa pyogenesa na makrolide iznosi oko 7%, također ostaje indikacija za primjenu makrolida / azalida, osobito u slučaju alergije na beta-laktamske antibiotike i u slučajevima kada se liječenje može otpočeti nakon prijma bakteriološkog nalaza obriska ždrijela. Azitromicin ima nedvojbeno mjesto i u liječenju nerespiratornih infekcija poput bolesti mačjeg ogreba (infekcije uzrokovane Bartonellom henselae), antimikrobnog liječenja bakterijskog proljeva i proljeva putnika, solitarnoga migratornog eritema (lajmske borelioze), nekih neonatalnih infekcija (klamidijskog konjunktivitisa i pneumonije), kao i nekih drugih infekcija (infekcije kože i mekih česti (impetigo, erizipel, celulitis, folikulitis, paronihija), kriptosporidioza, profilaksa meningokokne bolesti, egzacerbacije kod cistične fibroze)., Despite the increasing macrolide resistance among the most common respiratory pathogens, especially in countries with a long-term use of macrolides, azithromycin plays an important role in the treatment of frequent infections in children. Respiratory infections, primarily community-acquired pneumonia, among children above the age of 5 in particular, as well as therapeutic combination for the treatment of pneumonia in younger children with suspected mixed infection or severe/complicated course of clinical disease, remain an important indication area for azithromycin. Moreover, evidence of the role of atypical bacteria (Mycoplasma pneumoniae and Chlamydia pneumoniae) in the aetiology of acute tonsillopharyngitis and asthma, as well as the continuous presence of whooping cough in population, point out clear indications for the use of macrolides/azalides in paediatric medicine. The treatment of streptococcal tonsillopharyngitis in Croatia, where macrolide resistance in Streptococcus pyogenes is about 7%, also remains an indication for the administration of macrolides/azalides, particularly in the case of beta-lactame antibiotic allergy and in the cases where treatment can be initiated following the receipt of bacteriological findings for throat swab. Azithromycin should undoubtedly be used in the treatment of non-respiratory infections such as cat-scratch disease (infections caused by Bartonella henselae), antimicrobial therapy of bacterial diarrhoea and traveller's diarrhoea, solitary erythema migrans (Lyme borreliosis), some neonatal infections (chlamydial conjunctivitis and pneumonia), as well as some other infections (skin and soft tissue infections such as impetigo, erysipelas, cellulitis, folliculitis and paronychia, and cryptosporidiosis, meningococcal prophylaxis, cystic fibrosis exacerbations).

Zbog jedinstvene farmakokinetike, odnosno kliničke učinkovitosti, jednostavnog doziranja i dobre podnošljivosti azitromicin je dvadesetak godina imao status izuzetno poželjnog antibiotika, posebno u liječenju akutnih respiratornih infekcija. Pojavom sve učestalije rezistencije pneumokoka na penicilin i makrolide taj je primat sada potisnut, no i danas je azitromicin vrlo važan i nezaobilazan antibiotik. Zbog neučinkovitosti beta-laktamskih antibiotika azitromicin je ostao antibiotikom prvog izbora u liječenju atipičnih pneumonija i hripavca. Također ima vrlo važnu ulogu u liječenju težih oblika pneumonija u kombinaciji s beta-laktamskim antibioticima. U liječenju bakterijskih infekcija gornjeg dijela dišnog sustava (faringitis, sinusitis, otitis media) te akutnih egzacerbacija kronične opstruktivne plućne bolesti azitromicin je alternativni izbor pri alergiji na betalaktamske antibiotike. Nova saznanja o imunomodulaciji i protuupalnom ucinku makrolida pozicionirala su uporabu azitromicina u cističnoj fibrozi i drugim kroničnim bolestima dišnog sustava., For over twenty years, due to its unique pharmacokinetic profile, i. e. clinical efficacy, simple dosing and good tolerability, azithromycin was the preferred antibiotic, especially for the treatment of acute respiratory infections. As a result of an increasing trend in pneumococcal resistance to penicillin and macrolides, azithromycin has lost this prime position, but it still remains an important and indispensable antibiotic. Due to the inefficacy of beta-lactam antibiotics, azithromycin remained the first-line antibiotic for the treatment of atypical pneumonia and pertussis. Furthermore, combined with beta-lactam antibiotics, it plays a major role in the treatment of severe cases of pneumonia. Azithromycin is an alternative choice for the treatment of upper respiratory tract infections (pharyngitis, sinusitis, and otitis media) and acute COPD exacerbations in patients allergic to beta-lactam antibiotics. New discoveries regarding immunomodulatory and anti-inflammatory effect of macrolides have led to the use of azithromycin in patients with cystic fibrosis and other chronic respiratory diseases.

Urogenitalne infekcije i danas su važan javnozdravstveni problem. Opterećenje su ne samo za pojedinca već i za cijelu populaciju i sav zdravstveni sustav. Azitromicin sa svojim jedinstvenim farmakokinetskim i farmakodinamskim osobinama lijek je izbora u liječenju brojnih spolno prenosivih i urogenitalnih infekcija. Potvrdile su to brojne kliničke studije, kao i učinkovitost azitromicina i 30 godina nakon prve registracije. Kronični klamidijski prostatitis nova je klinička indikacija za Sumamed. Ovaj pregledni rad daje preporuke za primjenu azitromicina u različitim urogenitalnim infekcijama, a važnost tog lijeka potvrđena je njegovim mjestom u vodećim svjetskim smjernicama., Urogenital infections are still an important public health concern. They represent a burden not only for an individual but also for the entire population and the complete health system. With its unique pharmacokinetic and pharmacodynamic properties, Azithromycin represents a drug of choice for the treatment of numerous sexually transmitted and urogenital infections. This has been corroborated by a number of clinical trials, as well as by the efficacy that azithromycin has demonstrated since its first registration 30 years ago. Chronic chlamydial prostatitis is a new clinical indication for Sumamed. This overview provides recommendations for the use of azithromycin in various urogenital infections. The inclusion of azithromycin in the world's leading guidelines testifies to its importance.

Azitromicin je antibiotik široka spektra djelovanja iz skupine makrolidnih antibiotika i ima snažno imunomodulatorno djelovanje. Bez obzira na zasad malen broj radova o ovom iznimno važnom svojstvu azitromicina, iz dostupne se literature lako razaznaje njegov potencijal imunomodulatornog djelovanja na razini stanica i tkiva te u različitim kliničkim entitetima, ponajprije u patologiji različitih akutnih i kroničnih respiratornih poremećaja i infekcija. Azitromicin se nakuplja u nizu stanica i djeluje imunomodulatorno na njih preko različitih imunosnih mehanizama (epitelne stanice, fibroblasti, monociti/makrofagi, dendritičke stanice, NK-stanice, T-limfociti, neutrofili, mastociti, eozinofili). Ograničen broj kliničkih studija upućuje na imunomodulatorno djelovanje azitromicina u različitim infekcijama, u prvom redu respiratornim, a onda i infekcijama kože i potkožnog tkiva, spolno prenosivim bolestima i infekcijama te bolestima gastrointestinalnog trakta. Većina njih u svojim se zaključcima dijelom oslanja na različite kalkulacije o imunomodulatornom djelovanju azitromicina, bazirane na studijama in vitro ili ex vivo. Učinkovito antimikrobno djelovanje azitromicina na brojne mikroorganizme, udruženo s njegovim potentnim imunomodulatornim djelovanjem, čini ovaj lijek snažnim terapeutikom ne samo u akutnim nego i u kroničnim infekcijama, ali i u nekim kliničkim entitetima i kroničnim bolestima za koje nije dokazan infektivni uzročnik., Azithromycin is a broad-spectrum antibiotic from a group of macrolide antibiotics that has a strong immunomodulatory effect. Relatively few papers focusing on this extremely important property of azithromycin have been published so far. Nevertheless, available literature reveals its enormous potential for immunomodulatory effect at the cell and tissue level and in various clinical entities, primarily in the pathology of various acute and chronic respiratory disorders and infections. Accumulating within a whole string of cells, azithromycin uses different immune mechanisms (epithelial cells, fibroblasts, monocytes/macrophages, dendritic cells, NK cells, T lymphocytes, neutrophils, mast cells, eosinophils) to exert its immunomodulatory effect. A limited number of clinical studies indicate the immunomodulatory effect of azithromycin in various infections, primarily respiratory infections, as well as in skin and subcutaneous tissue infections, sexually transmitted diseases and gastrointestinal infections and diseases. In most of these studies, conclusions regarding the immunomodulatory effect of azithromycin are, in part, based on various calculations supported by in vitro or ex vivo studies. Given its effective antimicrobial activity against numerous microorganisms, together with its potent immunomodulatory effect, azithromycin is an effective therapeutic drug indicated for the treatment of both acute and chronic infections, as well as certain clinical entities and chronic diseases for which no infectious agent has been proven.

Skupina istraživača-znanstvenika iz PLIVE stvorila je 1981. godine novi antibiotik pod nazivom azitromicin, po svemu različit od ostalih do tada primjenjivanih antibiotika. U rujnu 1988. godine registriran je PLIVIN originalni antibiotik, azitromicin, poznatiji pod zaštićenim imenom Sumamed. Osobine azitromicina od samih početaka upućivale su na to da se radi po svemu jedinstvenom antibiotiku koji ima izvrsnu učinkovitost na najčešće uzročnike infekcija, ciljano djelovanje na mjestu infekcije, vrlo se jednostavno i kratko primjenjuje i ima odličnu podnošljivost. Iz farmakoloških osobina i kliničkih istraživanja proizišle su i originalne marketinške kampanje koje su mijenjale paradigmu razumijevanja antimikrobnog učinka i djelovanja na mjestu infekcije, kao i način uzimanja antimikrobnih lijekova. Marketinške kampanje odnosile su se na jednostavnost i kratkoću primjene lijeka te izvrsnu učinkovitost i podnošljivost Sumameda. U radu su prikazani počeci promocije Sumameda., In 1981 a group of researchers at Pliva created azithromycin, a new antibiotic completely different from the antibiotics in use at the time. In September 1988 PLIVA registered azithromycin as its original antibiotic under the brand name Sumamed. From the very beginning the properties of azithromycin indicated that this was a unique antibiotic with excellent efficacy against the most common infectious agents, targeted action at the site of infection, simple dosing, short-term use and excellent tolerability. Pharmacological characteristics and results of clinical trials were used in the original advertising campaigns which resulted in a paradigm shift in the understanding of antimicrobial activity and action at the site of infection, as well as of administration of antimicrobial drugs. Advertising campaigns referred to the simple and short-term use as well as excellent efficacy and tolerability of Sumamed. The paper focuses on the beginnings of promotion of Sumamed.

Bez obzira na to što je rezistencija bakterija na antibiotike (antimikrobna otpornost) jedan od glavnih izazova moderne medicine, klamidijska je infekcija dosad pokazivala dobar klinički odgovor na lijek prvog izbora – azitromicin. Ipak, u literaturi postoje ponovljeni opisi laboratorijski dokazane rezistencije spolno prenosive bakterije Chlamydia trachomatis (C. trachomatis) koja uzrokuje ovu infekciju. Osim toga, Hrvatska je zemlja s visokom stopom propisivanja azitromicina i drugih antimikrobnih lijekova, što može utjecati na pojavu otpornosti ili slabijeg odgovora na terapiju. U skladu s time cilj prvoga hrvatskog istraživanja antimikrobnog profila C. trachomatis bio je utvrditi djelotvornost azitromicina i drugih antibiotika prema urogenitalnim kliničkim izolatima C. trachomatis u laboratorijskim uvjetima. U staničnome modelu McCoyevih stanica određivane su minimalna inhibitorna koncentracija (MIC) i minimalna klamidicidna koncentracija (MCC), a svi ispitivani sojevi bili su osjetljivi na azitromicin (MIC < 4 μg/ml). Premda rezistentni sojevi bakterije nisu nađeni, kod određenih izolata uočene su znatno više vrijednosti MCC-a za azitromicin, što pokazuje trend moguće smanjene osjetljivosti. Bez obzira na zahtjevnu metodologiju, potrebno je nastaviti s ovakvim istraživanjima, poglavito u zemljama s velikim opterećenjem propisivanja antibiotika., Regardless of the fact that antimicrobial resistance represents one of the most pressing issues of modern medicine, thus far chlamydial infection responded well to the first-line therapy i.e. azithromycin. Nonetheless, laboratory-confirmed resistance of sexually-transmitted pathogen Chlamydia trachomatis (C. trachomatis) causing this infection has been repeatedly described in the literature; furthermore, Croatia belongs to the group of countries that exhibit high prescribing rates of azithromycin and other antimicrobial drugs, which can, in turn, give rise to frank resistance or decreased susceptibility. Accordingly, the first Croatian antibiotic susceptibility study of Chlamydia trachomatis aimed to assess in vitro effectiveness of azithromycin and other antibiotics against clinical chlamydial isolates from genitourinary tract. McCoy cell culture model was used to assess minimal inhibitory concentration (MIC) and minimal chlamydicidal concentration (MCC), and all the tested isolates were susceptible to azithromycin (MIC < 4 μg/ml). Although no resistant strains have been found, certain isolates showed significantly higher values of MCC for azithromycin, pointing towards the trend of possible decreased susceptibility. Regardless of the complex methodology, it is pivotal to continue with this kind of resistance surveillance, especially in countries with high antibiotic prescribing rate.

Svrha ovog rada bila je provesti fizikalnu karakterizaciju različitih tipova liposoma s uklopljenim lipofilnim ili hidrofilnim lijekom (azitromicin ili diltiazemklorid). Poseban naglasak stavljen je na ispitivanje membranske elastičnosti budući da su formulacije namijenjene primjeni na kožu i sluznicu. Ispitivanjem se željelo utvrditi kako fosfolipidni sastav, prisustvo surfaktanta (natrijev deoksikolat) ili suotapala (propilenglikol) u liposomima utječe na fizikalna svojstva liposoma: veličinu, naboj na površini te elastičnost/rigidnost njihovih dvoslojeva. Liposomi s azitromicinom pripravljeni su film metodom, dok su liposomi s diltiazemkloridom pripremljeni proliposomskom i metodom razrjeđenja poliola. Pokazano je da metoda priprave i fosfolipidni sastav značajno doprinose fizikalnim svojstvima liposoma. Film metodom dobivene su izrazito heterogene disperzije liposoma, a provedena ekstruzija rezultirala je liposomima značajno manjeg srednjeg promjera i niskog indeksa polidisperznosti. Ispitivanja membranske elastičnosti pokazala su najveću elastičnost deformabilnih liposoma s azitromicinom. Elastičnost propilenglikol liposoma s azitromicinom je bila manja od deformabilnih liposoma, ali značajno veća od anionskih, konvencionalnih, te pogotovo kationskih liposoma. Potvrđeno je da fosfolipidni sastav liposoma, prisustvo surfaktanta ili suotapala te lipofilnost uklopljene djelatne tvari značajno utječu na stupanj membranske elastičnosti liposoma. The main objective of this study was physical characterization of the different types of liposomes loaded with either a lipophilic or hydrophilic drug (azithromycin or diltiazem chloride). The focus was placed on determination of membrane (liposome bilayer's) elasticity, because the investigated nanoformulations are destined for skin and mucosal drug delivery. The specific aims were to determine influence of the phospholipid composition, surfactant (sodium deoxycholate) or co-solvent (propylene glycol) on liposomal physical characteristics: size, surface charge and bilayer’s elasticity. Liposomes containing azithromycin were prepared by the film method, while liposomes with diltiazem chloride were prepared by proliposome and polyol dilution method. The results have shown that the phospholipid composition and method used for the preparation of liposomes significantly influence physical characteristics of the liposomes. Liposomes prepared by the film method were significantly more heterogenic than liposomes prepared by proliposome and polyol dilution method, respectively. Extrusion of the originally prepared liposomes resulted in the liposomes of significantly smaller mean diameters and polydispersity indexes. Deformable liposomes with azithromycin were characterized with the most elastic bilayers. Propylene glycol liposomes had less elastic bilayers than deformable liposomes, but significantly more elastic bilayers than anionic, conventional and cationic liposomes encapsulating azithromycin. On the contrary, propylene glycol liposomes with diltiazem chloride were more elastic than propylene glycol liposomes encapsulating azithromycin. This study confirmed that phospholipid composition, presence of surfactant and co-solvent, as well as the lipophilicity of the drug have a significant impact on liposome membrane elasticity.

Istraživanja potrošnje antibiotika od osobitog su interesa zbog dokazane povezanosti rezistencije bakterija s njihovom potrošnjom. Cilj ovog rada je istražiti je li došlo do promjene trendova potrošnje antibiotika u RH u odnosu na one prikazane u European Surveillance of Antimicrobial Consumption (ESAC) studiji. Pratila se farmakološka potrošnja antibiotika (J01AJ01X) na temelju podataka iz godišnjih Izvješća o potrošnji lijekova, Hrvatske agencije za lijekove i medicinske proizvode tijekom dva razdoblja, razdoblja trajanja ESAC studije (2005.-2009.) i razdoblja nakon završetka studije (2010.-2014.). U odnosu na ESAC studiju u kojoj je zabilježen stalni rast potrošnje antibiotika, nakon završetka studije potrošnja se stabilizirala na oko 23 defi nirane dnevne doze na 1000 stanovnika na dan. Iako se potrošnja stabilizirala, RH još uvijek spada u skupinu zemalja s većom potrošnjom antibiotika od sjevernoeuropskih zemalja, osobito s nepovoljnom strukturom potrošnje. Dok je u našoj studiji udio uskog spektra penicilina iznosio manje od 10 %, u nordijskim zemljama njegov udio je prelazio 50 % ukupne penicilinske potrošnje. Kod nas je zabilježena osobito velika potrošnja amoksicilina s klavulonskom kiselinom i azitromicina, pojava koja nije prisutna u zapadnoeuropskim zemljama. Iako se već primjenjuju mjere smanjenja potrošnje antibiotika u RH, dobiveni rezultati ukazuju na potrebu još sustavnijeg pristupa problemu njihove prekomjerne potrošnje., Introduction: Antibiotic resistance is a very important public health problem throughout the world, which is undoubtedly connected with the excessive use of antibiotics. According to monitoring of antibiotic consumption in Europe, Croatia as part of the European Surveillance of Antimicrobial Consumption (ESAC) project was among countries with stable high outpatient antibiotic consumption from 1997 to 2009 and consequently high rates of resistance to many antibiotics. Research on the use of antibiotics is especially important for family medicine; more than 90% of antibiotics were prescribed outside the hospital, mostly by family physicians (general practitioners). The main aim of this study was to investigate ten-year (2005-2014) trends in antibiotic consumption in Croatia divided into two periods, i.e. during (2005-2009) and after (2010-2014) ESAC study, and to compare the data obtained to those from other EU countries. Methods: The study was observational, based on routinely collected data from annual reports of the Croatian Agency for Medicinal Products and Medical Devices (HALMED) for the 2005-2014 period. Drug consumption in the reports was presented according to ATC/ DDD methodology and antibiotics were presented under the J code. Results: The results showed an increasing trend of antibiotic consumption until 2009, followed by a stable trend from 2010, with 22-23 daily doses per 1000 inhabitants per day. In comparison to other EU countries, Croatia is a country with high antibiotic consumption. Similar to other countries, penicillins were the most widely used antibiotics, accounting for 48%-55% of total pharmacological consumption. Cephalosporins ranked second with 34%-22%, showing a downward trend of consumption. Macrolides with an upward trend were on the third place and quinolones on the fourth place with a variable trend. However, the structure of consumption was not comparable to other EU countries. For instance, co-amoxiclav accounted for 53%-70% of the penicillin group, with an upward trend, and azithromycin for about 50% of macrolide consumption, with a varying trend. The increasing trend of consumption of third-generation cephalosporins and quinolones also differed from other EU countries. Consumption of ciprofl oxacin showed an upward trend, and norfl oxacin a downward trend. Conclusion: Although the trend has been stable since 2010, the consumption of antibiotics in Croatia is still higher than in many European countries. Special attention should be paid to the overuse of co-amoxiclav and azithromycin. These data are the reasons to think about introducing systematic interventions aimed at reducing the use of antibiotics.

Svrha ovog rada je utvrditi potencijal tretmana hladnom plazmom za uklanjanje azitromicina i atrazina iz modelne vode, te utjecaj izvedbe reaktora i pH vrijednosti na uĉinkovitost tretmana hladnom plazmom. Otopina azitromicina i atrazina u mineralnoj vodi tretirana je u ĉetiri razliĉita reaktora pri pH 3, 7 i 10. Tretman hladnom plazmom pokazao se djelotvornim za uklanjanje azitromicina i atrazina iz modelne vode. Najuĉinkovitija razgradnja azitromicina od 98 % hladnom plazmom iz modelne vode postignuta je nakon 1 h u reaktoru s JET plazmom pri pH 10, a najuĉinkovitija razgradnja atrazina hladnom plazmom iz modelne vode, 55 % postignuta je nakon 20 min u reaktoru s plinskom plazmom uz miješanje peristaltiĉkom pumpom i upuhivanje argona pri pH 3. The purpose of this study is to determine the potential of cold plasma treatment for removal of azithromycin and atrazine from water, as well as the impact of the reactor design and the pH value on the effectiveness of cold plasma treatment. A solution of azithromycin and atrazine in mineral water was treated in four different plasma reactors, at three different pH values, pH 3, 7 and 10. Treatment with cold plasma proved to be effective for the removal of azithromycin and atrazine in water. The most effective degradation of azithromycin, 98 % was achieved after 1 h with JET plasma at pH 10, while the most effective degradation of atrazine, 55 % was achieved after 20 min treatment with gas plasma with blowing argon into reactor at pH 3.

Cilj ovog rada bio je ispitati oslobađanje azitromicina iz kationskih liposoma uklopljenih u kitozanski gel. U tu svrhu pripravljene su film metodom dvije preparacije kationskih liposoma; DPPC/DODAB liposomi te SPC liposomi obloženi kitozanom, a za usporedu su poslužili negativno nabijeni SPC liposomi. Sve su formulacije karakterizirane u pogledu srednjeg promjena, indeksa polidisperznosti i zeta potencijala, prije i nakon homogenizacije disperzija korištenjem ekstrudera i ultrazvučne sonde. Nakon odijeljivanja neuklopljenog lijeka liposomi su umiješani u kitozanski gel pripravljen iz kitozana velike molekulske mase. Koncentracija liposoma u gelu bila je 30% (w/w). Srednji promjer liposoma, nakon provedene homogenizacije, kretao se između 130 (SPC liposomi) i 250 nm (DPPC/DODAB); jedino su oni obloženi kitozanom bili su nešto veći zbog specifičnosti postupka izrade. Zeta potencijali kationskih liposoma bili su veći od +30 mV, dok je za SPC liposome zeta potencijal bio manji od -30 mV, što ukazuje na fizičku stabilnost svih pripravljenih disperzija. Rezultati in vitro ispitivanja oslobađanja azitromicina iz liposoma-u-gelu ukazuju na brzi trend oslobađanja lijeka u prva 4 sata ispitivanja i polaganiji tijekom idućih 20 sati. Nije uočena statistički značajna razlika u profilu oslobađanja lijeka između ispitivanih kationskih i anionske formulacije liposoma. Usporedba s kontrolnim gelom (sadrži otopinu azitromicina umjesto liposoma), ukazuje na polaganiji trend oslobađanja azitromicina iz svih liposomskih pripravaka. The aim of this study was to examine the release profile of azithromycin from liposomes embedded in chitosan hydrogel. For that purpose, two type of cationic liposomes were prepared by film hydration method: DPPC/DODAB liposomes and SPC liposomes coated with 0,3% (w/w) chitosan solution. Negatively charged SPC liposomes served for comparison. All liposomal preparations were physically characterized before and after homogenization by measuring mean diameter, size distribution and zeta potential. Homogenization of different liposome dispersions was preformed using an ultrasonic probe sonicator or extruder. After separation of unencapsulated drug, the liposomes were mixed into a gel prepared from high-molecular weight chitosan. The concentration of liposomes in the gel was 30% (w/w). After homogenization, the mean diameter ranged between 130 nm (SPC liposomes) and 250 nm (DPPC/DODAB). Liposomes coated with chitosan were slightly higher due to the specificities of the manufacturing process. Zeta potentials of the cationic liposomes were higher than +30 mV, while the zeta potential of the SPC liposomes was less than -30 mV, indicating the physical stability of all the prepared liposome dispersions. In vitro release results indicate a trend of rapid drug release in the first 4 hours and slower in the next 20 hours. There were no statistically significant differences in the drug release from all the tested cationic and anionic liposomes. When compared to control (gel containing solution of azithromycin instead of the liposomes), slower release of azithromycin from all the tested liposome formulations was proven.

Cilj ovog rada bio je pripraviti kationske liposome s uklopljenim azitromicinom, provesti njihovu fizikalnu karakterizaciju te ispitati profil oslobađanja uklopljenog lijeka. Kationski liposomi (DOPC/DODAB; DPPC/DODAB; DPPC/DC-Chol) s uklopljenim azitromicinom su pripravljeni metodom hidratacije suhog fosfolipidnog sloja (filma). Homogenizacija liposomskih disperzija je provedena primjenom ultrazvučnog sonikatora sa sondom frekvencije 40 Hz tijekom 20 sekundi. Nakon soniciranja dobiveni su liposomi srednjeg promjera oko 300 nm, uz znatnu bolju razdiobu veličina liposoma (PI u rasponu od 0,360 do 0,471). Svim je preparacijama izmjeren zeta potencijal, a izmjerene pozitivne vrijednosti zeta potencijala u iznosu od +56,7 do +75,2 mV upućuju na fizičke stabilne liposomske disperzije. In vitro ispitivanje oslobađanja azitromicina je provedeno na Franz difuzijskoj ćeliji u trajanju od 24 sata te je količina oslobođenog azitromicina određena HPLC metodom. Za usporedbu su uzeti vodeno-etanolna otopina azitromicina kao kontrola i anionski SPC-liposomi. Nakon 24 sata iz testiranih kationskih liposoma se oslobodila približno ista količina (%) azitromicina. Iz ranije dobivenih podataka za anionske liposome, vidljivo je da se oslobodilo znatno manje azitromicina, ali je kod njih postignuto kontrolirano i produljeno oslobađanje, za razliku od kationskih gdje je takva vrsta oslobađanja postignuta samo djelomično. Može se zaključiti da je potrebno provesti dodatna in vitro ispitivanja oslobađanja kako bi se optimizirali uvjeti za postizanje potpunog kontroliranog i produženog oslobađanja kod kationskih liposoma. The aim of this study was to prepare cationic liposomes with encapsulated azithromycin, perform physical characterization and determine the drug release profile. Cationic liposomes (DOPC/DODAB; DPPC/DODAB; DPPC/DC-Chol) with encapsulated azithromycin were prepared by the film hydration method. Homogenization of liposome dispersions was performed using an ultrasonic probe sonicator with a frequency of 40 Hz for 20 seconds. After sonication, the resulting liposomes were with a mean diameter of 300 nm and with much better size distribution (PI ranging from 0.360 to 0.471). The measured positive value of zeta potential in a range of +56.7 to +75.2 mV indicated the formation of physically stable liposomal dispersions. In vitro release studies were performed on the Franz diffusion cell during 24 hours and the amount of the released azithromycin was determined by HPLC method. For comparison, aqueous-ethanolic solution of azithromycin as the control and the anionic-SPC liposomes were prepared and tested under the same conditions. After 24 hours, all of the tested cationic liposomes released approximately the same amount (%) of azithromycin. From the previously obtained data for anionic liposomes, it is evident that they have released significantly less amount of azithromycin, but they have achieved a controlled and prolonged release, as opposed to cationic liposomes where such release is achieved only partially. It could be concluded that it is necessary to do additional in vitro release testing in order to optimize the conditions for achieving complete controlled and prolonged release of the drug from cationic liposomes.

Cilj studije bio je opisati kliničku učinkovitost azitromicina u liječenju infekcija gornjeg dijela dišnog sustava, ispitati duljinu simptoma nakon početka terapije i pokazati moguće nuspojave liječenja azitromicinom. U studiju je uključeno 422 ispitanika: 184 s dijagnozom akutnog faringitisa, 156 s dijagnozom akutnog sinuitisa, te 82 bolesnika s akutnom upalom srednjeg uha. Klinička učinkovitost nakon tri dana liječenja azitromicinom temeljila se na poboljšanju kliničke slike i izlječenju bolesnika. Za svaku dijegnozu je izračunat klinički zbroj prije početka liječenja, te četvrti i dvanaesti dan nakon početka liječenja. Studija je pokazala kako je trodnevno liječenje azitromicinom dovelo do smanjenja simptoma u 93,6% bolesnika. Godinu dana prije uključenja u studiju antibiotici su bili prethodno propisivani u 56,6% bolesnika, a 20,5% bolesnika je bilo liječeno makrolidima. Klinička učinkovitost je bila pozitivna u 95,6% bolesnika, a neuspjeh liječenja zabilježen je u 4,4% bolesnika. Prijavljeno je 48 nuspojava kod 38 (9,1%) bolesnika. Najčešćee nuspojave bile su: bol u trbuhu zabilježena kod 14 (1,1%) bolesnika, proljev kod 12 (0,9%), mučnina kod 5 (0,4%), te povraćanje kod 5 (0,4%) bolesnika. Prekid terapije zbog nuspojava zabilježen je kod jednog bolesnika. Rezultati studije pokazuju da u liječenju infekcija gornjeg dišnog sustava azitromicinom ima visoku učinkovitost i mali broj nuspojava. Azitromicin je pouzdan lijek izbora u liječenju infekcija gornjeg dijela dišnog sustava, s brzim povlačenjem simptoma infektivne bolesti i relativno malim brojem nuspojava., The aim of the study was to describe clinical effectiveness of azithromycin in the management of upper respiratory tract infections, to examine the duration of symptoms after beginning of therapy and to mark possible adverse events of azithromycin treatment. The overall intention-to-treat (ITT) study population included 422 patients: 184 with diagnosis of acute pharyngitis (AP), 156 with acute sinusitis (AS) and 82 with acute otitis media (AOM). Clinical effectiveness was based on the results of clinical improvement and cure after three day treatment with azithromycin, calculating the clinical score for each diagnosis before treatment (at the inclusion), at the 4th day (end of the therapy) and at the 12th (end of the study). In this study azithromycin led to the relief of symptoms after three days in the 93.6% of patients. Antibiotics were prescribed within one year before inclusion in 56.6% of patients and 20.5% were previously treated with macrolides. Clinical effectiveness in ITT population was 95.6% and there were 4.4% failures. Overall, 38 (9.1%) patients reported 48 adverse events. The most common were: abdominal pain reported in 14 (1.1%) patients, diarrhea in 12 (0.9%), nausea in 5 (0.4%), vomiting in 5 (0.4%). The therapy was withdrawn because of an adverse event in one patient. The results of our study show that azithromycin in the treatment of upper respiratory tract infections has a high clinical effectiveness and small number of adverse events. Azithromycin is a reliable antibiotic treatment in upper respiratory tract infections, giving fast resolution of symptoms with little adverse events in patients with presumed bacterial infections.

Akutne respiratorne infekcije najčešće su infektivne bolesti i razlog najčešćih posjeta ordinacijama opće prakse. Za azitromicin je poznato da ima širok spektar djelovanja na većinu uzročnika pneumonija uključujući gram-pozitivne, gram-negativne i atipične mikroorganizme s dobrom kliničkom učinkovitosti pri peroralnoj primjeni, što omogućava ambulantno liječenje bolesnika mlađih od 65 godina bez kroničnih bolesti. Danas je poznato da azitromicin osim antimikrobnog ima i znatno imunomodulatorno i u okviru toga protuupalno djelovanje. Ovdje ce biti prikazano u prvom redu potencijalno protuupalno djelovanje azitromicina u infekcijama respiratornog trakta., Acute respiratory infections are the most common infectious diseases and the most common reason for visiting a general practitioner clinic. Azithromycin is known to have a broad spectrum of action against most causes of pneumonia, including Gram-positive, Gram-negative and atypical microorganisms. Its clinical efficacy in oral administration is also good, which allows outpatient treatment of patients below 65 years of age and without chronic diseases. Today we know that in addition to antimicrobial, azithromycin also has significant immunomodulatory and anti-inflammatory properties. This article primarily focuses on its potential anti-inflammatory effects in respiratory tract infections.

Povrh poznatog bakteriostatskog učinka makrolidi, uključujući azitromicin, mogu ublažiti kroničnu upalu i modulirati imunosni odgovor domaćina, a da ne utječu na njegovu sposobnost obrane od infekcija. Ovakva imunomodulacijska svojstva makrolida sve se više rabe u liječenju kroničnih upalnih bolesti dišnih putova u djece i odraslih. Danas je dugotrajna niskodozna primjena azitromicina široko prihvaćena i preporučuje se u liječenju cistične fibroze i bronhiektazija. Makrolidi se ispituju i rabe u liječenju kronične neutrofilne upale dišnih putova koja se često viđa u bolesnika s astmom rezistentnom na kortikosteroide i kroničnom opstruktivnom plućnom bolesti. Međutim potrebna su daljnja klinička ispitivanja koja trebaju prepoznati grupe bolesnika za koje je izgledno da će reagirati na takvo imunomodulacijsko liječenje., Besides their known bacteriostatic activity, macrolides, including azithromycin, can reduce chronic inflammation and modulate immunity without impairing host response against infection. These immunomodulatory properties of macrolides have been increasingly used to treat chronic inflammatory airway diseases in children and adults. Low-dose, long-term therapy with azithromycin is now recommended and widely adopted for the treatment of cystic fibrosis and bronchiectasis. Macrolides are also being investigated and used to treat chronic neutrophilic airway infl ammation, often seen in patients with steroid-resistant asthma and chronic obstructive pulmonary disease. However, further clinical trials are required to identify groups of patients most likely to respond to such immunomodulatory therapy.

Povrh poznatog bakteriostatskog učinka makrolidi, uključujući azitromicin, mogu ublažiti kroničnu upalu i modulirati imunosni odgovor domaćina, a da ne utječu na njegovu sposobnost obrane od infekcija. Ovakva imunomodulacijska svojstva makrolida sve se više rabe u liječenju kroničnih upalnih bolesti dišnih putova u djece i odraslih. Danas je dugotrajna niskodozna primjena azitromicina široko prihvaćena i preporučuje se u liječenju cistične fibroze i bronhiektazija. Makrolidi se ispituju i rabe u liječenju kronične neutrofilne upale dišnih putova koja se često viđa u bolesnika s astmom rezistentnom na kortikosteroide i kroničnom opstruktivnom plućnom bolesti. Međutim potrebna su daljnja klinička ispitivanja koja trebaju prepoznati grupe bolesnika za koje je izgledno da će reagirati na takvo imunomodulacijsko liječenje., Besides their known bacteriostatic activity, macrolides, including azithromycin, can reduce chronic inflammation and modulate immunity without impairing host response against infection. These immunomodulatory properties of macrolides have been increasingly used to treat chronic inflammatory airway diseases in children and adults. Low-dose, long-term therapy with azithromycin is now recommended and widely adopted for the treatment of cystic fibrosis and bronchiectasis. Macrolides are also being investigated and used to treat chronic neutrophilic airway infl ammation, often seen in patients with steroid-resistant asthma and chronic obstructive pulmonary disease. However, further clinical trials are required to identify groups of patients most likely to respond to such immunomodulatory therapy.

Antimikrobno liječenje indicirano je u bolesnika akutnim bakterijskim prostatitisom, kroničnim bakterijskim prostatitisom, upalnim oblikom sindroma kronične zdjelične boli bez dokažljive infekcije kojih su dio i bolesnici s bakterijskim prostatitisom u kojih uzročnik nije dokazan klasičnim mikrobiološkim metodama, te u odabranih bolesnika s neupalnim oblikom sindroma kronične zdjelične boli bez dokažljive infekcije i s asimptomatskim upalnim prostatitisom. Azitromicin je lijek prvog izbora u liječenju prostatitisa uzrokovanog Chlamydijom trachomatis i Ureaplasmom urealyticum. Preporučuje se u ukupnoj dozi od 4,0 do 4,5 g primijenjen intermitentno tijekom 3-4 tjedna. Kada se kronični bakterijski prostatitis očituje težim kliničkim tijekom, preporučuje se kombinirana antimikrobna terapija ciprofloksacinom i azitromicinom koji se tada primjenjuje u pulsnoj terapiji i u ukupnoj dozi od 4,5 do 6,0 grama. U tih bolesnika azitromicin je indiciran zbog svoje protuupalne i imunomodulatorne aktivnosti te djelovanja na bakterijski biofilm., Antimicrobial treatment is indicated in patients with acute bacterial prostatitis, chronic bacterial prostatitis, inflammatory symptoms of chronic pelvic pain syndrome without detectable infection (which also includes patients with bacterial prostatitis in whom the causative pathogen has not been proven by classical microbiological methods), and in selected patients with non-inflammatory chronic pelvic pain syndrome without detectable infection and patients with asymptomatic inflammatory prostatitis. Azithromycin is the drug of choice for the treatment of prostatitis caused by C. trachomatis and U. urealyticum. It is recommended in a total dose of 4.0 to 4.5 grams intermittently administered over 3-4 weeks. If chronic bacterial prostatitis has a severe clinical course, a combined antimicrobial therapy with ciprofloxacin and azithromycin is recommended, where azithromycin is applied as a pulse therapy in a total dose of 4.5 to 6.0 grams. In these patients azithromycin is administered because of its anti-inflammatory and immunomodulatory activity and its effects on bacterial biofilm.

Mediterranean Spotted Fever (MSF) is a tick-borne rickettsiosis caused by Rickettsia conorii. in the North Dalmatia is an endemic area with high prevalence of R.conorii infected ticks (70.5%) and antirickettsial antibodies among inhabitants (40%). Due to high risk of MSF antibiotic prophylaxis in persons with tick bites seems justified.

Azitromicin (Sumamed®, PLIVA) antibiotik je proizveden u Plivinu Istraživačkom institutu. Zahvaljujući svojim iznimnim svojstvima, postao je danas najprodavaniji antibiotik u svijetu i svakako najpropisivaniji antibiotik za liječenje najčešćih bakterijskih spolno prenosivih i urogenitalnih infekcija, jednokratno u dozi od 1 g po. Lijek je izbora za liječenje akutne infekcije Chlamydijom trachomatis, Ureaplasmom urealyticum i Mycoplasmom genitalium. Kliničkom primjenom azitromicina postignut je u svjetskim razmjerima bitan napredak u liječenju SPI. Danas je međutim jasno da u SPI postoje indikacije u kojima je jednokratna primjena 1 g azitromicina nedostatna te se doze i duljina primjene moraju mijenjati. Tako se u liječenju perzistentne klamidijske infekcije primjenjuje u dozi 1x1 g na tjedan tijekom 3 tjedna. I u liječenju zdjelične upalne bolesti preporučuje se terapija tijekom 3 tjedna, najčešće 1x1 g na tjedan tijekom 3 tjedna ili 1x500 mg u iv. infuziji tijekom 1-2 dana, a zatim 250 mg po. do ukupno 7 dana liječenja. U produljenoj primjeni 1x1 g na tjedan tijekom 3 tjedna primjenjuje se i kod postveneričnoga reaktivnog artritisa i Reiterova sindroma., A zithromycin (Sumamed, PLIVA) is an antibiotic discovered in PLIVA’s Research Institute. Thanks to its exceptional properties, it has become the best-selling antibiotic worldwide and the most prescribed antibiotic for the treatment of the most common bacterial sexually transmitted and urogenital infections. Administered in a single 1 g oral dose, azithromycin is a drug of choice for acute infections due to Chlamydia trachomatis, Ureaplasma urealyticum and Mycoplasma genitalium. At the global level, considerable progress has been made in the treatment of sexually transmitted infections thanks to the clinical use of azithromycin. However, it is clear today that there are some indications where a single 1 g dose is insufficient and that azithromycin strengths and the duration of treatment should be changed. Therefore, azithromycin is administered in a dose of 1 g/week over three weeks in persistent chlamydial infection. A three-week therapy is also recommended in pelvic inflammatory disease, most often 1 g/week over three weeks or 500 mg/day IV over 1 – 2 days, followed by oral 250 mg doses up to 7 days. The long-term administration of 1 g/week over three weeks is also indicated in postvenereal reactive arthritis and Reiter’s syndrome.

Enteroagregacijska Escherichia coli koja proizvodi shiga-toksin (STEC O104:H4) uzrokovala je epidemiju enterokolitisa u Njemačkoj u svibnju 2011. godine. Epidemija je karakterizirana visokom učestalošću hemolitičko-uremijskog sindroma (HUS) koji se javio u više od 20% oboljelih. Antimikrobno liječenje upotrebljava se u liječenju enterokolitisa uzrokovanog enteroagregacijskom Escherichiom coli (E. coli), no nije dopušteno u liječenju infekcija uzrokovanih STEC-om zbog povećane učestalosti HUS-a. Tijekom epidemije u Njemačkoj uzrokovane STEC-om značajan broj bolesnika liječen je azitromicinom. Primjena azitromicina skratila je kliconoštvo i nije utjecala na incidenciju HUS-a., An outbreak of Shiga toxin–producing enteroaggregative Escherichia coli (STEC O104:H4) infection with a high incidence of haemolytic uremic syndrome (HUS) occurred in Germany in May 2011. Antibiotic therapy is widely used to treat enteroaggregative E. coli infection but antibiotic treatment of STEC infection is discouraged because it might increase the risk of HUS development. In the German outbreak, a noticeable number of patients received azithromycin treatment. Treatment with azithromycin was associated with a lower frequency of long-term STEC 0104 H4 carriage, and it did not affect the incidence of HUS.

Kronični bakterijski prostatitis (KBP) još je i danas važan terapijski problem s obzirom na razmjerno nizak postotak izlječenja. Jedan od razloga svakako je i slabo prodiranje većine antimikrobnih lijekova u tkivo i sekrete prostate. Stoga je razumljivo da se posebna pozornost posvetila farmakokinetskim karakteristikama antimikrobnih lijekova koji dolaze u obzir za liječenje ove složene infekcije. Za uspješno liječenje svake bakterijske infekcije, pa tako i KBP-a, važno je identificirati uzročnika i utvrditi njegovu osjetljivost. U KBP-u izolacija je uzročnika problem između ostaloga i zbog još prisutnih dilema koji je biološki uzorak najrelevantniji za mikrobiološku dijagnostiku. Zbog toga postoje ne sasvim razumljive razlike u postotcima izoliranih uzročnika KBP-a među provedenim studijama. Iz toga razloga uspješnost empirijske terapije u KBP-u niža je od željene. Određeni je napredak u liječenju KBP-a postignut upravo na temelju boljeg razumijevanja i uporabe farmakokinetskih karakteristika antimikrobnih lijekova. Primjenom fluoriranih kinolona u liječenju KBP-a izazvanog Escherichijom coli postotak izlječenja s 40% iz doba kada je lijek izbora u liječenju KBP-a bio trimetoprim, povećan je na iznad 60%. Preostaje vidjeti hoće li fluorirani kinoloni 4. generacije zbog bolje farmakokinetike i postizanja visokih koncentracija u prostati pridonijeti daljem poboljšanju liječenja KBP-a. Uzimanjem u obzir farmakokinetskih karakteristika azitromicina unaprijeđeno je i liječenje KBP-a izazvanog Chlamydijom trachomatis. Činjenica da se u znatnog broja oboljelih od KBP-a u liječenju uz antimikrobne lijekove rabe i protuupalni lijekovi, blokatori alfaadrenergičkih receptora upućuje na složenost ovoga kliničkog entiteta. Rezultati kliničkih studija upućuju na potrebu razjašnjenja još uvijek nepoznatih faktora zbog kojih je liječenje KBP-a nedovoljno uspješno unatoč dugotrajnoj primjeni jednog ili više antimikrobnih lijekova., Chronic bacterial prostatitis (CBP) is still an important therapeutic problem due to relatively low treatment success rates. One of the reasons is certainly a poor penetration of most antimicrobial drugs into prostatic tissue and secretions. Special attention given to pharmacokinetic properties of antimicrobial drugs, which could treat these complex infections, is therefore understandable. For a successful treatment of any bacterial infection, including CBP, pathogens and their sensitivity should be identified. The isolation of a pathogen in CPB is still a problem due to an ever present dilemma about the most relevant biological sample for microbiological diagnosis. Therefore, there are some not quite understandable differences in the percentages of CPB pathogens isolated in studies conducted so far. Consequently, the success rates of the empirical CPB therapy are below those desired. Some progress in CBP treatment has been achieved thanks to a better understanding and use of pharmacokinetic properties of antimicrobial drugs. The percentage of successfully treated patients with CBP due to E. coli grew from 40% in those treated with trimethoprim as first-line-treatment to 60% in those treated with fluoroquinolones. It remains to be seen whether 4th generation fluoroquinolones will further improve the CBP treatment due to their enhanced pharmacokinetics and high prostate levels. The treatment of CBP due to C. trachomatis has been also improved thanks to the understanding of the azithromycin pharmacokinetic properties. The fact that anti-inflammatory drugs and alpha-adrenergic blockers are used in a considerable number of CBP patients in addition to antimicrobial drugs indicates the complexity of this clinical entity. The results of clinical studies show that the reasons lying behind the insufficiently successful CBP treatment despite a long-term use of one or more antimicrobial drugs remain to be elucidated.

Prostatitis is an inflammatory condition of the prostate that presents with urethral symptoms, prostatic symptoms and sexual dysfunction. It is diagnosed by clinical symptoms and signs, the microscopy of expressed prostatic secretion (EPS) and culture of EPS and segmented urine samples. Prostatitis is chronic when symptoms have been present for at least 3 months. As a part of several scientific research projects we have been prospectively investigating prostatitis syndrome at the Outpatient Department of Urogenital Infections and Sexually Transmitted Diseases, Dr Fran Mihaljevic University Hospital for Infectious Diseases, Zagreb, Croatia, since March 1, 1999 and is still ongoing. We examined more than 3450 patients over 18 years of age with symptoms of chronic prostatitis and no evidence of lower genitourinary tract abnormalities. Azithromycin fulfills the criteria for the treatment of chronic prostatitis and sexually transmitted infections-high efficacy against the most common bacterial agents of urogenital and sexually transmitted infections, acceptable toxicity and tolerability, simple oral use, excellent penetration through chronic inflammatory changed prostate capsule, achieves high concentrations in prostatic tissue and slowly eliminates for 4-8 weeks, has anti inflammatory and immunomodulatory effect, and is active against bacterial biofilm. At total of 307 patients with chronic prostatitis caused by Chlamydia trachomatis were treated with azithromycin in a total dose of 4.0, 4.5 or 6.0 g for 3-4 weeks. Eradication of Chlamydia trachomatis was achieved in about 80%, with 70% of patients clinically cured. There were no statistically significant differences between these three dosage regimens. A total of 82 patients with chronic prostatits caused by Ureaplasma urealyticum were treated with azithromycin in a total dose of 4.5 g for 3 weeks. The eradication of Ureaplasma urealyticum was 85% and clinical cure 76%. Conclusion: azithromycin is an effective and safe antimicrobial drug for the treatment of chronic prostatitis caused by Chlamydia trachomatis or Ureaplasma urealyticum. It is recommended in a total dose of 4.5 g po. during 3 weeks, administered for 3 days weekly in a dose 1x500 mg po. per day. The results of recently published clinical studies on the treatment of chronic bacterial prostatitis with combination therapy with ciprofloxacin and azithromycin are impressive. We also have a limited, but positive experience.

Seksualno prenosive bolesti pokazuju izraziti potencijal širenja, a uz istodobno diskretne ili odsutne simptome bolesti bolesnik je u stanju pridonijeti zarazi u više osoba. Stigma koja prati spolno prenosive bolesti nadalje odgađa traženje liječničke pomoći, a time i efi kasno liječenje. Mnogi bolesnici ostaju nedijagnosticirani i neliječeni. Razvoj testova koji se zasnivaju na amplifikaciji nukleinskih kiselina uzročnika omogućio je razdoblje senzitivnih i robustnih dijagnostičkih postupaka. Nažalost, mnogi od tih testova nisu komercijalno dostupni ili su preskupi osobito za sredine koje ih najviše trebaju. Nemogućnost brze, točne i cijenom pristupačne dijagnostike zahtijeva eksploataciju druge mogućnosti, a to je jednostavna, prihvatljiva i sigurna terapija seksualno prenosivih bolesti., Sexually transmitted diseases exhibit a high spreading potential. Moreover, due to discrete or absent symptoms a patient can spread STDs to a number of other persons. Stigma associated with sexually transmitted diseases also delays a visit to a physician and effective treatment. Many patients remain undiagnosed and untreated. The development of tests based on the amplification of pathogen nucleic acids has resulted in more sensitive and robust diagnostic procedures. Unfortunately, many of these tests are either commercially unavailable or too expensive, especially for those areas where they are needed most. The lack of timely, accurate and affordable diagnostics requires the exploitation of other options, i.e. a simple, acceptable and safe therapy of sexually transmitted diseases.

Chlamydia trachomatis najčešći je uzročnik spolno prenosivih infekcija bakterijske etiologije i svake godine diljem svijeta uzrokuje 92 milijuna novih infekcija. Što se tiče zemalja Srednje i Istočne Europe, epidemiološka slika klamidijske genitalne infekcije ne može se sa sigurnošću procjenjivati. Naime, prijavljivanje klamidijskih infekcija ne provodi se sustavno, studije o prevalenciji su sporadične i teško metodološki usporedive, a preventivni programi na asimptomatskoj populaciji uglavnom ne postoje. Procjenjuje se da je incidencija klamidijske genitalne infekcije u Srednjoj i Istočnoj Europi 21 do 276 slučaja na 100.000 stanovnika. Javnozdravstvena važnost klamidijske genitalne infekcije očituje se u činjenici da je njezina prevalencija najviša u mladoj populaciji, da je najvećim dijelom asimptomatska te da neliječena infekcija može rezultirati kasnim posljedicama u obliku kronične upalne zdjelične bolesti, sterilnosti ili izvanmaternične trudnoće. Osim toga, pokazalo se da su troškovi liječenja komplikacija klamidijskih infekcija veći od troškova liječenja svih drugih spolno prenosivih bolesti, osim liječenja HIV/AIDS-a. Danas se klamidijska infekcija otkriva jednostavnim, pouzdanim i neinvazivnim testovima, a terapija izbora, 1 g azitromicina primijenjen jednokratno, dokazala se učinkovitom i sigurnom, posebice u slučajevima akutnih infekcija. Azitromicin se također pokazao lijekom izbora i za kronične infekcije, klamidijski prostatitis, kao i za liječenje značajnih komplikacija, kao što je npr. upalna zdjelična bolest. Kako bi pridonijela podizanju svijesti i znanja o klamidijskoj genitalnoj infekciji, PLIVA je 2002. godine na Brijunima okupila 40 stručnjaka iz Srednje, Istočne i Zapadne Europe, kao i Sjedinjenih Američkih Država koji se, s različitih aspekata, bave klamidijom. U tom smislu osnovana je Chlamydia Trachomatis Study Group (CTSG) - radna skupina za istraživanje C. trachomatis, koja je, u međuvremenu, prerasla u međunarodnu mrežu znanstvenika povezanih u međunarodnu udrugu., Chlamydia trachomatis is the most common causative agent of sexually transmitted infections of bacterial etiology, causing annually 92 million new infections worldwide. As far as Central and Eastern European ("CEE") countries are concerned, the epidemiological picture of chlamydial genital infections cannot be reliably estimated since there is no systematic registration, prevalence studies are sporadic and methodologically hardly comparable, and prevention programs focusing on asymptomatic population are scarce. The incidence of chlamydial genital infections in CEE is estimated to be 21 to 276 cases per 100,000 inhabitants. The public health importance of chlamydial genital infections lies in the fact that their prevalence is the highest in young population, that they are mostly asymptomatic and that, if left untreated, they may later result in chronic pelvic inflammatory disease, sterility or ectopic pregnancy. Additionally, the costs of treatment of chlamydial infection complications have shown to be higher than those of treatment of all other sexually transmitted diseases, with the exception of HIV/AIDS. Today chlamydial infections are diagnosed by simple, reliable and non-invasive tests, and the therapy of choice is a single dose of 1 g azithromycin, which has proven to be safe and effective, especially in acute infections. Azithromycin also proved to be the drug of choice in the treatment of chronic infections, chlamydial prostatitis, and in the treatment of major complications, such as pelvic inflammatory disease. In order to contribute to raising the awareness and knowledge about chlamydial genital infections, PLIVA gathered 40 experts treating different aspects of chlamydial infections from CEE, Western Europe and the United States on the Brijuni Islands in 2002. They established a Chlamydia Trachomatis Study Group (CTSG), i.e. the group for research of C. trachomatis, which in the meantime grew into an international association of scientists.