La tesi di dottorato è stata sviluppata all'interno del progetto nazionale Valorvitis ed è suddivisa in due parti. La prima riguarda la caratterizzazione chimica di sei estratti di vinacce d’uva ottenute diverse varietà di uva: tre a bacca rossa( Pinot Noir, Barbera e Nebbiolo) e tre a bacca bianca (Moscato, Muller Thurgau e Chardonnay). Ogni estratto è stato caratterizzato per contenuto e composizione in composti fenolici. La seconda attività consiste nello studio di un metodo alternativo di estrazione-purificazione basato sull'impiego di Gas Aphron Colloidali (CGAs) con la finalità di ridurre i costi di processo, la quantità di solvente da utilizzare durante la fase di estrazione e, allo stesso tempo, aumentare il grado di purezza dell’estratto in termini di contenuto polifenolico. / The present PhD thesis work was carried out under the framework of the Valorvitis.Extracts have been obtained from different red (Pinot Noir, Barbera and Nebbiolo) and white (Moscato, Muller Thurgau and Chardonnay) grape marcs and quantitatively analysed for the content of total phenolics, tannins, anthocyanins, cinnamic acids, flavonoids, flavonols, sugars (glucose and fructose) and for the anti-oxidant power (AOP). A new extraction-purification process based on CGAs application has been investigated. CGAs were first reported by Sebba (1987) as micro bubbles, with dimension among 10–100 μm, composed of a gaseous inner core surrounded by a thin surfactant film, which are created by intense stirring of a surfactant solution above its critical micellar concentration. Surfactants are able to catch phenolics thanks to both electrostatic and hydrophobic interactions depending on the surfactant (ionic or non-ionic) and on the phenolic compound. In this project the low-cost non-ionic food grade surfactant Tween 20 was used.

<p>Glavni cilj istraživanja ove doktorske disertacije je bio valorizacija sporednog proizvoda žalfije (Salvia officinalis L.) iz fabrike čaja za dobijanje visoko-vrednih ekstrakata željenih osobina koji bi imali primenu u gotovim proizvodima prehrambene, kozmetičke i farmaceutske industrije.<br />U cilju ekstrakcije polifenolnih jedinjenja su primenjene konvencionalne (maceracija) i savremene (ultrazvučna ekstrakcija, mikrotalasna ekstrakcija i ekstrakcija subkritičnom vodom) ekstrakcione tehnike. U postupaku maceracije je optimizovana koncentracija etanola u ekstragensu, dok je za optimizaciju savremenih ekstrakcionih tehnika primenjen eksperimentalni dizajn i metoda odzivne povr&scaron;ine. Prinos polifenola (ukupnih fenola i ukupnih flavonoida) i antioksidativna aktivnost određena različitim in vitro testovima su bili ispitivani odzivi. Za svaki od navedenih ekstrakcionih postupaka je ispitivan uticaj glavnih ekstrakcionih parametara: ultrazvučna ekstrakcija (temperatura, vreme ekstrakcije i snaga ultrazvuka), mikrotalasna ekstrakcija (koncentracija etanola, vreme ekstrakcije i odnos rastvarač-droga) i ekstrakcija subkritičnom vodom (temperatura, vreme ekstrakcije i koncentracija HCl u ekstragensu). Tečni ekstrakti dobijeni ultrazvučnom, mikrotalasnom i subkritičnom ekstrakcijom su pokazali izuzetno visoku antioksidativnu aktivnost određenu &bdquo;hvatanjem&ldquo; DPPH i O2∙- radikala i kapacitetom redukcije Fe3+ jona i imali su znatno veći sadržaj polifenola u odnosu na ekstrakte dobijene maceracijom.<br />U cilju ekstrakcije terpenoidnih jedinjenja su primenjeni postupci Soxhlet ekstrakcije, destilacije vodenom parom i ekstrakcije superkritičnim CO2. Postupak superkritične ekstrakcije je optimizovan metodom odzivne povr&scaron;ine i optimalni uslovi ekstrakcije su bili pritisak 290 bar, temperatura 55˚C i protok CO2 od 0,4 kg/h, dok su predviđene i eksperimentalne vrednosti prinosa ekstrakcije bile 8,90 i 8,84%. Hemijskom analizom lipofilnih ekstrakata i etarskog ulja je utvrđeno da su oksidovani monoterpeni (&alpha;-tujon i kamfor), oksidovani seskviterpeni (viridiflorol) i polifenolni diterpeni (epirosmanol) dominantna jedinjenja prisutna u ovim uzorcima.<br />Tečni ekstrakti žalfije dobijeni konvencionalnim i savremenim ekstrakcionim tehnikama su spray drying tehnikom osu&scaron;eni u cilju prevođenja u stabilniju formu suvog ekstrakta. Suvim ekstraktima su određene fizičko-hemijske osobine (hemijski sastav, sadržaj vlage, higroskopnost, moć rehidratacije, WAI i WSI) i biolo&scaron;ka aktivnost (antimikrobna i antioksidativna aktivnost) i procenjena je njihova mogućnost primene u prehrambenoj i farmaceutskoj industriji.</p> / <p>The main aim of this dissertation was valorization of sage (Salvia officinalis L.) by-product from filter-tea factory for recovery of high-value extracts with desirable properties for application in food, cosmetics and pharmaceutical products.<br />Conventional (maceration) and novel (ultrasound-assisted, microwave-assisted and subcritical water extraction) extraction techniques were applied for polyphenols recovery. Ethanol concentration was optimized in maceration process, while experimental design and response surface methodology were applied for optimization of novel extraction techniques. Polyphenols yield (total phenols and total flavonoids) and antioxidant activity, determined by various in vitro assays, were investigated responses. Influence of main process parameters was determined for each technique: ultrasound-assisted extraction (temperature, extraction time and ultrasonic power), microwave-assisted extraction (ethanol concentration, extraction time and sample-solvent ratio) and subcritical water extraction (temperature, extraction time and HCl concentration in solvent). Liquid extracts obtained by novel extraction technique exhibited higher antioxidant activity determined by scavenging of DPPH and O2∙- radicals and reduction of Fe3+ and also provided higher polyphenols yield comparing to extracts obtained by maceration. Soxhlet extraction, hydrodistillation and supercritical fluid extraction were applied for terpenoids recovery. Supercritical fluid extraction process was optimized by response surface methodology and optimal conditions were pressure of 290 bar, temperature of 55˚C and CO2 flow rate of 0.4 kg/h, while predicted and experimentally obtained values of total extraction yield at these conditions were 8.90 i 8.84%, respectively. According to chemical analysis, the most abundant compounds in lipid extracts and essential oil were oxygenated monoterpenes (&alpha;-thujone and camphor), oxygenated sesquiterpenes (viridiflorol) and diterpene polyphenols (epirosmanol).<br />Sage liquid extracts obtained by conventional and novel extraction techniques were spray dried in order to obtain dry extract form. Physico-chemical properties (chemical profile, moisture content, hygroscopicity, rehydratation time, WAI and WSI) and biological activity (antimicrobial and antioxidant activity) were determined in dry extracts and their potential application in food and pharmaceutical formulations was discussed.</p>

Circa l’1% della popolazione mondiale è colpita da celiachia. I celiaci sono costretti a seguire una dieta priva di glutine e molto spesso quest’ultima risulta essere sbilanciata e/o carente in molti nutrienti. Recentemente, l’uso di matrici alternative al frumento, come pseudocereali, legumi e cultivar di riso pigmentate sta riscuotendo grande interesse a causa del loro elevato quantitativo di composti bioattivi (polifenoli). Quindi, considerando l’importanza attuale dei polifenoli sia nella formulazione tecnologica che nella promozione di aspetti salutistici degli alimenti senza glutine, lo scopo di questa tesi è stata basata su: 1) profilazione dei polifenoli in matrici prive di glutine (farine non di frumento, legumi, pseudocereali e frutta secca) e loro proprietà antiossidanti in vitro; 2) valutazione dell’impatto di trattamenti termici e di fermentazioni microbiche sul profilo fenolico di queste matrici prive di glutine; 3) valutazione del ruolo dei polifenoli come inibitori degli enzimi amilolitici; e 4) valutazione del destino dei polifenoli caratterizzanti alimenti senza glutine durante processi in vitro simulanti digestione gastrointestinale e fermentazione fecale. I polifenoli sono stati analizzati sfruttando tecniche di metabolomica mirata/non-mirata. / Around 1% of world population is affected by coeliac disease. Coeliac people are constrained to follow a strict gluten free (GF) diet and very often this latter is unbalanced and lacks in many nutrients. In the last years, the exploitation of alternative crops or underutilized species, such as pseudocereals, legumes and pigmented cereal cultivars, is gaining interest because of their amount and profile of bioactive compounds, such as polyphenols. Therefore, considering the actual importance of polyphenols for both the formulation of GF foods and their health-promoting properties, the current PhD thesis was based on: 1) the profiling of polyphenols in GF raw materials (such as non-wheat flours, legumes, pseudocereals and nuts) and their in vitro antioxidant activities; 2) the evaluation of the impact of different heat treatments and microbial fermentations on the phenolic profile of GF raw materials; 3) the investigation of polyphenols in GF foods as inhibitors of digestive enzymes; and 4) the assessment of the fate of polyphenols characterizing GF foods during simulated in vitro gastrointestinal and fermentation processes. Polyphenols were analysed by means of targeted/untargeted metabolomics-based approaches (i.e., high resolution chromatography and mass spectrometry platforms).

<p>U okviru ovog rada izvr&scaron;eno je ispitivanje različitih procesa ekstrakcije herbe industrijske konoplje. Za ekstrakciju određenih bioaktivnih jedinjenja primenjene su klasične tehnike ekstrakcije, maceracija i Soxhlet ekstrakcija, dok su od savremenih metoda primenjene ultrazvučna ekstrakcija, mikrotalasna ekstrakcija, ekstrakcija vodom u subkritičnom stanju i ekstrakcija ugljendioksidom u superkritičnom stanju. Praćenjem uticaja ulaznih promenjivih (procesnih parametara) na odabrane izlazne promenjive (sadržaj bioaktivnih jedinjenja ili antioksidativnu aktivnost) za svaku tehniku ekstrakcije pojedinačno utvrđeni su optimalni uslovi ekstrakcije. Nakon izvr&scaron;enog poređenja odabrana je najadekvatnija tehnika ekstrakcije za pripremu tečnog ekstrakta koji će biti transformisan u suvi ekstrakt primenom metode su&scaron;enja raspr&scaron;ivanjem. Prilikom dobijanja suvog ekstrakta ispitan je uticaj različite koncentracije nosača na kvalitet ekstrakata i efikasnost su&scaron;enja. Dobijeni ekstrakti su okarakterisani u pogledu fizičkih i hemijskih osobina. Sadržaj kanabinoida u tečnim ekstraktima suve herbe industrijske konoplje ispitanih u ovoj doktorskoj disertaciji je bio u opsegu od 0,4314 mg/mL CBD-a i 0,0364 mg/mL THC-a (u optimalnom ekstraktu dobijenom maceracijom) do 1,0420 mg/mL CBD-a i 0,0430 mg/mL THC-a (u optimalnom ekstraktu dobijenom mikrotalasnom ekstrakcijom). U ekstraktima koji su nepolarnog karaktera i koji su dobijeni primenom Soxhlet ekstrakcije sadržaj CBD-a i THC-a je bio 64,40 i 2,90 mg/g, dok je primenom superkritične ekstrakcije dobijen sadržaj CBD-a i THC-a u opsegu od 76,1193 do 163,1111 mg/g i od 4,1746 do 6,5803 mg/g. Tečni ekstrakti dobijeni primenom vode i etanola kao ekstragensa bili su bogati flavonoidima i fenolima, a utvrđeno je i da imaju značajnu antioksidativnu aktivnost, dok su ekstrakti dobijeni Soxhlet ekstrakcijom i ekstrakcijom ugljendioksidom u superkritičnom stanju bili bogati lipofilnim jedinjenjima i terpenima. Ovako dobijeni ekstrakti predstavljaju visokovredne proizvode koji imaju veliki potencijal za upotrebu u farmaceutskoj industriji.</p> / <p>Various extraction processes of the aerial parts of industrial hemp were examined. From classic extractions, maceration and Soxhlet extraction were applied, while from novel methods, ultrasound-assisted extraction, microwave-assisted extraction, subcritical water extraction and supercritical carbon dioxide extraction were applied. By monitoring the influence of the input variables on the selected output variables for each extraction, the optimal extraction conditions were determined separately. After a comparison of all the extraction processes tested, microwave-assisted extraction was selected to prepare the liquid extract for spray drying. The effect of different carrier concentrations was examined during the preparation of the dry extract. The extracts obtained were characterized in terms of physical and chemical properties. The content of cannabinoids in liquid extracts of dry Cannabis sativa herba examined ranged from 0.4314 mg/mL for CBD and 0.0364 mg/mL for THC in the optimum extract obtained by maceration to 1.0420 mg/mL for CBD and 0.0430 mg/mL for THC in the optimum extract obtained by microwave-assisted extraction. The CBD and THC content in extract obtained by Soxhlet extraction was 64.4000 mg/mL and 2.9000 mg/mL, respectively, while the highest content of CBD and THC in the supercritical carbon dioxide extract was 163.11 and 6.5803 mg/g. In addition to cannabinoids, the liquid extracts obtained were rich in phenols and flavonoids, and have considerable antioxidant activity, while the extracts obtained by Soxhlet extraction and supercritical carbon dioxide were rich in lipophilic compounds and terpenes. The extracts mentioned above are high value products which have great potential for implementation in the pharmaceutical industry.</p>

<p style="text-align: justify;">U okviru ove doktorske disertacije ispitana je mogućnost ekstrakcije bioaktivnih jedinjenja iz biljne pra&scaron;ine A. uva-ursi, koja nastaje kao sporedni proizvod u fabrici filter čaja. Za ekstrakciju bioaktivnih jedinjenja primenjene su različite tehnike ekstrakcije (klasična ekstrakcija, ultrazvučna ekstrakcija i ekstrakcija subkritičnom vodom), kao i različiti ekstragensi: voda na sobnoj temperaturi, 30-70% etanol i subkritična voda. Sadržaj bioaktivnih jedinjenja u ekstraktima ispitan je primenom odgovarajućih spektrofotometrijskih metoda i HPLC metode. U ekstrakciji biljne pra&scaron;ine A. uva-ursi detaljno je matematički analiziran uticaj procesnih parametara na ekstrakciju subkritičnom vodom bioaktivnih jedinjenja iz materijala ovog tipa i u tom smislu je posmatran uticaj temperature, vremena i dodatka kiseline, a takođe su određeni uslovi pri kojima je, primenom ove tehnike, moguće ostvariti maksimalnu ekstrakciju bioaktivnih jedinjenja od značaja. Pored toga prikazana je i mogućnost primene spray drying tehnike za su&scaron;enje tečnih i proizvodnju suvih ekstrakta A. uva-ursi u formi praha. Na kraju, na osnovu koncentracije bioaktivnih jedinjenja i njihovog delovanja, dat je predlog nekoliko novih preparata na bazi ekstrakata A. uva-ursi, u tečnoj i čvrstoj formi.</p> / <p>This dissertation investigated the possibility to extract bioactive compounds from A. uva-ursi herbal dust, which occurs as a by-product at the filter-tea<br />factory. Various extraction techniques (classical extraction, ultrasound assisted extraction and subcritical water extraction), were applied for the extraction of bioactive compounds, as well as different solvents: water at room temperature, 30-70% ethanol and subcritical water. The content of bioactive compounds in the extracts was investigated using appropriate spectrophotometric methods and the HPLC method. In the extraction of A. uva-ursi herbal dust a detailed mathematical analysis of the influence of process parameters on subcritical water extraction of bioactive compounds from this type of material was conducted. In this regard the influence of temperature, time and acid addition was observed and the conditions under which it is possible to achieve maximum extraction of significant bioactive compounds, by applying this technique, were determined. In addition, the possibility of applying the spray drying technique for drying the liquid and producing dry A. uva-ursi extracts in the form of powder is also presented. Eventually, based on the concentration of the bioactive compounds and their activity, a proposal for a several new preparations in liquid and solid form, based on A. uva-ursi extracts is given.</p>

<p>Ugljenične nanocevi (UNC) imaju sve veću primenu u elektronici, kompjuterskoj i optičkoj industriji, kao i u biomedicini. Dok proizvodnja jednoslojnih UNC nanocevi beleži sve veći rast poslednjih godina, rizik koji nosi izlaganje ovom nanomaterijalu ostaje nerazja&scaron;njen. Oskudni i često kontradiktorni podaci o toksičnim efektima jednoslojnih UNC ukazuju na potrebu za daljim ispitivanjima. U na&scaron;im istraživanjima ispitivane su promene u ćelijskom odgovoru kao i morfolo&scaron;ke promene nakon delovanja jednoslojnih ugljeničnih nanocevi na ćelijskoj liniji humanih fetalnih fibroblasta pluća MRC-5 i ćelijskoj liniji humanog adenokarcinoma pluća A549. U ovoj studiji kori&scaron;ćene su jednoslojne ugljenične nanocevi koje su sadržale rezidualne nečistoće poput gvožđa. Citotoksičnost jednoslojnih UNC (engl. single-walled carbon nanotubes &ndash; SWCNT) je ispitivana kolorimetrijskim MTT testom. Tokom 24 i 48h niske koncentracije jednoslojnih ugljeničnih nanocevi (&lt;250 &mu;g/mL) pokazale su nisku toksičnost na proliferaciju i vijabilnost u obe ispitivane ćelijske linije. Ipak, pri visokim koncentracijama UNC (250-750 &mu;g/mL) antiproliferativni efekat je bio blizu IC50 vrednostima. Na osnovu rezultata dobijenih MTT testom može se zaključiti da su maligne A549 ćelije osetljivije na delovanje jednoslojnih UNC u odnosu na normalne MRC-5 ćelije. Kombinacija ugljeničnih nanocevi sa prirodnim polifenolima (resveratrolom i proantocijanidolima) nije značajno uticala na citotoksičnost u MRC-5 ćelijama, za razliku od A549 ćelija gde je tretman kombinacijama umanjio toksičnost ugljeničnih nanocevi. Transmisionom elektronskom mikroskopijom ispitivan je efekat jednoslojnih ugljeničnih nanocevi na ćelijsku morfologiju i preživljavanje. Intracelularni agregati ugljeničnih nanocevi primećeni su u obe ćelijske linije, čime je potvrđeno da ugljenične nanocevi ulaze u ćelije. Imajući u vidu da nanomaterijali poput ugljeničnih nanocevi indukuju oksidativni stres i njime posredovanu apoptozu, na protočnom citometru je određivano prisustvo ćelija u apoptozi i nekrozi. Tretman ćelija sa jednoslojnim ugljeničnim nanocevima nije doveo do značajnog porasta broja apoptotskih ili nekrotičnih ćelija, &scaron;to ide u prilog niskoj toksičnosti ovog nanomaterijala, odnosno ukazuje na alternativne mehanizme toksičnosti. Međutim kombinacija jednoslojnih ugljeničnih nanocevi sa antioksidantima, resveratrolom i proantocijanidolima indukuje veći procenat apoptoze i nekroze u odnosu na tretman samo sa nanocevima. Promene u ekspresiji gena praćene su lančanom reakcijom polimeraza (PCR). Komparativna analiza rezultata genske ekspresije MRC-5 i A549<br />ćelija nakon tretmana sa jednoslojnim ugljeničnim nanocevima pojedinačno i u kombinaciji sa antioksidantima ukazala je na kompleksnost i raznolikost biolo&scaron;kog odgovora ispitivanih ćelija. U na&scaron;em istraživanju ispitivana je i promena aktivnosti enzima antioksidativne za&scaron;tite i količine glutationa u ćeliji. Primena jednoslojnih ugljeničnih nanocevi u MRC-5 ćelijama dovodi po smanjenja specifične aktivnosti enzima SOD i GR, povećava specifičnu aktivnost GPx i ne utiče na promenu specifične aktivnosti GST i količine glutationa u ćeliji Primena jednoslojnih ugljeničnih nanocevi u A549 ćelijama dovodi po smanjenja specifične aktivnosti enzima SOD, ne utiče na promenu specifične aktivnost enzima GR, GST i GPx, i dovodi do povećanja količine glutationa u ćeliji. Ćelijska vijabilnost, morfolo&scaron;ke promene, redoks homeostaza i ekspresija ispitivanih gena bile su promenjene nakon tretmana sa jednoslojnim ugljeničnim nanocevima. Iako su dobijeni rezultati značajni za procenu toksičnosti ugljeničnih nanocevi, neophodna su dalja istraživanja koja treba da doprinesu boljem razumevanju toksičnih efekata ugljeničnih nanocevi.</p> / <p>Carbon nanotubes are being actively introduced in electronics, computer science, and optics as well as for various biomedical applications. While production of single-walled carbon nanotubes (SWCNT) has escalated in recent years, the knowledge on risk associated with exposure remains unclear. Contradictory data on the toxic effects of single-walled carbon nanotubes highlights the urgent need for further studies. In this study we investigated the alterations in cellular response along with morphological changes induced by single-walled carbon nanotubes in human lung fibroblast cell line MRC-5 and adenocarcinoma human alveolar basal epithelial cells A549. In this study we used SWCNT containing large amounts of residual metallic impurities such is iron, and the iron concentration increased in dose dependent manner in cells exposed to SWCNT. Cytotoxicity was evaluated by MTT assay and SWCNT showed little cytotoxic effect on the proliferation and viability of two cell lines tested at the concentrations used (&lt;250 &mu;g/mL) within 24 and 48h. However exposing both cell lines to high concentrations (250-750 &mu;g/mL) resulted in near IC50 values. Based on MTT test SWCNT were more cytotoxic to A549 cell line. Cytotoxicity of SWCNT in combination with natural polyphenols (resveratrol and proanthocyanidins) did not noticeably affect the cytotoxicity of SWCNT to MRC-5 cells. However introduction of polyphenols did reduce the cytotoxicity of SWCNT to A549 cells. Transmission electron microscopy was used to complement cytotoxicity assays and to investigate the pathological effect of internalized SWCNT on cell morphology and survival. Intracellular bundles of CNTs, possibly aggregated/agglomerated were observed in both cell lines, confirming internalization after 24h exposure. Since nanoparticles like carbon nanotubes are toxic mainly because they cause oxidative stress, often associated with an increased apoptosis we checked for apoptotic and necrotic cells using flow cytometry. Incubation with SWCNT did not result in pronounced apoptosis or necrosis supporting its low toxicity and possibly alternative mechanism of cell damage. However incubation with SWCNT in combination with resveratrol and proanthocyanidins induced higher levels of both apoptosis and necrosis than SWCNT alone. Changes in gene expression following exposure to SWCNT were evaluated by polymerase chain reaction PCR array which indicated complex and diverse change in expression of genes involved in apoptosis, cell proliferation and oxidative stress. Finally we investigated the modulation of the antioxidant enzyme system and the changes in the cytosolic levels of GSH. SWCNT reduced the specific activity of SOD and GR enzymes, increased GPx activity. No changes in intracellular levels of GSH were observed in MRC-5 cell line. Same treatment in A549 cell reduced the specific activity of SOD, had no effect on GR, GST and GPx activity, but increased intracellular levels of GSH. Cell viability, morphologic changes, redox homeostasis and gene expression were affected by the presence of SWCNT. Although our findings are useful in predicting human response against SWCNT exposure, further study is needed for better understanding of the effects of SWCNT.</p>

<p>U okviru disertacije kreirani su novi, nutritivno obogaćeni, fini pekarski proizvodi na bazi bra&scaron;na od cerealija i pseudocerealija, kroz čiju se proizvodnju valorizuju sporedni proizvodi prehrambene industrije &ndash; trop cvekle. Rezultati istraživanja pružaju nova saznanja o stepenu redukcije bioaktivnih jedinjenja (polifenola i betalaina) tokom postupka proizvodnje i skladi&scaron;tenja finih pekarskih proizvoda obogaćenih ekstraktom tropa cvekle, kao i njihov uticaj na reakcije neenzimskog tamnjenja.<br />Uspe&scaron;no realizovana istraživanja ukazala su na stepen stabilinosti bioaktivnih jedinjenja iz tropa cvekle inkapsulacijom na protein soje i na opravdanost primene ove metode (tehnike) u proizvodnji funkcionalne hrane. Na ovaj način se obezbeđuju funkcionalni proizvodi koji sadrže dovoljnu količinu visokovrednih jedinjenja koja imaju veću bioraspoloživost i finalan zdravstveni efekat. Zbog niza prednosti inkapsulirane forme ekstrakta imaju veliki potencijal primene u industriji u vidu boja i aditiva, tj. funkcionalnih dodatakau prehrembenim ali i kozmetičkim proizvodima. Istraživanja u okviru disertacije omogućavaju dobijanje inovativnih proizvoda koji pro&scaron;iruju asortiman nutritivno vrednih proizvoda i doprinose kvalitetnijoj ishrani potro&scaron;ača.<br />Upotrebom nusproizvoda prerade voća i povrća smanjuje se količina otpada i doprinosi za&scaron;titi životne sredine, povećava dobit proizvođača kroz pobolj&scaron;anje iskori&scaron;ćenja sirovine i konkurentnosti na trži&scaron;tu.</p> / <p>Within the thesis a new, nutrient-rich, fine bakery products based on flour of cereals and pseudocereals were created, through the production of which valorizing by-products of the food industry are produced- pomace beetroot. Results of the research provide new knowledge about degree of reduction bioactive compounds (polyphenols and betalains) during process of production and storage of fine bakery products enriched with beetroot pomace extracts, as well as their influence on non-enzymatic tanning reactions.<br />Successfully realized researches have indicated degree of stability bioactive compounds from beetroot pomace by encapsulation on soy protein and justification of application this method (technique) in the production of functional food. In this way, functional products witch contain a sufficient amount of high-value compounds which have higher bioavailability and final health effect are provided. Due to a number of advantages, encapsulated form of the extract have a great potential for application in industry like food color and additives, ie. functional supplements for food and cosmetic products. Research in the framework of the thesis allow obtaining innovative products which expand the range of nutritionally valuable products and contribute to the quality consumers diet.<br />Using by-products of processing fruits and vegetables reduces the amount of waste and contributes to the protection of environment, increases the profit of producers by improving the efficiency of raw materials and market competitiveness.</p>

<p>Nesavesna upotreba antibiotika izazvala je razvoj antimikrobne rezistencije zabrinjavajućih razmera. Porast rezistencije prema konvencionalnim antimikrobicima nametnuo je potrebu pronalaska novih supstanci, koje će biti delotvorne protiv otpornih sojeva. U poslednje četiri decenije, dominira trend da se lekovi i lekovita sredstava prave na bazi prirodnih proizvoda, a naročito biljnog materijala. Hemijski sastav plodova porodice ruža (Rosaceae) upućuje na to da bi određene vrste ove porodice biljaka mogle ispoljavati antimikrobno dejstvo. Plodovi voća porodice Rosaceae istraživani u okviru ove disertacije su: malina (Rubus idaeus), kupina (Rubus fruticosus), tre&scaron;nja (Prunus avium), vi&scaron;nja (Prunus cerasus) i aronija (Aronia prunifolia). Predmet istraživanja ove doktorske disertacije je antimikrobno tj. antibakterijsko, antigljivično, antialgalno i antivirusno dejstvo sokova i ekstrakata plodova odabranog voća porodice Rosaceae gajenog i prikupljenog na teritoriji Srbije na &scaron;irok spektar humanih i animalnih patogenih mikroorganizama.Sokovi su dobijeni ceđenjem svežih plodova voća. Ekstrakti su dobijeni iz ostataka voća nakon ceđenja (tropa) metodom dvostruke etanolne ekstrakcije. Nakon ceđenja i ekstrakcije sprovođeno je uparavanje do suva sokova i ekstrakata. HPLC metodom je ispitano prisutstvo 23 hemijske komponente u svakom od sokova i ekstrakata. Antimikrobna analiza je sprovedena na 21 mikroorganizmu, od čega su 15 bile bakterije, 2 gljivice, 2 alge i 2 virusi. Od navedenih mikroorganizama 14 je bilo autohtonih izolata, a 7 referentnih sojeva. Ispitivanje antibakterijskog, antifungalnog i antialgalnog dejstva plodova odabranog voća je sprovedeno difuzionom i mikrodilucionom metodom. Analiza antivirusne aktivnosti plodova odabranog voća je sprovedena posmatranjem inhibicije citopatogenog efekta (CPE) virusnih čestica na ćelijsku kulturu. Sve studije su sprovedene u triplikatu. Rezultati hemijske analize su obrađeni statistikom metodom srednje vrednosti, uz računanje standardne devijacije. Eksperimentalno dobijeni podaci u okviru ispitivanja antibakterijskog, antigljivičnog i antialgalnog dejstva su uvr&scaron;teni u statističku analizu. Metodom aproksimacije je utvrđena tačna koncentracija soka ili ekstrakta koja daje željeni efekat na ispitivani mikroorganizam. Rezultati analize antivirusnog dejstva su statistički obrađeni metodom srednje vrednosti. Kako bi se utvrdio stepen povezanosti rezultata dobijenih u okviru antimikrobnih analiza sa hemijskim supstancama prisutnim u plodovima odabranog voća sprovedena je regresiona analiza za vaki pojedinačni slučaj. Antibakterisjko, antiungalno i antialgalno dejstvo sokova i ekstrakata je upoređeno sa istim dejstvima konvencionalnih antimikrobnih lekova. Najveću ukupnu količinu ispitivanih supstanci je sadržao ekstrakt kupine (3965,56 mg/ml), a najmanju ekstrakt tre&scaron;nje (161,44 mg/ml). Od svih ispitivanih hemijskih supstanci, samo su: antocijani, katehin, galna kiselina i vitamin C, prisutni u svim ispitivanim sokovima i ekstraktima. Sve ispitivane biljke sadrže veću količinu supstanci u ekstraktu, osim kod tre&scaron;nje, kod koje se veća količina supstanci nalazi u soku. U najvećoj meri su u sokovima i ekstraktima plodova biljaka prisutni antocijani i to najvi&scaron;e kod kupine od ekstrakata, a kod maline od sokova. Antimikrobno (antibakterijsko, antifungalno, antialgalno i antivirusno) dejstvo je dokazano za svaki od sokova i/ili ekstrakata plodova odabranog voća. Po prosečnim vrednostima rezultata dilucione analize antimikrobnog i dejstva na MDBK ćelije, protiv najvećeg broja mikroorganizama najdelotvorniji je bio ekstrakt kupine, a neaktivan ili najslabije aktivan (aktivan u najvećoj koncentraciji) sok tre&scaron;nje. Prosečne vrednosti antimikrobnog i dejstva na MDBK ćelije su bile u opsegu od 0,71 mg/ml do 100 mg/ml. Sve ispitivane supstance su pokazale antibakterijsku aktivnost u ispitivanim koncentracijama na sve ispitivane bakterije, osim soka maline na Escherichia-u coli i soka aronije na Klebsiella-u pneumoniae. Ekstrakti su bolje delovali u većini slučajeva, uz nekoliko izuzetaka. Sve ispitivane supstance su pokazale antfiungalnu aktivnost na Candida-u albicans ATCC 24433, osim soka aronije. Na autohtoni soj Candida-e albicans su delovali samo ekstrakti kupine i tre&scaron;nje. Ekstrakti su bolje delovali u većini slučajeva, osim kupine na Candida-u albicans ATCC 24433, u kojem je dejstvo soka bilo jednako sa dejstvom ekstrakta. Iako su skoro svi ispitivani sokvi i ekstrakti ispoljili antialgalno dejstvo, na Prototheca-u wickerhamii nisu delovali vi&scaron;nja i sok maline, dok sok aronije nije delovao antialgalno ni na jednu od ispitivanih Prototheca. Ekstrakti su bolje delovali u većini slučajeva, osim maline kod koje je, u oba slučaja, bolje delovao sok. Sve ispitivane supstance su pokazale antivirusnu aktivnost na Bovine herpesvirus i Bovine viral diarrhea virus, kao i toksičnost na MDBK ćelije, osim sokova maline i tre&scaron;nje, koji su bili neaktivni protiv Bovine herpesvirus-a. Ekstrakti su bolje delovali u većini slučajeva, osim tre&scaron;nje na MDBK ćelije i Bovine viral diarrhea virus, na koje je jače delovao sok. U potpunosti jednako delovanje soka i ekstrakta je uočeno kod aronije za sva tri dejstva i maline za toksičnost na MDBK ćelije. Pri ispitivanju difuzionom metodom antibiotik je ispoljio jače dejstvo u odnosu na sokove i ekstrakte, osim ekstrakata kupine, maline i vi&scaron;nje na Trueperella-u pyogenes. Pri ispitivanju dilucionom metodom, dejstvo ekstrakata i sokova na Gram pozitivne bakterije je bilo uporedivo sa dejstvom Streptomicina, dok je dejstvo antibiotika na Gram negativne bakterije bilo jače u odnosu na sokove i ekstrakte, izuzetev jačeg dejstva ekstrakta maline protiv Escherichia-e coli. Nistatin je sve antifungalne i antialgalne efekte na gljivice i alge obuhvaćene istraživanjem ispoljio u manjim koncentracijama u odnosu na ispitivane sokove i ekstrakte.Najpovezanije sa antibakterijskim dejstvom sokova i ekstrakata na odgovarajuće mikroorganizme su bile: kumarinska, ferulna i galna kiselina, katehin. Najveći procenat povezanosti sa antimikrobnom aktivnosti ploda imali su antocijani na Enterococcus faecalis (71,94%). Najmanje povezani sa antimikrobnom aktivnosti na odgovarajuće mikroorganizme su bili: hlorogenska kiselina, kempferol, kumarinska i siringinska kiselina, vitamin C, vanilinska kiselina, epikatehin, katehin, rutin, krisin, elaginska i p-hidroksibenzoeva kiselina. Saznanja dobijena u okviru ove disertacije pro&scaron;iruju obim znanja o antimikrobnom dejtvu voća i predstavljaju osnovicu za svrsishodniju primenu voća kako u ishrani tako i u terapiji. Međutim, dalja istraživanja u smislu pro&scaron;irenja spektra ispitivanih mikroorganizama, antimikrobnih studija pojedinačnih komponenti iz plodova i mehanizama dejstva su potrebna.</p> / <p>Nonconscientious use of antibiotics has caused the development of antimicrobial resistance of worrying proportions. The increase in resistance to conventional antimicrobials has imposed a need to find new substances that will be effective against resistant strains. Over the last four decades, the trend is to medicines and medicinal products be made on the basis of natural products, and especially plant material. The chemical composition of the Rose family (Rosaceae) fruits suggests that certain species of this family of plants may exhibit an antimicrobial effect. The fruits of the Rosaceae family studied in the framework of this dissertation are: Raspberry (Rubus idaeus), Blackberry (Rubus fruticosus), Cherry (Prunus avium), Cherry (Prunus cerasus) and Aronia (Aronia prunifolia). The subject of the research of this doctoral dissertation is antimicrobial ie. antibacterial, antifungal, antialgal and antiviral effects of juices and fruit extracts, of the selected fruit of the Rosaceae family collected on the territory of Serbia, on a wide range of human and animal pathogenic microorganisms. Juices were obtained by the process of squeezing fresh fruits. The extracts were obtained from residues of fruits after extraction (pomace) by the double ethanol extraction method. After squeezing and extraction, evaporation to dryness of juices and extracts was carried out. The presence of 23 chemical components in each of the juices and extracts was examined by the HPLC method. Antimicrobial analysis was carried out on 21 microorganisms, of which 15 were bacteria, 2 fungi, 2 algae and 2 viruses. 14 of these microorganisms were autochthonous isolates and 7 reference strains. The antibacterial, antifungal and antialgal effect of the selected fruit was tested by the diffusion and microdilution method. The analysis of the fruits antiviral activity was carried out by observing the inhibition of cytopathogenic effect (CPE) of viral particles per cell culture. All studies were conducted in triplicate. The results of the chemical analysis were processed by means of a mean value method, with calculation of the standard deviation. The experimentally obtained data within the antibacterial, antifungal and antialgal examination were included in the statistical analysis. The method of approximation was used in order to determine the exact concentration of the juice or extract, which gives the desired effect on the examined microorganisms. The results of the antiviral analysis were statistically processed using the mean value method. In order to determine the degree of correlation of the results obtained in the framework of antimicrobial analysis with the chemical substances present in the fruits of the selected fruit, regression analysis was carried out for each individual case. Antibacterial, antifungal and antialgic effects of juices and extracts are compared with he same effects of conventional antimicrobial drugs, Streptomycin and Nystatin. The largest total amount of test substances was contained by blackberry extract (3965.56 mg/ ml), and the smallest by extract of sweet cherry (161.44 mg/ ml). Of all the investigated chemicals, only: anthocyanins, catechins, gallic acids and vitamin C were present in all examined juices and extracts. All investigated plants contain a higher amount of substances in the extract, except for sweet cherries, in which a greater amount of substances is present in the juice. In the juices and extracts of fruits anthocyanins are present to the greatest extent, mostly in the blackberry extract, and in raspberry juices. According to the average values of the results of the dilution analysis of the antimicrobial and the effect on the MDBK cell, the most effective was the blackberry extract and the most active inactive (active in the highest concentration) cherry juice against the most numerous microorganisms. The average antimicrobial values and effects on the MDBK cells were in the range of 0.71 mg/ ml to 100 mg/ ml. All investigated substances showed antibacterial activity in the tested concentrations on all examined bacteria, except raspberry juice on Escherichia coli and aronia juice on Klebsiella pneumoniae. Extracts acted better in most cases, with few exceptions. All investigated substances showed antifungal activity on Candida albicans ATCC 24433, except aronia juice. On the autochthonous Candida albicans, only blackberry and sweet cherry extracts were active. Extracts worked better in most cases, except for blackberry on Candida albicans ATCC 24433, in which the effect of the juice was equally efficient as the extract. Although almost all of the examined juices and extracts exhibited an antialgal effect, the sour cherry and raspberry juice did not work on the Prototheca wickerhamii, while the aronia juice did not express antialgal activity to any of tested Prototheca isolates. Extracts worked better in most cases, apart from raspberries, wich juice was stronger in both cases. All investigated substances showed antiviral activity on Bovine herpesvirus and Bovine viral diarrhea virus, as well as the toxicity to MDBK cells, except for raspberry and cherry juices, that were inactive against Bovine herpesvirus. The extracts worked better in most cases, except sweet cherries on MDBK cells and Bovine viral diarrhea virus, whose juice had stronger effect. The equally effective action of juice and extract was observed in aronia for all three effects and raspberries for toxicity to MDBK cells. In the diffusion test, antibiotic showed a stronger effect than juices and extracts, except for extracts of blackberry, raspberry and sour cherries on Trueperella pyogenes. In a dilution study, the effect of extracts and juices on Gram-positive bacteria was comparable to that of Streptomycin, while the effect of the antibiotic on Gram negative bacteria was stronger than juices and extracts, with exception of the stronger effect of raspberry extract against Escherichia coli. Nystatin has expressed its all antifungal and antialgal effects on fungi and algae included in the study in smaller concentrations compared to the examined juices and extracts. The most consistent with the antibacterial effect of juices and extracts on the appropriate microorganisms were: coumarin, ferulan and gallic acid, catechin. The highest percentage of association with antimicrobial activity of the fruits was given by anthocyanins to Enterococcus faecalis (71.94%). The least related to the antimicrobial activity on the appropriate microorganisms were: chlorogenic acid, champferol, coumarin and saringic acid, vitamin C, vanillic acid, epicatechin, catechin, rutin, chrysin, elaginic and p-hydroxybenzoic acid. The knowledge gained through this dissertation expands the scope of knowledge on antimicrobial activity of fruits and forms the basis for more effective use of fruits in both therapy and nutrition. However, further research in terms of extending the range of examined microorganisms, antimicrobial studies of individual fruits components and mechanisms of action are required.</p>