Submitted by Maike Costa (maiksebas@gmail.com) on 2017-03-06T11:14:23Z No. of bitstreams: 1 arquivo total.pdf: 786129 bytes, checksum: 1e08eca0cb38b5c895cff1f2cbb20f5c (MD5) / Made available in DSpace on 2017-03-06T11:14:24Z (GMT). No. of bitstreams: 1 arquivo total.pdf: 786129 bytes, checksum: 1e08eca0cb38b5c895cff1f2cbb20f5c (MD5) Previous issue date: 2016-12-13 / Oral candidiasis is a fungal infection caused by yeasts of the genus Candida, with C. albicans being the main pathogen. The emergence of resistant species against the available therapeutic arsenal as well as its adverse effects has driven new research on natural products. In view of this problem, the present study sought to investigate the antifungal activity of essential oil (OE) obtained from the leaves of Melissa officinalis L (lemon balm), isolated and associated with antifungal agents licensed on C. albicans strains. Two strains of the American Type Culture Collection (ATCC) and eight from the oral cavity (LM) were selected. The microdilution and sowing of subcultures were used to determine, respectively, the Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (CFM). The antifungigram was performed using the disc diffusion technique with the antifungal agents nystatin, amphotericin B, miconazole and fluconazole, evaluating them in isolation and associated with OE. MIC and MFC ranged from 64 to 128 μg/mL and 256 to 512 μg/mL, respectively. The presence of resistance of the strains to the antifungal agents nistatin (LM-38 and LM-65), amphotericin B (ATCC-76485, LM-3A, LM-38, LM-62 and LM -65) and fluconazole (LM-1A, LM- 1B, LM-38, LM-62 and LM-65). In the association tests, the results showed the influence of the essential oil on the antifungal activity, with synergism (S), indifference (I) and antagonism (A) being observed. The percentages found for each antifungal in the order (S, I and A) were: nystatin (30%, 30% , 40%), amphotericin B (30%, 50%, 20%), fluconazole (20% ,40% , 40%) and miconazole (100%, 0,0). In view of the above, it is concluded that the OO of M. Officinalis may become a promising product in the fight against oral candidiasis, since it has a strong antifungal activity against Candida albicans and that the use of this product associated with the licensed antifungal can, in some situations, potentiate its effectiveness of use in treatment. / A candidíase oral é uma infecção fungica causada por leveduras do gênero Candida, sendo a espécie C. albicans apontada como principal patógeno. O surgimento de espécies resistentes frente ao arsenal terapêutico disponível assim como seus efeitos adversos tem impulsionado novas pesquisas voltadas para os produtos naturais. Diante dessa problemática o presente trabalho buscou investigar a atividade antifúngica do óleo essencial (OE) obtido das folhas de Melissa officinalis L (erva-cidreira), isolado e associado a antifúngicos licenciados sobre cepas de C. albicans. Para tanto, foram selecionadas duas cepas da American Type Culture Collection (ATCC) e oito provenientes de cavidade oral (LM). Utilizou-se a técnica da microdiluição e semeadura de subcultivos para determinar, respectivamente, a Concentração Inibitória Mínima (CIM) e a Concentração Fungicida Mínima (CFM). O antifungigrama foi realizado a partir da técnica de difusão de disco com os antifúngicos nistatina, anfotericina B, miconazol e fluconazol avaliando-os de forma isolada e associada ao OE. A CIM e CFM variaram, respectivamente, de 64 a 128 μg/mL e 256 a 512 μg/mL. Quanto ao teste de sensibilidade de forma isolada observou-se a presença de resistência das cepas aos antifúngicos nistatina (LM-38 e LM-65), anfotericina B (ATCC- 76485, LM-3A, LM-38, LM-62 e LM-65) e fluconazol (LM-1A, LM-1B, LM-38, LM- 62 e LM-65). Nos ensaios de associação os resultados mostraram influência do óleo essencial sobre a atividade dos antifúngicos sendo observado sinergismo (S), indiferença (I) e antagonismo (A). As porcentagens encontradas para cada antifúngico, na ordem (S, I e A), foram: nistatina (30%, 30% e 40%), anfotericina B (30%, 50%, 20%), fluconazol (20%, 40% e 40%) e miconazol (100%,0,0). Perante o exposto conclui-se que o OE de M. Officinalis pode se tornar um produto promissor no combate a candidose oral, visto que apresenta forte atividade antifungica contra Candida albicans e que o uso desse produto associado à antifungicos licenciados pode, em algumas situações, potencializar sua efetividade de uso no tratamento.

<p>Cilj rada je bio ispitivanje funkcionalnih karakteristika kombuha napitka<br />od melise (<em>Melissa officinalis</em> L.). Antibakterijska aktivnost kombuha<br />napitaka optimalne konzumne i većih kiselosti ispitana je prema<br />bakterijama izolovanim iz hrane i vode za piće. Nosilac antimikrobne<br />aktivnosti je sirćetna kiselina, a na ostale nosioce ukazuje delovanje<br />neutralisane kombuhe i čajnog napitka prema nekim test bakterijama.<br />Spektrofotometrijskom metodom određen je sadržaj ukupnih fenolnih<br />jedinjenja, a HPLC analizom određen je kvalitativni i kvantitativni sastav<br />fenolnih jedinjenja u fermentacionim tečnostima, čajnim i kombuha<br />napicima od melise i crnog čaja. Antioksidativna aktivnost istih uzoraka<br />ispitana je na DPPH i OH radikale ESR spektralnom metodom. Uzorci<br />fermentacione tečnosti i kombuha napitka od melise imali su veću<br />antioksidativnu aktivnost prema oba radikala u odnosu na uzorke sa<br />crnim čajem. Konzumna kombuha od melise imala je veću<br />antioksidativnu aktivnost od čajnog napitka. Aktivne komponente<br />kombuha napitka od melise su verovatno ruzmarinska kiselina i<br />kvercetin. U ispitivanju antiproliferativne aktivnosti konzumnog<br />kombuha napitka i čajnog napitka od melise na tri ćelijske linije humanih<br />karcinoma: HeLa (epitelni karcinom grlića materice), MCF-7<br />(adenokarcinom dojke) i HT-29 (adenokarcinom debelog creva) utvrđeno<br />je da nije do&scaron;lo do stimulacije proliferacije ispitanih ćelijskih linija pri<br />koncentracijama većim od 100 &mu;g/ml. Pored istraživanja biolo&scaron;ke<br />aktivnosti ispitana je mogućnost simultane mlečno-kiselinske i kombuha<br />fermentacije. Dodatkom starter kultura i <em>Lactobacillus spp</em>. izolata u<br />fermentacionu tečnost dolazi do povećanja sadržaja L- i D- mlečne<br />kiseline, iako su ćelije bakterija mlečne kiseline, osim izolata iz kiselog<br />testa (<em>L. hilgardii</em>), pokazale malu otpornost na uslove tokom<br />fermentacije i čuvanja pripremljenih napitaka. Izvr&scaron;ena je i identifikacija<br />bakterija sirćetnog vrenja izolovanih iz lokalnih čajnih gljiva PCR<br />metodom. Dva izolata verovatno pripadaju vrsti <em>Gluconobacter oxydans</em>,<br />a treći vrsti <em>Gluconacetobacter hansenii</em>.</p> / <p>The aim of this study was to investigate functional characteristics of akombucha beverage from lemon balm (<em>Melissa officinalis</em> L.) tea. Antibacterial activity of kombucha beverages with optimum and higheracidities was tested against bacteria isolated from food and drinking water. The main active component of antibacterial activity was acetic acid, while slight activity of neutralized kombucha and unfermented tea against some test bacteria indicated presence of other antibacterial components. Total phenol concentration in unfermented tea samples, fermentation broths and kombucha beverages from lemon balm and black tea was determined spectrophotometrically whereas qualitative and quantitative concentration of polyphenolic compounds was determined by HPLC method. Antioxidant activity on DPPH and hydroxyl radical in the same samples was determined on an ESR spectrometer. Fermentation broth and kombucha beverage from lemon balm had higher antioxidant activity against both radicals than the samples from black tea. Kombucha beverage from lemon balm with optimum acidity had higher antioxidant activity than unfermented lemon balm tea. The main active components of antioxidant activity were probably rosmarinic acid and quercetin. Antiproliferative activity of lemon balm tea and kombucha was measured by sulforhodamine B colorimetric assay on HeLa (cervix epitheloid carcinoma), HT-29 (colon adenocarcinoma), and MCF-7 (breast adenocarcinoma) cell lines. By applying concentrations higher than 100 &mu;g/ml, tested samples did not stimulate proliferation of cell lines. The possibility of simultaneous lactic acid and kombucha fermentation was tested as well. When starter cultures and Lactobacillus spp. isolates were applied, the content of Land D- lactic acid increased during fermentation, although lactic acid bacteria (except <em>L. hilgardii</em> isolated from sour dough) showed low resistance to the conditions during fermentation and beverages storage. Acetic acid bacteria isolated from local tea fungus were identified by PCR method. Two isolates might be <em>Gluconobacter oxydans</em> and one - <em>Gluconacetobacter hansenii.</em></p>

<p>Cilj ispitivanja čiji su rezultati prikazani u ovoj doktorskoj disertaciji bio<br />je da se ispita mogućnost dobijanja kombuha napitka od/sa CoffeeBerry&reg;<br />ekstraktom i da se ispitaju njegove funkcionalne karakteristike.<br />CoffeeBerry&reg; ekstrakt kao bogat izvor biolo&scaron;ki aktivnih jedinjenja bi<br />obezbedio dodatne funkcionalne karakteristike kombuha napitku u<br />poređenju sa onim pripremljenim na tradicionalan način od zaslađenog<br />crnog čaja. U disertaciji je nakon optimizacije sastava podloge za<br />kultivaciju čajne gljive ispitana (in vitro) antioksidativna, antimikrobna i<br />citotoksična aktivnost kombuhe/fermentacione tečnosti obogaćene<br />CoffeeBerry&reg; ekstraktom. Antioksidativna aktivnost je ispitana ESR<br />(elektron-spin rezonantnom) spektroskopijom na reaktivne hidroksil i<br />stabilne DPPH (1,1-difenil-2-pikrilhidrazil) radikale, kao i kvalitativni i<br />kvantitativni sastav fenolnih jedinjenja, potencijalnih nosilaca biolo&scaron;ke<br />aktivnosti (HPLC metodom). Antimikrobna aktivnost je ispitana agardifuzionom<br />metodom na odabrane referentne i izolovane (&bdquo;divlje&ldquo;)<br />sojeve bakterija (Gram-pozitivnih i Gram-negativnih) i kvasaca.<br />Citotoksična aktivnost uzoraka na rast odabranih ćelijskih linija: Hep2c<br />(Human larynx carcinom), RD (Rhabdomyosarcoma) i L2OB (mi&scaron;ija<br />tumorska fibrolastna linija u koju su transfektovani neki humani geni)<br />ispitana je MTT testom. U navedenim ispitivanjima je tradicionalna<br />kombuha dobijena od zaslađenog crnog čaja kori&scaron;ćena kao kontrolni<br />uzorak. Rezultati su pokazali da CoffeeBerry&reg; ekstrakt značajno<br />doprinosi antioksidativnoj i citotoksičnoj aktivnosti napitka &scaron;to govori o<br />punoj opravdanosti upotrebe ovog ekstrakta kao funkcionalnog dodatka<br />podlozi za kultivaciju čajne gljive. Kombuha napici od crnog čaja i sa<br />dodatkom CoffeeBerry&reg; ekstrakta pokazali su približno isto delovanje na<br />bakterijske sojeve, dok je delovanje na kvasce izostalo u potpunosti.<br />Primarni nosilac antibakterijske aktivnosti napitaka je sirćetna kiselina.</p> / <p>The aim of this study was to investigate fermentation of sweetened<br />medium prepared with CoffeeBerry&reg; еxtract and functional<br />characteristics of a kombucha beverage enriched with CoffeeBerry&reg;<br />еxtract. Total phenol concentration in unfermented samples,<br />fermentation broths and kombucha beverages made of CoffeeBerry&reg;<br />еxtract and black tea was determined spectrophotometrically whereas<br />qualitative and quantitative concentration of polyphenolic compounds<br />was determined by HPLC method. Antioxidant activity on DPPH and<br />hydroxyl radicals in the same samples was determined on an ESR<br />spectrometer. Fermentation broth and kombucha beverage enriched with<br />CoffeeBerry&reg; еxtract had higher antioxidant activity against both<br />radicals than control samples. Antimicrobial activity of kombucha<br />beverages optimal acidity was tested against selected strains of bacteria<br />(Gram-positive and Gram-negative) and yeast. The main active<br />component of antibacterial activity was acetic acid, but samples did not<br />show any activity against yeast Candida albicans and Saccharomyces<br />cerevisiae. Antiproliferative activity of fermentation broth with and<br />without CoffeeBerry&reg; еxtract was measured by MTT test on following<br />cell lines: Hep2c (Human larynx carcinom), RD (Rhabdomyosarcoma) i<br />L2OB (murine tumor fibrolast lines trаnsfеcted by some human genes).<br />The highest antiproliferative activity shown fermentation broth enriched<br />with CoffeeBerry&reg; еxtract, wherein Hep2c cells was the most<br />susceptible.</p>