Title Sprječavanje genotoksičnog učinka ohratoksina i aflatoksina B1 ekstraktima vrsta roda Echinacea L. i Castanea Mill. Title (english) Prevention of genotoxic effect of ochratoxin and aflatoxin B1 by Echinacea L. and Castanea Mill. extracts Author Huska Jukić Mentor Tomislav Klapec (mentor)Mentor Ivan Kosalec (komentor)Committee member Daniela Čačić Kenjerić (predsjednik povjerenstva)Committee member Tomislav Klapec (član povjerenstva)Committee member Ivan Kosalec (član povjerenstva)Committee member Ivica Strelec (član povjerenstva)Granter Josip Juraj Strossmayer University of Osijek Defense date and country 2012-03-16, Croatia Scientific / art field, BIOTECHNICAL SCIENCES Universal decimal classificationUDC ) 615 - Pharmacology. Therapeutics. Toxicology Abstract Cilj ovoga rada bio je ispitati sadržaj polifenola u ekstraktima vrsta Echinacea purpurea (L.) Moench kao i Castanea sativa Mill. te odrediti njihov antioksidativni i antigenotoksični učinak. Antioksidativna aktivnost ekstrakata i glavnih sastavnica je utvrđena primjenom elektronske spinske rezonancije i spektrofotometrijskih metoda hvatanja DPPH, superoksidnog i hidroksilnog radikala. Ispitan je i redoks učinak i citotoksičnost ekstrakata u kulturi stanica raka kolona (SW480).
Antigenotoksični i antimutageni učinak procijenjen je kometnim i Amesovim testom uz mutagene i genotoksične mikotoksine – aflatoksin B1 (AFB1) i ohratoksin A (OTA). Provedena je i senzorska analiza tretiranih uzoraka kikirikija i grožđica za preliminarnu procjenu primjenjivosti tretmana ekstraktima. Ekstrakt herbe ehinaceje je sadržavao više ukupnih polifenola u odnosu na ekstrakt lista kestena (10,57 % prema 6,74 %). Ekstrakt ehinaceje je sadržavao višestruko veću količinu fenolnih
kiselina (prvenstveno cikorijska i kaftarinska kiselina), dok je ekstrakt kestena bio najbogatiji taninima i flavonoidima (derivati galne i elaginske kiseline). Ekstrakt kestena je pokazao znatno veću sposobnost neutralizacije DPPH radikala (EC50 = 1,87 μg/mL) u odnosu na ekstrakt ehinaceje (EC50 = 15,67 μg/mL). Slični rezultati utvrđeni i ispitivanjem hvatanja hidroksilnog i superoksidnog radikala, pri čemu je ekstrakt kestena bio desetak puta učinkovitiji. Utvrđena je i snažna antioksidativna
aktivnost dominantnih sastavnica koja ukazuje na fenolne kiseline (ehinaceja) i elaginsku kiselinu (kesten) kao sastojke koji najviše doprinose učinku ekstrakata. Inhibitorni učinak ekstrakata ehinaceje na mutageno djelovanje aflatoksina B1 zabilježeno je samo kod najniže koncentracije na soju bakterije Salmonella typhimurium osjetljivom na pomak okvira čitanja. S druge strane, utvrđen je snažan inhibitoran učinak ekstrakta kestena, izraženiji na soju osjetljivom na pomak okvira čitanja. Supresija
genotoksičnog učinka AFB1 i OTA utvrđena je i kometnim testom, pri čemu je ustanovljena značajna redukcija dužine repa, repnog intenziteta i repnog momenta u leukocitima istovremeno tretiranim mikotoksinima i različitim koncentracijama ekstrakata. Dok ekstrakt ehinaceje tek pri dužoj inkubaciji i višim koncentracijama djeluje citotoksično na stanice raka kolona, ekstrakt kestena je pokazao znatno snažniji učinak već kod najniže koncentracije. Ispitivanjem supresije oksidativnog stresa
ekstraktima u kulturi stanica raka zabilježeno je podsticanje oksidativnog stresa kod nižih koncentracija i redukcija pri višim koncentracijama, a slično djelovanje je ustanovljeno i u stanicama tretiranim vodikovim peroksidom. Rezultati upućuju na indukciju antioksidantne zaštite stanica. Senzorska analiza tretiranih kikirikija je utvrdila koncentraciju ekstrakta koja nije izazvala značajnije promjene organoleptike u odnosu na kontrolni uzorak. Tretirane grožđice su bile promijenjenog mirisa i arome, iako bez utjecaja na prihvatljivost samog proizvoda. Dodatak ovdje ispitanih ekstrakata potencijalnog antigenotoksičnog učinka hrani bi, dakle, pored uloge u povećanju održivosti samih namirnica antioksidantnim djelovanjem, mogao modulirati toksičnost prisutnih mikotoksina nakon konzumacije.
Abstract (english) The purpose of this thesis is to examine polyphenol content in Echinacea purpurea (L.) Moench and
Castanea sativa Mill. extracts, and to determine antioxidant effects of those extracts, trough electron
spin resonance and colorimetric method of DPPH•; a superoxide hydroxyl radical. Redox effect and
cytotoxicity of the extract in colon cancer cell culture SW480 has also been tested. Antigenotoxic
antimutagenic effect has been evaluated using comet and Ames test, with mutagenic and genotoxic
micotoxines AFB1 and OTA present. Sensory analysis of peanut and raisin samples has also been
performed for the preliminary evaluation of extract treatment. Echinacea herb extract contained higher
total polyphenol content than chesnutt leaves extract (10,57% compared to 6,74%). Echinacea herb
extract had significantly higher phenolic acid content (cicoric and caftarinic acid), while chestnut
extract was rich in tanine and flavonide compounds. Chestnut extract had higher capability to
neutralize DPPH radicals (EC50 = 1,87 μg/mL) compared to Echinacea extract (EC50 = 15,67 μg/mL).
Similar results were obtained when hydroxyl and superoxide radical capture capability was examined;
chestnut extract was about 10 times more effective. Strong antioxidative activity of dominant
compounds has been established, which points out to presence of phenolic acids and elaginic acid in
Echinacea and chestnut respectively, as well as other compounds which contribute to the extract
effect. Inhibitory effect of Echinacea extract on mutagenic effect of aflatoxin B1 has been recorded at
the lowest concentrations, when Salmonella typhimurium bacteria has been treated. On the other hand
there was a strong inhibitory effect demonstrated by the chestnut extract, once it was applied to the
appropriate bacteria culture. The suppression of genotoxic effect of AFB1 and OTA has been
determined using the comet test, whereby significant reduction of the tail length has been determined,
as well as tail moment and intensity in leukocytes treated by mycotoxines and different concentrations
of extracts simultane ously. Chestnut extract has shown strong effects even at low concentrations,
while Echinacea extract had cytotoxic effect on colon cancer cells only after prolonged exposure at
high concentrations. When suppression of the oxidative stress trough extract application has been
examined, it has been established that there is oxidative stress relation with lower concentrations,
while reduction has been established at high concentrations; similar effects have been established once
cells were treated with hydrogen peroxide. The results suggest induction of antioxidative protection of
the cell. Sensory analysis of treated peanuts has established the concentration with no significant
effects on organoleptics compared with the control sample. Treated raisins had a change in aroma and
scent, but without substantial effect on the consumability of the product itself. These additives not only would extend the useful life of various food products; they can also serve as a modulator for toxicity of various micotoxins after those food products have been consumed.
Keywords
polifenoli
antioksidacijska aktivnost
kikiriki
ekstrakt
genotoksičnost
Keywords (english)
polyphenols
antioxidant activity
genotoxicity
extract
peanut
Language croatian URN:NBN urn:nbn:hr:109:362347 Study programme Title: Food Engineering Type of resource Text Extent 217 str.: ilustr., graf. prikazi: 30 cm File origin Born digital Access conditions Access restricted to students and staff of home institution Terms of use Created on 2016-01-26 12:45:48
