Karakterizacija i korelacija bioaktivnih fenolnih jedinjenja crvenih vina Balkana i njihova antioksidaciona i antimikrobna svojstva
Characterization and correlation of bioactive phenolic compounds of red wines from the balkans and their antioxidant and antimicrobial properties
Author
Radovanović, Aleksandra N.Mentor
Jovančićević, Branimir
Committee members
Tešević, Vele
Roglić, Goran

Vajs, Vlatka
Metadata
Show full item recordAbstract
U radu su prikazani rezultati ispitivanja fenolnog sastava, antioksidacione i
antimikrobne aktivnosti crvenih vina, dobijenih iz vinogradarskih i vinskih rejona
Balkana (Bosne i Hercegovine, Hrvatske, Srbije, Crne Gore, Makedonije i Bugarske),
proizvedenih u periodu od 2007 do 2011 godine od različitih proizvođača. Izabrana
su crvena vina od internacionalnih: Cabernet Sauvignon, Merlot, Pinot Noir,
Frankovka i Game i autohtonih Vitis vinifera sorti grožđa: Vranac i Kratošija.
Ispitivanje fenolnog sastava izvršeno je spektroskopskim metodama i HPLC-om. Na
taj način identifikovani su jedanaest fenolnih kiselina (galna, vanilinska, siringinska,
hlorogenska, elagilna, trans-kaftarna, trans-kutarna, trans-kafena, p-kumarna i
ferulna kiselina) određivani DAD detektorom na 280 i 320 nm, trans-resveratrol na
320 nm i dvadeset i devet flavonoidna jedinjenja ((+)-katehin, procijanidin B2, (-)-
epikatehin, (-)-epigalokatehingalat, kvercetin-3-glikozid, rutin, miricetin, morin,
kvercetin, kemferol..., luteolin, apigenin i naringin) određivani na 360 nm i 275/322 nm
sa fluorescentnim detektorom i kao veoma značajna flavonoidna grupa antocijani
(malvidin-3-glikozid, peonidin-3-glikozid, delfinidin-3-glikozid, cijanidin-3-glikozid i
petunidin-3-glikozid, njihovi 3-acetilglikozidi i p-kumaroilglikozidi derivati, kao i Vitisin
A i malvidin-3-vinilfenolglikozid) na 520 nm. Rezultati ispitivanja svih analiziranih
uzoraka vina ukazuju da fenolni sadržaji zavise od više faktora (sorte grožđa, mesta
položaja vinograda, agrohemijskih uslova, vremena i godine berbe grožđa,
primenjene tehnologije proizvodnje, načina čuvanja, skladištenja, starenja vina itd).
HPLC analiza je pokazala da je najdominantnija fenolna kiselina galna kiselina
(65,06 - 88,75% od ukupnih fenolnih kiselina), a od flavonoida najdominantniji su:
(+)-katehin (55,30 - 83,13% od ukupnih flavan-3-ola), kvercetin-3-glikozid (57,58 –
60% od ukupnih flavonola) i malvidin-3-glikozid sa njegovim acetil- i p-kumaroil
derivatima (86,47 – 100% od ukupnih antocijana). Ispitivani uzorci vina pokazuju
antioksidativno delovanje na DPPH radikale u opsegu EC50 vrednosti od 47,17 do
145,83 mL/g ili u procentima od 69,55 do 91,83%. Korelacionom analizom je
utvrđeno da postoji dobra korelacija (0,6912 - 0,8595) između antioksidativne
aktivnosti vina i spektroskopskih, odnosno HPLC detektovanih koncentracija nekih
fenolnih jedinjenja (galne kiseline, (+)-katehina, kvercetin-3-glikozida i transresveratrola).
Antioksidaciona aktivnosti vina je analizirana i novom redoks
metodom, upotrebom bakar (II)-1,10-fenatrolina kao oksidanta. Dobijeni rezultati su
u saglasnosti sa rezultatima dobijenim DPPH metodom (0,9775 - 0,9776).
Antimikrobna aktivnost izabranih uzoraka vina je ispitivana disk-difuzionom i mikrodilucionom
metodom u odnosu na šest Gram-pozitivnih i šest Gram-negativnih
bakterija, i kvascu Candida albicans. Dobijeni rezultati za minimalnu inhibitornu
koncentraciju (MIC) i minimalnu bakterijsku koncentraciju (MBC) daju sledeći
redosled vina: Cabernet Sauvignon > Vranac > Merlot > Pinot Noir ~ Frankovka ~
Višesortna Župska vina. Ispitivana vina pokazuju antimikrobnu aktivnost u odnosu na
gram-pozitivne bakterije na sledeći način: Clostridium perfringens > Bacillus subtillis
~ Listeria monocytogenes ~ Sarcina lutea > Staphylococus aureus ~ Micrococcus
flavus. Antimikrobna aktivnost protiv gram-negativnih bakterija je sledeća: Proteus
vulgaris > Pseudomonas aeruginosa ~ Salmonella enteritidis > Klebsiella pneumonia
> Shigella sonnei. Inhibiciona zona delovanja Cabernet Sauvignon vina pokazuju
inhibiciju do 20,0 mm za Escherichia coli i 19,0 mm za Bacillus subtillis, što je
uporedljivo sa inhibicionom zonom standardnih antibiotika (16,0 – 35,0 mm) i nekih
fenolnih jedinjenja (14,5 – 21,8 mm). Korelacionom analizom je potvrđeno da postoji
dobra korelacija između antimikrobnog delovanja prema Escherichia coli i
Staphylococus aureus i nađenih koncentracija za (+)-katehina (0,6625 - 0,7433) i
ukupnih fenolnih kiselina (0,5121 - 0,7333). Urađena analiza antioksidacione i
antimikrobne aktivnosti ispitivanih crvenih vina je pokazala da se najbolje korelacije
uspostavljaju sa ukupnom koncentracijom detektovanih flavonoida, pa se može
zaključiti da je biohemijsko delovanje crvenih vina, posledica sinergestičkog
delovanja većeg broja biološkoaktivnih komponenata. Dobijeni rezultati bi dali
mogućnost za proizvodnju kvalitetnih crvenih vina sa povećanim fenolnim
sadržajem, koji bi uz bolji marketing bili konkurentni na svetskom tržištu.
This paper presents the results of analyzes of the phenolic composition, antioxidant
and antimicrobial activity of red wines, obtained from wine-growing and wine regions
of the Balkans (Bosnia and Herzegovina, Croatia, Serbia, Montenegro, Macedonia
and Bulgaria), and produced from 2007 to 2011 from different wineries. The selected
red wines were international: Cabernet Sauvignon, Merlot, Pinot Noir, Frankovka and
Gamay, and indigenous Vitis vinifera grape varieties: Vranac and Kratošija.
Investigation of the phenolic composition is carried out by spectroscopic methods,
and HPLC. In this way, we have identified eleven phenolic acids (gallic, vanillic,
syringic, chlorogenic, ellagic, trans-caftaric, trans-coutaric, trans-caffeic, p-coumaric
and ferulic acid) with DAD detector at 280 and 320 nm, trans-resveratrol at 320 nm,
and the twenty-nine flavonoid compounds ((+)-catechin, procyanidins B2, (-)-
epicatechin, (-)-epigallocatechin gallate, quercetin-3-glycoside, rutin, myricetin,
morin,... quercetin, kaempferol, luteolin, apigenin and naringin) determined at 360 nm
with DAD and at 275/322 nm with a fluorescence detector. As significant flavonoid
group anthocyanins (malvidin-3-glucoside, peonidin-3-glucoside, delphinidin-3-
glucoside, cyanidin-3-glucoside and petunidin-3-glucoside and their 3-acetyl
glucoside and p-coumaroyl glucoside derivatives, as well as Vitisin A and malvidin-3-
glucoside-4-vinylphenol) were detected at 520 nm. The results of the analyzed wine
samples indicate that the phenolic content depends on several factors (grape variety,
place of vineyard, agrochemical conditions, time and year of harvest, the applied
production technology, methods of preservation, storage, aging wines, etc). HPLC
analysis showed that the dominant phenolic acid is gallic acid (65.06 to 88.75% of
the total phenolic acids), and the most dominant of flavonoids are: (+)-catechin
(55.30 to 83.13% of the total flavan-3-ols), quercetin-3-glucoside (57.58 to 60% of
the total flavonols) and malvidin-3-glucoside of its acetyl- and p-coumaroyl glucoside
derivatives (86.47 to 100% of the total anthocyanins). The investigated samples of
wines show antioxidant activity in the DPPH radicals in the range of EC50 values of
47.17 to 145.83 mL/g and a percentage of 69.55 to 91.83%. Correlation analysis
showed that there is a good correlation (0.6912 to 0.8595) between the antioxidant
activity of wines and spectroscopic, or HPLC detected concentrations of some
phenolic compounds (gallic acid, (+)-catechin, quercetin-3-glycoside and trans –
resveratrol). Antioxidant activity of wines is determinate with a new redox method,
using copper (II) -1,10-phenanthroline as oxidant. The obtained results are in
accordance with the results obtained by DPPH method (0.9775 to 0.9776). The
antimicrobial activities of selected wine samples were studied using the disk-diffusion
and micro-dilution method, according to the six Gram-positive and six Gram-negative
bacteria and the yeast Candida albicans, too. The results of the minimum inhibitory
concentration (MIC) and minimum bacterial concentration (MBC) give the following
order of wines: Cabernet Sauvignon > Vranac > Merlot > Pinot Noir ~ Frankovka ~
Blends from Župa. All tested wines showed antimicrobial activity against Grampositive
bacteria, as follows: Clostridium perfringens > Bacillus subtillis ~ Listeria
monocytogenes ~ Sarcina lutea > Staphylococus aureus ~ Micrococcus flavus.
Antimicrobial activity against Gram-negative bacteria is as follows: Proteus vulgaris >
Pseudomonas aeruginosa ~ Salmonella enteritidis > Klebsiella pneumonia > Shigella
sonnei. The inhibition zone in which the Cabernet Sauvignon wines show inhibition
to 20.0 mm for Escherichia coli and 19.0 mm for Bacillus subtillis, which is
comparable to the standard antibiotic inhibitory zone (16.0 to 35.0 mm), and some of
the phenolic compounds (14, 5 to 21.8 mm). Correlation analysis has confirmed that
there is a good correlation between the antimicrobial activity towards Escherichia
coli, and Staphylococcus aureus and the results for the concentrations of (+)-
catechin (0.6625 to 0.7433) and total phenolic acids (0.5121 to 0.7333). The analysis
of antioxidant and antimicrobial activities of the tested red wines showed that the
best correlation is made with the total concentration of flavonoids detected, and it
can be concluded that the biochemical activity of red wines is connected with the
synergetic action of a number of biologically active components. The results gives
the possibility to produce high-quality red wines with high phenolic content, which
along with better marketing would be competitive in the world market.