UTICAJ BAKRA I CINKA U SIRI NA SADRZAJ VISIH ALKOHOLA U VINU
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1 APTEFF, 31, (2) UDC : : BIBLlD: , (2) 31, pp UTICAJ BAKRA I CINKA U SIRI NA SADRZAJ VISIH ALKOHOLA U VINU Nadeida E. Ruiic U radu su prikazani rezultati ispitivanja uticaja povecanih kolicina bakra i cinka u siri, kao i selekcionisanin sojeva kvasca upotrebljenihza [ermentaciju, na kolicinu visin alkohola u vinu. Bakar prisutan u siri u kolicini od II mgjdm' negativno je uticao na sintezu visih alkohola, bez obzira na vrstu i soj upotrebljenog vinskog kvasca za [ermentaciju. Negativan utica] se vise ispoljio kod sira sa niskom ph vrednoscu (ph-3,2) i kod uzoraka koje je [ermentisao kvasac Saccharomyces cerevisiae - soj MO Uproizvedenim vinima je konstatovano 12,3 do 35,3% manje visih alkohola nego u kontrolnim vinima. Cink prisutanu siriu koliciniad 24 mgldm' slimulisa je sintezu visilialkohola lokom alkohalne [ermentacije. Utica] cinka je bio izrazitijiu uzorcima.~ire sa visom ph vrednoscu. i u onim kaje je [ermentisao kvasac Saccharomyces bayanus - soj 5-4. U vinima proizvedenim od sira obogacenih cinkom konstatovano je 11,8 do 18,9% vise visih alkohola nego u kontrolnim vinima. KUUCNE REC1: sira, bakat; cink, kvasac, vino, viii alkoholi UYOD Veci deo visih alkohola u vinu nastaje tokom alkoholne fermentacije kao rezultat aktivnosti kvasaca(1). Neki od njih su veoma prijatnog mirisa i pozitivno uticu na aramu i buke vina. Irna, medutim, i takvih (n-butilalkohol, izoamilalkohol) koji pogorsavaju senzorna svojstva vina (2, 3,4). Hemijski sastav sire, kvasac koji obavlja fcrrnentaciju, kao i uslovi pod kojima se ona odvija, uticu na kolicinu visih alkohola u proizvedenom vinu. Prema literaturnim podacima, putevi sinteze visih alkohola tokom fermentacije su razliciti. Nastaju dezaminacijom i dckarboksilacijom aminokiselina, ali vecim delom transformacijom ugljenih hidrata, tacnijc produkata koji od njih nastaju kao metaboliti kvasca (4,6,7,8). Kada u siri nema dovoljno azota, zapravo aminokiselina, nagomilavaju se ketokiseline a ove clekarboksilacijom i redukcijom prevocle clovisih alkohola. Ukoliko su aminokiseline u visku transforrnacija do visih alkohola sc odvija njihovom dezaminacijom i clekarboksilacijom (5,9) Bela vina sadrze 15-4 mg/drn' visih alkohola (4), nasa vina u proseku 325 mg/drn' (1), a crna vina ocl3 do 6 mg/dm". Najvisc ima izobutil alkohola (cini 1/3 ad svih visih alkohola dr Nadczda E. Ruzic, rcdovan profcsor, Univcrzitct u NOVOlll Sadu, Tchnoloski fakultct, 21 Novi Sad, Bulcvar Cara Lazara I, Jugoslavija, 239
2 kod belih i 1/4 kod crnih vina) i izoamil alkohola (1). Poznato je da kolicina visih alkohola u vinu zavisi od hemijskog sastava sire, njene pripreme pre alkoholne fermentacije, prisustva pokozice (11, 12), temperature fermentacije, vinskog kvasca (4, 5). Zainteresovalo nas je zbog toga koliko prisustvo povecanih kolicina bakra i cinka u siri utice na sintezu visih alkohola tokom fermentacije, pogotovo sto oba katjona imaju znacajnu ulogu kao metaloaktivatori biohemijskih procesa.(13). Bakar i cink u siri i vinu su, inace, fizioloskog i tehnoloskog porekla. U grozde dospevaju preko korenovog sistema vinove loze, u siru preko sredstava za zastitu vinove loze, preko oprcme u vinskom podrumu, prcko tehnoloske vode i cnoloskih sredstava. EKSPERIMENTALNI DEO Ogledi su izvedeni sa pasterizovanom sirom koja je u tabeli 1oznacena kao sira II. Sira je podeljena na tri dela pa je ph u prvoj trecini podesen na zeljenu vrednost rastvorom vinske kiseline (sira I), a u drugoj trecini rastvorom kalijumhidroksida (sira III). Sadrzaj secera u sirama je korigovan dodatkom saharoze. Tako su dobijena tri uzorka sire ciji je sastav prikazan u tabeli 1. Nakon toga sirama je dodata odredcna kolicina bakra iii cinka (u obliku acetata) tako da jc sadrzaj bakra povecan za 1 mg/drri', a cinka za 2 mg/dm'. Naznacene kolicine bakra i cinka u uzorcima sire su i do 1 puta vece od onih koje sc mogu naci na grozdu i u siri. Tabela 1. Sastav sire Sira I Sira II* Sira III Secer, % 18,6 18,8 18,3 Ukupna kiselost, g/dm' 8,2 5,9 4,4 ph 3,2 3,2 3,8 Bakar, rng/dm',98 1,3,84 Cink, mg/dm' 3,92 3,98 3,6 * sira sa kojom su postavljeni ogledi Analiza sire je obavljena uobicajenim metodama (14). Bakar i cink u sir: odredcni su potenciometrijskom striping analizom (15). Uzorci sire su zasejani sledecim aktivnim pojedinacnim kulturama selekcionisanog vinskog kvasca (kolekcija Tehnoloskog fakulteta u Novom Sadu): Saccharomyces cerevisiae - sojevi 25-C i MO i Saccharomyces bayanus - sojevi S-4 i V-2. Zasejavanje je obavljeno tako da je u jednom mililitru sire bilo oko 3 x 1f> celija kvasca. Broj celija je odreden brojanjem u Thomovoj komori (16). Fermentacijaje obavljena na temperaturi od 2 llc u bocama od Idrn', zatvorcnim hidraulickim vcntilima. Tokom alkoholne Iermentacije, u odredenim vremcnskim intervalima, u uzorcima sire je odredivan sadrzaj visih alkohola. Pracen je i tok alkoholnc fermentacije svakodnevnim merenjem mase uzoraka sire (prikazan je kao kolicina CO? izdvojena u funkciji vremena). Visi alkoholi u siri su tokom alkoholne fermentacije odredivani spektrofotometrijskom metodom (17), au vinu spektrofotometrijskom i metodom gasnc hromatografijc. Uslovi rada gasnc hromatografije su bili sledeci: 24
3 aparat za gasnu hromatografiju: VARIAN, model 144, gas nosac i protok: azot - 25 ml/rnin, vodonik - 24 rnl/min i vazduh - 2 ml/min, kolona: Hallcomid M-18, duzine 4 m i promera 1/8 inca, uslovi rada: 1-11 x 4, kretanje papira 1 em/min, temperatura kolone: 95 C, temperatura injektora: 15 C, temperatura detektora: 18 C unutrasnji standard: 1-pentanol, kolicina ubrizganog uzorka(destilata):,5 mikrolitara, detektor: plameno jonizacioni (FID) a temperaturni program izoterman. U radu su prikazani ukupni visi alkoholi (iako su gasnom hromatografijom utvrdeni i pojedinacni), posto su kao takvi odredivani tokom alkoholne fermentacije spcktrofotometrijskom metodom. Kolicine visih alkohola u vinu prikazane su kao srednje vrednosti dobijenih rezultata i jednom i drugom metodom analize. Uticaj ispitivanih teskih metala na sintezu visih alkohola tokom fermentacije, proeenjeno je, rnoguce je posmatrati zasebno od rezultata analize istih sastojaka u proizvedenim vinima. REZULTATI I DISKUSIJA Alkoholna fermentacija uzoraka sire kojima je dodat bakar bila je slabijeg intenziteta nego kontrolnih sira, sto je i logicno jer je bakar u vecim koneentraeijama toksican za kvasae (18,19). Na slid 1 prikazani su rezultati pracenja intenziteta fermentacije kontrolnih uzoraka sire i uzoraka kojima je dodat bakar u uslovima koji su bili najpovoljniji za kvasae (ph sire-3,8). Vecina uzoraka otpocela je, kako pokazuju rezultati, da ferrnentise u toku prvih 24 sata po zasejavanju kvasca. Izuzetak su sire sa povecanim sadrzajern bakra zasejane sojem 25-C. One su otpocele da fermentisu sa izvesnim zakasnjenjem.. Prikazani rezultati pokazuju takode, da je od upotrebljenih kvasaca, soj S-4 pokazivao vecu rezistentnost prema povecanoj kolicini bakra od ostalih ispitivanih sojeva. Cink dodat siri nije ometao alkoholnu fermentaeiju. Ona je tekla istim intcnzitetom kao kod kontrolnih uzoraka. Zbog toga nije ni prikazana. ~ 2S-C, kontrola -.-2S-C, + bakar - - MO, kontrola ~~ MO, + bakar ~ 5-4, kontrola , + bakar -t-v-2, kontrola --V-2, + bakar danl Slika 1. Uticaj bakra na dinamiku alkoholne fermentacije sire (ph sire=3,8) 241
4 Bakar i cink, dodati oglednirn uzorcirna sire, uticali su i na sintezu visih alkohola tokorn alkoholne ferrnentacije. Uticaj bakra je prikazan sarno za uzorke sire zasejane kvascern Saccharomyces cerevisiae -soj MO (slika 2) i kvascern Saccharomyces hayanus-soj S-4 (slika 3). C') 25 S 2 " t;n S 15 '..c 1 ~ '"@ 5 'Vi.s;: -+- ph -3,, kontrola ~ ph -3,2, kontrola - - ph -3,8, kontrola --ph -3,, + bakar ~ ph -3,2, + bakar -.- ph -3,8, + bakar dani Slika 2. Uticaj bakra na sintezu visih alkohola tokorn ferrncntacije sire zasejane kvascem Saccharomyces cerevisiae - soj MO (bakar dodat u kolicini od 1 mg/drn') 16 M 14 E " ph-3,, kontrola -!Ii- ph-3,2, kontrola On 1 E - - ph-3,8, kontrola 8 -,.-ph-3,, +bakar "..c: 6 ~ ph-3,2, +bakar..::.:: co >cn 4.;;: dani -.- ph-3,8, +bakar Slika3. Uticaj bakra na dinamiku stvaranja visih alkohola tokom ferrnentacije sire zasejane kvascern Saccharomyces bsyunus - soj S-4 (bakar dodat u kolicini od 1 mg/dm') U uzorcima sire sa povecanom kolicinorn bakra sinteza visih alkohola je tekla sporije nego kod kontrolnih sira kojima bakar nije dodavan. Najintenzivnija je bila u pocetku fermentacije (drugog iii treceg dana), sto je zavisilo od soja kvasca kojim je zasejana sira. Na sintezu je bitno uticala i ph vrednost sire. U uzorcima sa nizom vrednoscu ph sintetizovano je manje visih alkohola nego u onim sa visom ph vrednoscu. Uticaj cinka na sintezu visih alkohola tokom alkoholne ferrnentacije prikazan je na slici 4 samoza kvasac Saccharomyces cerevisiae -soj MO posto su kod drugih ispitivanihsojeva dobijeni slicni rezultati. 242
5 C') S "Cl bo S (3..c:..><: w N ro ;cn...;; -+- ph-3,o, kontrola --{'M- ph-3,2, kontrola - - ph-3,8, kontrola --PH-3,O, + cink...-ph-3,2, + cink ph-3,8, + cink dani Stika 3. Uticaj cinka na dinamiku stvaranja visih alkohola tokom fermentacije sire zasejane kvascem Saccharomyces cerevisiae - soj MO (cink dodat u kolicini od 2 mg/drn ') U prisustvu vecih kolicina cinka u siri, tokom fermentacijc se sintetizovalo vise visih alkohola, sto se moze objasniti povecanjern intenziteta metabolizma kvasnih cclija u takvim uslovima. Ocigledan je i uticaj ph sire. Nairne, u sirama sa nizorn vrednoscu ph sintctizovano je manje visih alkohola nego u sirama sa visom ph vrednoscu, Rezultati koje smo dobili analizom visih alkohola u proizvcdenim vinirna prikazani su u tabelama 2 i 3. Tabela 2. Uticaj bakra I dodatog siri na sadrzaj visih alkohola u vinu Vinski kvasac ph sire Visi alkoholi, mg/drn' oznaka soja Kontrola Uzorak sa dodatim bakrom* 25-C , ,'2 13, prosek" 13L9±12.4% 19.l±14.%l MO prosek I86.9±8.8% 141.7±17.2% S prosek I39.3±7.5% 12,3±8,7% V , prosek 18,2±13,2% 89,1±14,3% I -bakar dodat siri u kolicini od 1 mg/drn' 2 -prosek (bez obzira na ph sire) i srednje odstupanje od proseka izrazeno u procentima 243
6 Tabela 3, Uticaj cinka' dodatog siri na sadrzaj visih alkohola u vinu Vinski kvasac ph sire Visi alkoholi, mg/drn' oznaka soja Kontrola Uzorak sa dodatim cinkom* 25-C 3,2 18,3 114,4 3,2 13,9 142,5 3,8 156,5 185,2 prosek" 131,9±12,4% 147,4±2,5% MO 3,2 162,4 18, 3,2 192, 228,3 3,8 26,4 252,4 prosek 186,9±8,8% 22,2± 12,2% S-4 3, ,8 3,2 137, 164, 3,8 155, 18,4 prosek 139,3±7,5% 161,4±8,9% V-2 3,2 86,8 118,5 3,2 99,4 148, 3,8 128,2 188,8 prosek 14,8±14,9% 151,8±16,3% 1_ cink dodat siri u kolicini od 2 mg/drn' 2 _ prosek (bez obzira na ph sire) i srcdnje odstupanjc od proseka izrazeno u procentima Sadrzaj visih alkohola u proizvedenim vinima varijanti kojima je dodat bakar manji je nego u kontrolnim vinima za 12,3 do 35,3%, zavisno od ph vina i soja upotrebljenog kvasca za fermentaciju (tabela 2). Vece razlike su konstatovane kod uzoraka cija je vrednost ph 3,2 (16,6 do 35,3%), a manje kod onih sa ph 3,8 (12,3 do 25%). Najmanji uticaj bakra i ph sire zabelezen je kod uzoraka koje je fermentisao kvasac Saccharomyces bayanus -soj 5-4 (sintetizovano je 12,3 do 16,6% manje visih alkohola nego u kontrolnim uzorcima), a najveci kod uzoraka koje je fermentisao kvasac Saccharomyces cerevisiae soj MO (15-35% manje visih alkohola u odnosu na kontrolne uzorke). U vinima proizvcdenim od sira sa povecanom kolicinom cinka konstatovano je vise visih alkohola nego u kontrolnim vinima (tabela 3). Cink jc bitan za aktivnost dekarboksilaza (13) koje ucestvuju i u dekarboksilaciji ketokiselina na putu sinteze visih alkohola. Verovatno je to i razlog intenzivnije sinteze ovih jedinjenja u sirama sa povecanorn kolicinorn cinka. U vinima proizvcdenim pomocu kvasca Saccharomyces bayanus - soja S-4 uticaj cinka je najvise izrazen (prosecno je sintetizovano 18,9% vise visih alkoholau odnosu na kontrolna vina, u ostalim vinima 11,8 do 17,8%vise). Kod istih vina je konstatovan i veci uticaj ph na sintezu visih alkohola. Najvise visih alkohola, inace, ustanovljeno je u vinima sa ph-3,8, koja je za kvasac najbliza optimalnoj. Uticaj cinka se nesto slabije ispoljio u vinima proizvedenim pomocu kvasca Saccharomyces cerevisiae - soja 25-C (sadrzaj visih alkohola je veci za 5,6 do 18,3% u odnosu na kontrolna vina). Generalno gledajuci, dobijeni rezultati se slazu sa konstatacijom da faktori koji uticu na povecanje razmene materija u celiji vinskog kvasca, a pod tim faktorima se podrazumeva i visa ph vrednost sire kao i manje prisustvo toksicnih metala, sto se u ovom slucaju odnosi na bakar, stimulisu sintezu visih alkohola tokom fermentacije (9,5). 244
7 ZAKLJUCAK Bakar prisutan u siri u kolicini od 11 mg/dm" uticao je negativno na sintezu visih alkohola tokom alkoholne fermentacije sire, bez obzira na vrstu i soj upotrebljenog vinskog kvasca za fermentaciju (sintetizovano je 12,3 do 35,3% manje visih alkohola u odnosu na kontrona vina). Negativan uticaj se vise ispoljio kod sira sa niskom ph vrednoscu (ph-3,2) i kod uzoraka koje je fermentisao kvasac Saccharomyces cerevisia - soj MO. Uticaj bakra na sintezu visih alkohola bio je najmanji kada je fermentaciju obavljao kvasac Saccharomyces bayanus - soj S-4.. Cink prisutan u siri u kolicini od 24 mg/dm" stimulisao je sintezu visih alkohola tokom fermentacije (sintetizovano je 11,8 do 18,9% vise visih alkohola nego u kontrolnim vinima). Uticaj cinka je najizrazitiji u vinima sa visorn ph vrednoscu i u onim koje je fermentisao kvasac Saccharomyces bayanus - soj S-4. Najmanji uticaj je konstatovan u vinima koja su proizvedena pornocu kvasca Saccharomyces cerevisiae - soja 25-C. LITERATURA 1. Kiskovskij E.N. i I,M. Skurihin: Himija vina. "Piscevaja prornisljennost", Moskva, (1976), 16l. 2. Etevant P.: Wine, H.Marrse: Volatile compounds in foods and beverages. Ed. Marcel Dekker, New York (1991), Dubois P.: Les aromes des vins et leurs defauts. Rev. Fr. Oenol. 146 (1994), Baumes R: Les constituants volatils du stade fermentaire in Oenologie-Fondaments scientifiques et technologiques, Ed. Multon "Sciences-Techniques Agroalimentaires", London,-New York, Paris (1998), Salmon J.M.: Relations levure-mileu, in Oenologie-Fondaments scientifiques et technologiques, Ed. Multon Sciences-Techniques Agroalimentaires, London,-New York, Paris (1998), Thoukis G.: La mechanisme de la formation de lalcool isoamylique en utilisant les techniques des marquers. Am. J. Enol.Vitic 9 (4) (1958), Ribereau-Gayon, J., Peynaud, E., Ribereau-Gayon, P. et P. Sudraud: Teorija i praktika vinodelija, tom 2., Ed. "Piscevaja prornisljennost", Moskva (1979), Nykanen L.: Formation and occurrence of flavor compounds in wine and distilled alcoholic beverages. AmJ.Enol. Vitic. 37 (1986), Bidan P.: Relation entre la teneur des vins en alcool superieurs et la teneur des mouts en substances azotees en particulier en acides amines. Bull. orv, (48) 536 (1975), Jovic S.: Uticaj nacina prerade grozda i vinifikacije na sadrzaj metanola i vaznijih visih alkohola u vinu. Dokt. disert. Univerzitet u Novom Sadu - 'Iehnoloski fakultet (1991), Ruzic N.: Stvaranje visih alkohola tokom alkoholne fermentacije sire. Prehranbeno -tehnoloska revija 17 (4) (1979), Heraiz T., G. Regalo, M. Heraiz, P. Martin-Alvarez, M. Cabezudo: The influence of the yeast and type of culture of the volatile composition of wines fermented without sulfur dioxide. Am. J. Enol. Vitic. 41 (4) (1979), Dedyukhina, E.G., Y.K. Eroshin: Essential Metal Ions in the control of microbial metabolism. Process Biochemistry 26 (1991),
8 14. Jazic Lj. in. Ruzic: Tehnologija vina-praktikum. Tehnoloski fakultet - Novi Sad (1982), 42,45, Suturovic Z., N. Marjanovic, N. Ruzic: Odredivanje cinka, kadmijuma, olova i bakra u crnim vinima potenciometrijskom striping analizom. Poljoprivreda 383/385 (1996), Georgiev J.Y., Toncev, T., Ivanov, S., Gerov, S., Mancev, A. Jankov,A., Bamhalov, G., P. Nackov: Praktikum po vinarska tehnologija, Ed. "Hristo Danov", Plovdiv (1964), Ribercau-Gayon J., E. Peunaud, P.Sudraud, P. Ribereau-Gayon: Science et techniques du yin, tome 1, Dunod, Paris (1972), White J.: Toxic ActionofCommon ElementsTowards Yeast Growth, in Yeast Technology, Acad. press London (1954), Ruzic N. i N. Marjanovic: Uticaj bakra na aktivnost vinskog kvasca. Zbornik Tehnoloskog Iakulteta-Novi Sad 19 (1988), EFFECT OF COPPER AND ZINC CONTENT IN GRAPE MUST ON HIGHER ALCOHOLS IN WINE Nadeida E. Ruiic Effects of added copper and zinc acetate in grape must on higher alcohols content in produced wines is discussed in this paper. Fermentation was carried at 2 C with individual four selected cultures of wine yeast: Saccharomyces cerevisiae - strains marked as 25-C and MO, as well as Saccharomyces bayanus - strains marked as S-4 and V-2. Greater quantity of copper in must (11 rng/dm") indicated lower amount of higher alcohols in wine (12,3 to 35,3% less higher alcohols than in control wines). Negative effects demonstrated in musts with lower ph value (ph 3,2) and in samples inoculated with yeast Saccharomyces cerevisiae -strain MO. Wines produced from musts enriched with zinc indicated, however, higher quantity of higher alcohols. Yeast Saccharomyces bayanus - strain marked as S-4 was more resistant toward copper than other investigated wine yeasts. Zinc added in musts (24 mg/dm') stimulated synthesis of higher alcohols during fermentation (wines enriched with zinc contents 11,8 to 18,9% higher alcohols more than control wines). Synthesis of higher alcohols was better in musts with higher ph value. Influence of zinc is expressive in wines produced by yeast Saccharomyces bayanus - strain S-4. Prispeo 18. januara 2. Prihvacen 23. februara
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