Characterisation of Colour Components and Polymeric Pigments of Commercial Red Wines by Using Selected UV-Vis Spectrophotometric Methods
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1 Characterisatin f Clur Cmpnents and Plymeric Pigments f Cmmercial Red Wines by Using Selected UV-Vis Spectrphtmetric Methds A. Versari*, G.P. Parpinell, A.U. Mattili Dipartiment di Scienze degli Alimenti, Universita di Blgna, P.zza Gidanich 6, Cesena 473, Italy Submitted fr publicatin: Nvember 6 Accepted fr publicatin: February 7 Key wrds: c-pigmentatin; plymeric pigments; Sangivese; tannins; wine. The cmpsitin f the clur cmpnents f cmmercial red wines frm Italy was analysed in terms f ttal clur, c-pigmentatin, SOrresistant pigments, small and large plymeric pigments, and tannins. A ttal f 18 wines, including Sangivese, Cabemet Sauvignn, Ner d' Avla, Merlt, Marzemin, Negramar, Aglianic, Cannnau and Rssese di dlceacqua frm five vintages (1999 t 3) were analysed. The cmpsitin f the wines shwed significant differences in terms f ttal clur and c-pigmentatin. Ttal clur at 5 nm was highest in Marzemin (7.5±1.5 AU, absrbance units), and then in Aglianic (7.1±.8 AU) and Cabemet Sauvignn (5.1±.5 AU). The level f large and small plymeric pigments in Sangivese wines shwed a range frm.5 t.4 and.1 t.7 AU 5 nm respectively. The variatin in the level f plymeric pigments is affected by a number f factrs, including vintage, grape cmpsitin, fermentatin and strage cnditins. INTRODUCTION The clur cmpnents f wine are imprtant parameters that cntribute t the sensry characteristics (e.g. clur and astringency) and the antixidant prperties f wine (Kanner et al., 1994; Hagerman et al., 1998; Waterhuse, ; L6pez-Ve1ez et al., 3). The clur f red wine is the result f an interesting and challenging, cmplex mixture f several cmpnents, including free mnmeric anthcyanins (Mazza, 1995), the enhancement f their clur due t c-pigmentatin with ther nnclured phenlics (Bultn, 1), and plymeric pigments (Smers, 1971). The clur f a red wine cntinues t change thrughut its life and can be affected by a number f winemaking practices and envirnmental cnditins. During fermentatin, and especially during aging, the mnmeric anthcyanins are thught t be gradually incrprated int plymeric pigments and this cnfers clur stability t a wine (Singletn and Esau, 1969; Smers and Evans, 1974; Ribereau-Gayn, 198). The mechanisms, the prducts and the kinetics f plymer frmatin in wine are nly partially knwn (Fulcrand et al., 4; Kennedy and Hayasaka, 4). Objective measurement f the cmpnents f wine clur is an essential part f the mdern cncept f winemaking. In this view, the availability f new methds fr the measurement f plymeric pigments prvides imprtant tls fr a better understanding f the chemistry f red wine clur. The spectrphtmetric assays develped thus far are based n the assumptin that the plymeric pigments are much less sensitive than the anthcyanins t S and t changes in ph. The classic spectrphtmetric assay f Ribereau-Gayn and Stnestreet (1965) measures the plymeric pigments f wine after the bisu1phite bleaching f anthcyanin mnmers carried ut n an acidified sample (PH <1). Smers and Evans (1974; 1977) have develped a set f spectrphtmetric measures t determine the amunt f ttal anthcyanins and plymeric pigments in red wines. This prcedure is based upn tw assumptins: (i) bisu1phite bleaches nly the mnmeric anthcyanins; and (ii) the clur f the plymeric pigments increases by a factr f % n acidificatin. Bakker et al. (1986) have shwn that ligmeric pigments are als bleached t sme extent by bisu1phite. Recently, Harbertsn et al. (3) prpsed a new methd fr the analysis f the plymeric pigments in wine based n a cmbinatin f prtein precipitatin and the bleaching effect f bisu1- phite. This methd allws fr the quantificatin f tw classes f nn-bleachable plymeric pigments: small plymeric pigments (SPP) that are nt precipitable with prtein, and large plymeric pigments (LPP), which are precipitable with prtein. The SPP fractin prbably cntains several anthcyanin-derived cmpunds in varius prprtins that have been frmed thrugh different mechanisms, such as crss-linking reactin, direct reactin and cycl-additin. The LPP fractin wuld be expected t cntain anthcyanins that have reacted directly with plymeric flavan-3-1s r plymeric pigments that have been frmed by acetaldehyde crss-linking. The levels f SPP and LPP have been mnitred in grape berries, and infrmatin n their cntent in cmmercial wines is limited t Zinfande1, Pint nir, Cabernet Sauvignn and Syrah (Adams et al., 4). The aim f this study was t characterise selected clur cmpnents, including c-pigmentatin, small and large plymeric pigments and tannins, f a variety f cmmercial red wines frm Italy, especially frm Sangivese grapes. *Crrespnding authr: address: andrea.versari@unib.it. Tel: ; Fax: s. Afr. J. Enl. Vitic., Vl. 8, N.1, 7 6
2 Clur and pigment detectin with UV-VIS Spectrmetry 7 MATERIALS AND METHODS Wine samples A ttal f 18 samples (n) f cmmercial bttled red wines frm nine regins in Italy, including Sangivese (n = 7 samples), Cabernet Sauvignn (n = 13), Ner d' Avla (n = 1), Merlt (n = 9), Marzemin (n = 7), Negramar (n = 6), Aglianic (n = 5), Cannnau (n = 4), and Rssese di dlceacqua (n = 4), were analysed (see Table 1). The map in Figure 1 shws the regins f rigin f the red wines analysed in this study. The wines were analysed at between three and 48 mnths f age (vintage years: 1999 t 3). Spectrphtmetric methd f Bultn Fr each wine (18 samples), the fllwing parameters were measured by spectrphtmetric assay accrding t the literature: ttal clur, c-pigmentatin and S-resistant pigments (Bultn et al., 1999). The samples were filtered thrugh a.45!lid plytetraflurethylene membrane (PfFE) syringe filter and the wine ph was adjusted t 3.6, befre spectrphtmetric analysis was dne using a Shimadzu 14 mdel (Shimadzu, Milan, Italy). Twenty micrlitres f 1% (v/v) acetaldehyde was added t ml f wine and, after 45 min at rm temperature, the ttal clur f the wine was measured at 5 nm. T anther ml f wine, 16,.u f 5% (w/v) S was added, and then the value f the S-resistant pigments was measured at 5 nm. Fr c-pigmentatin, the absrbance f wine at 5 nm was read after a sample dilutin f 1:19 in a 1% ethanl, ph 3.6 ptassium bitartrate buffer, crrected fr the dilutin. There are ph effects n the free mnmeric anthcyanin inisatin, the clur f the c-pigmented frm, and the clured plymers. The selected ph value (3.6) prvides the nly ratinal basis n which t cmpare clur cmpnents, which are independent f ph effects acrss all wines. Spectrphtmetric methd f Adams A set f 4 Sangivese wines frm the Emilia-Rmagna regin was further analysed by spectrphtmetric methds as described in the literature fr the fllwing parameters: small and large plymeric pigments (Harberts n et al., 3), and tannin (Harbertsn et al., ). Simultaneus analysis f the plymeric pigments and tannin in wines required tw 1.5 ml micrfuge tubes fr each sample. The first tube was made up by dispensing 1 ml f the acetic acid! FIGURE 1 Map f Italy shwing the rigin f the red wines analysed in this study. Regins: (A) Campania; (B) Emilia-Rmagna; (C) Liguria; CD) Puglia; (E) Sardegna; (F) Sicilia; (G) Trentin; (II) Umbria; (I) Venet. NaCI buffer (cntaining mm acetic acid and 17 mm NaCI adjusted t ph 4.9 with NaOH) int the tube and then adding 5,.u.. f the diluted wine (with a buffer f 1% aqueus ethanl (v/v), cntaining 5 gil ptassium bitartrate adjusted t ph 3.3 with HCI). One ml f the mixture was transferred t a cuvette and the absrbance at 5 nm was determined (reading A). Then 8,.u.. f.36 M ptassium metabisulphite was added and mixed, and the absrbance at 5 nm was redetermined after a 1 min incubatin (reading B). Fr the secnd tube, 1 ml f the acetic acid/nacl buffer cntaining bvine serum albumin at 1 mglml (BSA Fractin V pwder, Sigma, Milan, Italy) was put int a 1.5 ml micrfuge tube, and 5,.u.. f the diluted wine was added. The mixture was allwed t incubate at rm temperature fr 15 min with slw agitatin. After incubatin, the sample was cen- TABLE 1 Origin, type and number f red wines frm the different regins f Italy. Cde Regin Wine (number f samples) Ttal A Campania: Aglianic (5) 5 B Emilia-Rmagna: Merlt (), Sangivese (68), Cabemet Sauvignn (11) 81 C Liguria: Rssese di dlceacqua (4) 4 D Puglia: Sangivese (), Negramar (6) 8 E Sardegna: Cannnau (4) 4 F Sicilia: Merlt (), Ner d'avla (1) 1 G Trentin: Merlt (1), Marzemin (7) 8 H Umbria: Merlt (1) 1 Venet: Merlt (3), Cabemet Sauvignn () 5 Ttal 18 S. Afr. J. Enl. Vitic., Vl. 8, N.1, 7
3 8 Clur and pigment detectin with UV-VIS Spectrmetry trifuged fr 5 min at 13 5 g t pellet the tannin-prtein precipitate. One millilitre f the supernatant was put int a cuvette, 8!1L f.36 M ptassium metabisulphite was added and, after a 1 min incubatin, the absrbance was determined at 5 nm (reading C). The absrbance due t MP, SPP and LPP is given as (A B), C and (B-C) respectively. The absrbance due t plymeric pigments in the prtein-tannin precipitate was determined by redisslving the tannin-prtein precipitate (frm the secnd tube abve) and measuring the absrbance at 51 nm. The tannin-prtein pellet frm tube tw was first washed with 5!1L f the acetic acid/nacl buffer, and the sample was recentrifuged fr 1 min at 13 5 g. The wash slutin was discarded, after which 875!1L f a buffer cntaining 5% triethanlamine (TEA) (v/v) and 5% sdium ddecyl sulphate (SDS; lauryl sulphate, sdium salt, Sigma, Milan, Italy) (w/v) was added. After incubatin fr 1 min at rm temperature, the tube was vrtexed and the absrbance at 51 nm was determined. This absrbance represents the amunt f plymeric pigments that precipitated with the prtein. Fr tannin analysis, 15!1L f the ferric chlride hexahydrate (Sigma, Milan, Italy), was added and the absrbance at 51 nm was determined after a 1 min incubatin. The absrbance due t tannins in the sample is calculated as the final absrbance minus the backgrund (absrbance value due t the reagents withut the sample) and expressed in catechin equivalents by cmparisn with a standard curve pre- pared with standard slutin f (+ )-catechin (Extrasynthese, Genay, France), using eight cncentratin levels ranging frm 5 t 5 mgil (R =.999). All data are expressed in absrbance units (AU) with 1 cm length path and crrected by dilutins, unless therwise specified. Statistical analysis The statistical analysis f the data, including linear regressin analysis, analysis f variance (ANOVA) and pst-hc cmparisn by Tukey HSD test fr unequal sample size at 5% prbability level (p ~.5), was perfrmed using Statistica 5.1 (StatSft, Tulsa, OK). RESULTS AND DISCUSSION This investigatin was initiated as a survey f Sangivese wines because f the ecnmic imprtance f this grape cultivar in the Emilia-Rmagna regin. Taking int accunt the scientific and practical interest in the analysis f the clur cmpnents f red wine and the availability f additinal wines, it was decided t include wines frm tw internatinal varieties in this study, viz. Cabernet Sauvignn and Merlt, as well as wines frm lcal emerging grape varieties, such as Cannnau, Marzemin and Ner d' Avla. The first step in the study fcused n the analyses f ttal clur, c-pigmentatin, and S-resistant pigments in the wines using Bultn's methd. The cmpsitin f the wines shwed a TABLE Cmpsitin and statistical analysis f 18 red wines frm Italy. All parameters are expressed as AU at 5 nm. Wine N.sample Thtal clr Cpigmentatin S resistant pigments Mean ±SD* Mean±SD* Mean±SD** Sangivese ±1.31a.35±.38a.36±1. Cabemet Sauvignn 3 5.1±.5ab.41±.4a 3.5±.16 Cannnau ±.87a.71±.64ab 1.65±.76 Merlt 9 4.7±1.63a.6±.37ab.9±.99 Aglianic 5 7.1O±.9b.39±.3ab 3.3±.86 Negramar ±1.19a.8±.53ab.9±.78 Marzemin 7 7.5±1.5b 1.18±.63b 1.91±.4 Ner d'avla ±1.18a.3±.3a 1.85±.57 Rssese 4.48±.34a.6±.66ab.86±.3 *Grup cmparisns by means f Tukey HSD test fr unequal sample size. Means witbin each clumn with changed letter are significantly different using Tukey HSD test ~.5). **Nn-significant difference at p-ievel :::;.5. TABLE 3 Cmpsitin f 4 Sangivese wines frm Emilia-Rmagna regin, Italy. LPP SPP LPP+SPP LPP/sPP Thnnins AU 5 nm AU5nm AU 5 nm gil Year N samples mean±sd mean±sd mean±sd mean±sd mean±sd ±.37.99±.33.8± ±O.43.49± ± ±.3.35± ±O.38.67± ± ±.38.8± ±O.69.88± ± ±.4.46± ±O.37.8±.4 S. Afr. J. Enl. Vitic., Vl. 8, N.1, 7
4 Clur and pigment detectin with UV-VIS Spectrmetry significant difference in terms f ttal clur and c-pigmentatin, while there was n significant difference between the wines regarding S-resistant pigments (see Table ). Marzemin and Aglianic shwed the greatest ttal clur at 5 nm, fllwed by Cabernet Sauvignn, and then the thers wines. The level f cpigmentatin was highest in Marzemin wines, and lwest in Sangivese. The highest cntent f S-resistant pigments was fund in Cabernet Sauvignn (3.5±.16 AU 5 nm), whereas Sangivese wines shwed an intermediate level f S-resistant pigments (.36±1. AU 5 nm). In terms f clur, Marzemini and Aglianic wines seem t be suitable fr aging due t the high level f their ttal clur and S-resistant pigments. The general levels f S-resistant pigments fund in the analysed wines are cmparable with the values reprted in ther papers cncerning studies f plymeric pigments in different wines frm Canada (Mazza et al., 1999), Italy (Piergivanni et al., 1986; Bselli et al., 4), Spain (Gmez-Plaza, et al., 1999), the USA (Nagel and Wulf, 1979), and Australia (Smers and Evans, 1974). It is well dcumented in the literature that the grape cultivar affects the amunt f ttal clur and the c-pigmentatin f wine t a large extent (Bultn et al., 1999; Mazza et al., 1999; Bselli et al., 4; Versari et al., 4). The data presented here supprt this view. The c-pigmentatin f anthcyanins in wines is a cmpetitive equilibrium invlving several anthcyanins and many cfactrs. Wines made frm grapes high in cfactrs and/r with a prevalence f acylated frms f the nn-malvidin pigments in their anthcyanin pattern may have a higher level f c-pigmentatin (Bultn, 1). This is partly the reasn fr prer c-pigmentatin in the Sangivese wines, which lack the acylated pigments (Baldi and Rmani, 199), whereas Merlt and Cabernet Sauvignn wines cntain high levels f acylated anthcyanins (Mattivi et al., 1995), fllwed by Aglianic (Cal et al., 1994) and Cannnau (Castia et al., 199). Furthermre, c-pig- ~~-~~-~--~-~--~--~-~~ 9 mentatin is typical f yung wines, which can explain up t 5% f the ttal clur (Bultn, 1). T btain mre insight int the cmpsitin f Sangivese wines in terms f clur cmpnents, the S-resistant pigments were analysed using a mre selective methd prpsed by Harbertsn et al. (3), which allws the measurement f tw grups f plymeric pigments: SPP and LPP. This part f the investigatin was cnducted n a set f 4 Sangivese wines that were between three and 48 mnths ld (vintage years: 1999 t 3) at the time f these analyses. The S-resistant pigments accunted fr the average percentage f 6%,7%, 65% and 54% f the ttal clur f fur-year-ld (1999), three-year-ld (), tw-year-ld (1) and ne-year-ld () Sangivese wines respectively. Table 3 shws the level f LPP, SPP and tannins in Sangivese wines. The cmparisn between vintages shwed an increasing level f LPP, SPP and tannins in Sangivese wines during the first three years, i.e. 3 t. The variatin in the level f plymeric pigments is affected by a number f factrs, including vintage, grape cmpsitin, fermentatin and strage cnditins. In particular, the decrease in plymeric pigments may be due t precipitatin and/r the lighter hue f the larger plymers. There is general cnsensus n the increase in LPP cntent in red wines with age (De Beer et al., 4). The levels f LPP (range.3 t.3 AU) and SPP (range.6 t. AU) fund in Sangivese wines are generally higher than the values reprted fr wines made frm lcal red grape varieties in the Marche regin, Italy (Bselli et al., 4), and fr red wines frm the USA (De Beer et al., 4). The distributin f the LPP/SPP rati in Sangivese wines cvered a range f.15 t.78 (see Figure ), and was in agreement with the values reprted n Pint Nir (range.14 t.), Cabernet Sauvignn (.18 t 3.93), and Syrah (.8 t 1.71) (Harbertsn et al., 3). The highest crrelatin value was fund between the determinatin f S-resistant pigments by Bultn's assay at ph 3.6 and the sum f LPP and SPP as determined by Adams' assay using 4 -, , 18 R =.85 1 '. 1.n 4-; t.n E Z ::;, en ~ 16.. ~ 14 g "~ ~ ,-----,---~ LPP/SPP rati 3 4 S resistant pigments (AU 5 nm) FIGURE FIGURE 3 Frequency distributin f LPP/SPP rati fund in 4 Sangivese wines frm the Emilia-Rmagna regin (Italy). Crrelatin between the amunt f S-resistant pigments determined by Bultn's assay and LPP+SPP determined by Adams's assay fund in 4 Sangivese wines frm the Emilia-Rmagna regin (Italy). s. Afr. J. Enl. Vitic., Vl. 8, N.1, 7
5 1 Clur and pigment detectin with UV-VIS Spectrmetry cmbined prtein precipitatin and bisulphate bleaching at ph 4.9 (r =.91; R. =.85) (see Figure 3). This finding cnfirms that Adams's assay fractinates plymeric pigments int tw classes, thse that precipitate with prtein and thse that d nt (Harberts n et al., 3). CONCLUSIONS In cnclusin, the recent availability f selective UV-Vis methds fr the measurement f parameters that have nt been taken int accunt in traditinal wine clur measurements represents a valuable pprtunity fr researchers and winemakers. The chice f a prper analytical methd is critical, thught in many cases the results that are btained differ n the basis f the analytical prcedure. LITERATURE CITED Adams, D.O., Harbertsn, le & Piccitt, E.A., 4. Fractinatin f red wine plymeric pigments by prtein precipitatin and bisulphite bleaching. In: Waterhuse, A.L. & Kennedy, J.A. (eds). Red Wme Clr: Revealing the Mysteries. American Chemical Sciety 886, Washingtn, DC. pp Bakker, J., Prestn, N.W. & Timberlake, C.E, The determinatin f anthcyanins in aging red wines: Cmparisn f HPLC and spectral methds. Am. J. Enl. Vitic. 37, Baldi, A. & Rmani, A., 199. Studi su alcuni cmpsti plifenlici in uve, msti, vini della Tscana. Entecnic 6, Bselli, E., Bultn, R.B., Thmgate, J.H. & Frega, N.G., 4. Chemical and sensry characterizatin f DOC wines frm Marche (Italy) related t vintage and grape cultivars. J. Agric. Fd Chern. 5, Bultn, R., Neri, R., Levengd, J. & Vaadia, M., Cpigmentatin f anthcyanins in Cabemet Sauvignn and Merlt wines frm the Napa valley f Califrnia. In: Lnvaud-Funel, A. (ed). Prc. 6th Sympsium Internatinal d'cenlgie, Tec. & Dc. Publ., Paris, France. pp Bultn, R., 1. The cpigmentatin f anthcyanins and its rle in the clr f red wine: A critical review. Am. J. Enl. Vitic. 5, Cal, A., Tmasi, D., Craver, M.C. & Di Stefan, R., Cntribut alia caratterizzazine e classificazine varietale (Vitis sp.) attravers la determinazine degli antciani e degli acidi idrssicinnamiltartarici della buccia di varieta a bacca rssa. Rivista Vitic. Enl. (Italy) 3, Castia, T., Franc, M.A., Mattivi, E, Muggilu, G., Sferlazz, G. & Versini, G., 199. Characterizatin f grapes cultivated in Sardinia: chemmetric methds applied t the anthcyanic fractin. Sci. Aliment. 1, De Beer, D., Harbertsn, J.E, Kilmartin, P.A., Rginsky, v., Barsukva, T., Adams, D. O. & Waterhuse, A.L., 4. Phenlics: A cmparisn f diverse analytical methds. Am. J. Enl. Vitic. 55, Fulcrand, H., Atanasva, V., Salas, E. & Cheynier, V., 4. The fate f anthcyanins in wine: Are there determining factrs? In: Waterhuse, A.L. & Kennedy, J.A. (eds). Red Wme Clr: Revealing the Mysteries. American Chemical Sciety 886, Washingtn, DC. pp Gmez-Plaza, E., Gil-Munz, R., Lpez-Rca, 1M. & Martinez, A., Clr and phenlic cmpunds f a yung red wine as discriminating variables f its aging status. Fd Res. Int. 3, Hagerman, A.E., Riedl, K.M., Jnes, G.A., Svik, K.N., Ritchard, N.T., Hartzfeld, P.w. & Riechel, T.L., High mlecular weight plant plyphenlics (tannins) as bilgical antixidants. J. Agric. Fd Chern. 46, Habertsn, J.E, Kennedy, J.A.,. Tannin in skins and seeds f Cabemet Sauvignn, Shiraz and Pint Nir berries during ripening. Am. J. Enl. Vitic. 53, Harbertsn, J.E, Piccitt, E.A. & Adams, D.O., 3. Measurement f plymeric pigments in grape berry extracts and wine using a prtein precipitatin assay cmbined with bisulfite bleaching. Am. J. Enl. Vitic. 54, Kanner, J., Frankel, E.N., Granit, R., German, B. & Kinsella, le., Natural antixidants in grapes and wines. J. Agric. Fd Chern. 4, Kennedy, J.A. & Hayasaka, Y., 4. Cmpsitinal investigatin f pigmented tannin. In: Waterhuse, A.L. & Kennedy, J.A. (eds). Red Wme Clr: Revealing the Mysteries. American Chemical Sciety 886, Washingtn, DC. pp L6pez-V6Iez, M., Martinez-Martinez, E & Del Valle-Ribes, C., 3. The study f phenlic cmpunds as natural antixidants in wine. Crit. Rev. Fd Sci. Nutr. 43, Mattivi, E, Mnetti, A. & Niclini, G., Cmpsizine fenlica e caratterizzazine di vini rssi mnvarietali. Entecnic (Italy) 6, Mazza, G., Anthcyanins in grapes and grape prducts. Crit. Rev. Fd Sci. Nutr. 35, Mazza, G., Fukumt, L., Delaquis, P., Girare, P. & Ewert, B., Anthcyanins, phenlics, and clr f Cabemet Franc, Merlt, and Pint Nir wines frm British Clumbia. J. Agric. Fd Chern. 47, Nagel, C.w. & Wulf, L.w., Changes in the anthcyanins, flavnids, and hydrxycinnarnic acid esters during fermentatin and aging f Merlt and Cabemet Sauvignn. Am. J. Enl. Vitic. 3, Piergivanni, L., Vlntari, G. & Scibna A., Studi della frazine antcianica: mdificazini nel crs dell'invecchiament di un vin Barbera. Ind. Bevande (Italy) 6, -9, 17. Ribereau-Gayn, P The chemistry f red wine clr. In: Webb, A.D. (ed). The Chemistry f Winemaking. American Chemical Sciety, Advanced Chemistry Series, N. 137, Washingtn, DC. pp Ribereau-Gayn, P. & Stnestreet, E., Le dsage des anthcyanes dans Ie vin ruge. Bull. Sc. Chim. Fr. 9, Singletn, V.L. & Esau, P., Phenlic Substances in Grape and Wines and Their Significance. Academic Press, New Yrk, NY. Smers, T.C., The plymeric nature f wine pigments. Phytchemistry 1, Smers, T.C. & Evans, M.E., Wme quality: Crrelatins with clur density and anthcyanin equilibria in a grup f yung red wines. J. Sci. Fd Agric. 5, Smers, T.C. & Evans, M.E., Spectral evaluatin f yung red wines: Anthcyanin equilibria, ttal phenlics, free and mlecular S, "chemical age". J. Sci. Fd Agric. 8, Versari, A., Bultn, R. & Thmgate, 1, 4. The predictin f clr cmpnents f red wines using FTIR, wine analyses, and the methd f partial least squares. In: Waterhuse, A.L. & Kennedy, J.A. (eds). Red Wme Clr: Revealing the Mysteries. American Chemical Sciety 886, Washingtn, DC. pp Waterhuse, A.L.,. The phenlic wine antixidants. In: Cadenas, E. & Packer, L. (eds). Handbk f Antixidants. Marcel Dekker, New Yrk. pp s. Afr. J. Enl. Vitic., Vl. 8, N.1, 7
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