The effets of vineyrd mngement nd primry nd seondry fermenttions on grpe glyoonjugtes nd onjugte frtions By Chnning W. de Bordenve Thesis sumitted to the Fulty of the Virgini Polytehni Institute nd Stte University in prtil fulfillment of the requirements for the degree of Mster of Siene In Food Siene nd Tehnology Committee: Brue W. Zoeklein, Ph.D. (Chirperson) Joseph E. Mry Ph.D. Susn E. Dunn R.D., Ph.D. 9 August, 1999 Blksurg, Virgini Keywords: glyosides, vineyrd mngement, Cernet Suvignon, Pinot noir, Brettnomyes
The effets of vineyrd mngement nd primry nd seondry fermenttions on grpe glyoonjugtes nd onjugte frtions By Chnning W. de Bordenve Astrt Grpe-derived rom nd flvor preursors exist prtilly s non-voltile, sugr-ound glyosides. Hydrolysis of these ompounds my modify sensory ttriutes nd potentilly enhne wine qulity. In the first study, four levels of shoot thinning (ontrol, 20, 25, nd 30 shoots per meter) with nd without sl lef removl (2-4 leves per shoot) were estlished on mid-wire (90 m), ilterl ordon-trined, mture Cernet Suvignon (Vitis vinifer L.) grpevines in estern Virgini in 1996 to determine the effets on grpe hemistry, glyoonjugtes, nd onjugte frtions. Redued shoot density generlly resulted in higher erry weight nd lower solule solids ( Brix) t eh smpling dte. Titrtle idity nd ph were generlly unffeted y shoot thinning. The 25 shoots per meter tretment displyed the gretest rte of inrese in totl, red-free, nd phenoli-free glyoonjugtes, expressed s glyosides (µmol). Lef removl resulted in inresed ph, totl phenolis, nd totl nthoynins t eh smpling dte nd higher onentrtion of totl, red-free, nd phenoli-free glyosides. In seond study, three rop levels [high (6.4 nd 5.3 kg/vine), medium (5.1 nd 4.9 kg/vine), nd low (3.2 nd 2.6 kg/vine) ] were estlished on mture Cernet Suvignon grpevines during the 1995 nd 1997 sesons, respetively. Cluster thinning of vines trined to mid-wire (90m), ilterl ordon-system ws performed y hnd three weeks post-loom to determine the effets on grpe glyoonjugtes nd onjugte frtions (expressed s glyosyl-gluose). In 1995, redued rop level resulted in higher solule solids onentrtion, ph, nd totl nd red-free glyosides ut did not ffet erry weight or titrtle idity. In 1997, the redued rop level tretment hd higher erry weight nd lower solule solids, sugr per erry, nd nthoynins ompred with the high tretment throughout the smpling period. The low tretment hd the highest onentrtion of totl, red-free, nd phenoli-free glyosides per grm of fresh fruit weight on the lst smpling dte nd the highest totl, red-free, nd phenoli-free glyosides per grm
of fresh fruit weight when ompred t similr solule solids onentrtions. Duo-trio signifine testing resulted in no sensory differenes mong the tretments in 1997. In third study, Pinot noir (Vitis vinifer L.) wines were inoulted with one of six genetilly different strins of Brettnomyes intermedius (Ave, M, 216, Vin 1, Vin 4, nd Vin5). Wines stored sur lie nd those rked immeditely following the ompletion of seondry fermenttion were nlyzed to determine the influene of B. intermedius strins on totl, red-free, nd phenoli-free glyoside onentrtions (estimted y the nlysis of glyosyl-gluose), nd on seleted free voltiles. Sur lie wines inoulted with strin Vin 4 nd rked wines inoulted with Vin 4 nd Vin 5 hd the lowest totl glyoside onentrtion. Hydrolysis of red-free glyosides ppered gretest in sur lie wines inoulted with Vin 4 nd rked wines inoulted with Vin 4 nd Vin 5. Wines stored sur lie tht were inoulted with M nd Vin 1 nd rked wines inoulted with Vin 1, Vin 4, nd Vin 5 hd the lowest onentrtion of phenoli-free glyosides. Wines were nlyzed for voltile ompounds known to e produed y Brettnomyes spp. Inoulted wines were found to hve detetle onentrtions of ethyl-2- methylutyrte, isomyl lohol, ethyldenote, isovleri id, guiol, 2-pheylethnol, 4-ethylguiol nd 4-ethylphenol. There were signifint differenes in the onentrtions of these ompounds mong strins. Duo-trio testing demonstrted sensory differenes etween the ontrol nd ll inoulted wines. Differenes were lso found etween wines inoulted with strins Ave nd Vin 5, strins M nd 216, nd strins M nd Vin 4. iii
Aknowledgements I would like to thnk Dr. Brue Zoeklein for giving me the opportunity to work under him s grdute student nd for his guidne nd support during my studies. His motivtion nd relentless pursuit of preision hs helped to mke me more stute reserher nd etter writer. I would like to reognize nd thnk my other ommittee memers, Drs. Dunn nd Mry for their support nd diretion throughout my grdute studies. Thnk you to Yuhing Jsinski for her help with my lortory reserh, her guidne in nd out of the lortory, nd for tehing me proper lortory tehnique, ptiene, nd for her friendship. Thnks lso to Lur Dougls nd Bo Whiton for helping me with my lortory reserh nd for their editing skills nd friendship. I would like to thnk ll of the food siene nd tehnology fulty, stff, nd grdute students for lwys eing willing to provide help when it ws needed nd for their friendship. Speil ppreition to Hether MMhon for shring n offie with me for two yers nd for eoming one of the est friends I ve ever hd. Thnks lso to ll of my friends for their support nd enourgement, espeilly Ann Mnsfield, Melnie Petros, Jeremy Vehione, Amy L. Wright, Amy N. Wright, Erik Tikel, Ligon We, Anit Hok, Kren Benner, Chris Rhines, Greg Steeno, Dvid Ruggio, nd Kri Keen. The morl nd finnil support of my fmily nd espeilly my prents enourged me to persevere to the end. Thnk you to the Willimsurg Winery, Ltd., nd espeilly Steve Wrner nd Rihrd Crmihel for supporting my reserh nd tehing me out winemking. Thnks lso to Chris Hughes for his id with my sttistil nlyses. iv
Tle of Contents Astrt...ii Aknowledgements...iv List of Tles nd Figures...vi Introdution...x Chpter I: Review of the Literture...1 A. Grpevine Mirolimte nd Vineyrd Mngement... 1 B. Fermenttion, Yest Hydrolysis, nd Arom nd Flvor Compounds...4 C. Brettnomyes intermedius... 6 D. Sensory Evlution...8 Literture Cited...10 Chpter II: Effets of shoot thinning nd sl lef removl on Cernet Suvignon (Vitis vinifer L.) grpe hemistry, glyoonjugtes, nd onjugte frtions...17 Astrt...17 Introdution...17 Mterils nd Methods...18 Results nd Disussion...20 Shoot Thinning...20 Lef Removl...22 Conlusions...23 Literture Cited...38 Chpter III: The effet of rop lod on the onentrtion of Cernet Suvignon (Vitis vinifer L.) glyoonjugtes nd onjugte frtions....43 Astrt...43 Introdution...43 Mterils nd Methods...45 Results nd Disussion...47 Conlusions...51 Literture Cited...60 Chpter IV: Effets of Brettnomyes intermedius strins on Pinot noir (Vitis vinifer L.) glyosides nd seleted free voltiles...70 Astrt...70 Introdution...70 Mterils nd Methods...72 Results nd Disussion...74 Conlusions...77 Literture Cited...86 Vit...90 v
List of Tles nd Figures Chpter II. Effets of shoot thinning nd sl lef removl on Cernet Suvignon (Vitis vinifer L.) grpe hemistry, glyoonjugtes, nd onjugte frtions...17 Tle 1. Yield nd nopy desriptors omponents of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for ontrol...25 Figure 1. 100-erry weights (g) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....26 Figure 2. Solule solids ( Brix) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....26 Figure 3. Sugr per erry ( Brix/g) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....27 Figure 4. Totl glyosyl gluose per grm (µmol/g) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol...27 Figure 5. Totl glyosyl gluose per erry (µmol/erry) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....28 Figure 6. Phenoli-free glyosyl gluose per grm (µmol/g) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....28 Figure 7. Phenoli-free glyosyl gluose per erry (µmol/erry) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....29 Figure 8. Red-free glyosyl gluose per grm (µmol/g) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....29 Figure 9. Red-free glyosyl gluose per erry (µmol/erry) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....30 Figure 10. Totl phenolis (AU) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol....30 Figure 11. Estimted totl nthoynins (AU) of Cernet Suvignon grpevines shoot thinned to 20, 25, nd 30 shoots per meter (SPM) nd non-thinned ontrol vines in 1996. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05; N=8 for 20, 25, nd 30 SPM; N=10 for the ontrol...31 Figure 12. 100-erry weights of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...32 vi
Figure 13. Solule solids ( Brix) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...32 Figure 14. Sugr per erry ( Brix/g) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...33 Figure 15. Totl glyosyl gluose per grm (µmol/g) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...33 Figure 16. Totl glyosyl gluose per erry (µmol/erry) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...34 Figure 17. Phenoli-free glyosyl gluose per grm (µmol/g) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions....34 Figure 18. Phenoli-free glyosyl gluose per erry (µmol/erry) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions....35 Figure 19. Red-free glyosyl gluose per grm (µmol/g) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...35 Figure 20. Red-free glyosyl gluose per erry (µmol/erry) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions....36 Figure 21. Totl phenolis (AU) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...36 Figure 22. Totl nthoynins (AU) of ontrol nd lef-removed (LR) Cernet Suvignon grpes whih were smpled in 1996. LSD nlysis of tretment mens. Mens with different letters re signifintly different t P 0.05; N=5 for the ontrol nd N=12 for treted replitions...37 Chpter III: The effet of rop lod on the onentrtion of Cernet Suvignon (Vitis vinifer L.) glyoonjugtes nd onjugte frtions...43 Tle 1. Cnopy desriptor omponents of Cernet Suvignon grpevines luster thinned to Low (3.2 kg/vine), Medium (5.1 kg/vine), nd High (6.4 kg/vine) rop level in 1995. Grpevine mnipultion, mesurements nd lultions were mde y nd reported y Yoder (1996). LSD nlysis of tretment mens. Mens with different letters re signifint t P 0.05, N = 12. 53 Tle 2. Yield nd nopy desriptor omponents of Cernet Suvignon grpevines luster thinned to Low (2.6kg/vine), Medium (4.9 kg/vine), nd High (5.3 kg/vine) rop levels t four smpling dtes in 1997. LSD nlysis of tretment mens. Tretments with the sme letter re not signifintly different t P 0.05, N=12....54 Figure 1. Berry weight (g) of Cernet Suvignon grpevines luster thinned to low, medium, nd high rop levels in 1995 (A) nd 1997 (B). LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens....55 Figure 2. Solule solids ( Brix) of Cernet Suvignon grpevines luster thinned to low, medium, nd high rop levels in 1995 (A) nd 1997 (B). LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens....56 vii
Figure 3. Sugr per erry (g sugr per erry) of Cernet Suvignon grpevines luster thinned to low, medium, nd high rop levels in 1995 (A) nd 1997 (B). LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens....57 Figure 4. Titrtle idity (g/l) of Cernet Suvignon grpevines luster thinned to low, medium, nd high rop levels in 1995 (A) nd 1997 (B). LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens....58 Figure 5. Totl glyosyl gluose per grm of Cernet Suvignon grpevines luster thinned to Low (3.2 kg/vine), Medium (5.1 kg/vine), nd High (6.4 kg/vine) rop levels t seven smpling dtes in 1995. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens.59 Figure 6. Totl glyosyl gluose per grm (A) nd per erry (B) of Cernet Suvignon grpevines luster thinned to Low (2.6kg/vine), Medium (4.9 kg/vine), nd High (5.3 kg/vine) rop levels t four smpling dtes in 1997. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens...60 Figure 7. Red-free glyosyl gluose per grm of Cernet Suvignon grpevines luster thinned to Low (3.2 kg/vine), Medium (5.1 kg/vine), nd High (6.4 kg/vine) rop levels t seven smpling dtes in 1995. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens...61 Figure 8. Red-free glyosyl gluose per grm (A) nd per erry (B) of Cernet Suvignon grpevines luster thinned to Low (2.6kg/vine), Medium (4.9 kg/vine), nd High (5.3 kg/vine) rop levels t five smpling dtes in 1997. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens...62 Figure 9. Phenoli-free glyosyl gluose per grm (A) nd per erry (B) of Cernet Suvignon grpevines luster thinned to Low (2.6kg/vine), Medium (4.9 kg/vine), nd High (5.3 kg/vine) rop levels t five smpling dtes in 1997. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=12. Unlelled points hve no signifint differenes etween tretment mens....63 Figure 10. Effet of three rop levels of Cernet Suvignon grpevines luster thinned to low, medium, nd high rop levels in 1995 (A) nd 1997 (B). LSD nlysis of tretment mens. Tretments with the sme letter re not signifintly different t P 0.05, N=12. ns = not signifint t P 0.05....64 Tle 3. Chemil omposition of Cernet Suvignon rop lod wines mesured prior to sensory evlution. Tretments were luster thinned to Low (2.6kg/vine), Medium (4.9 kg/vine), nd High (5.3 kg/vine) rop levels in 1997. LSD nlysis of tretment mens. Mens with the sme letter re not signifintly different t P 0.05, N=3...65 Chpter IV: Effets of Brettnomyes intermedius strins on Pinot noir (Vitis vinifer L.) wine glyoonjugtes, onjugte frtions, nd sensory ttriutes...70 Tle 1. Conentrtion (mg/l) of seleted voltile ompounds produed y six strins of B. intermedius in Pinot noir wine. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...78 Tle 2. Seleted free voltile ompounds reported to e produed y Brettnomyes (Liker et l., 1999) nd their sensory desriptors; Soure: Aldrih, 1997.....78 Figure 1. Totl glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...79 Figure 2. Red-free glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...80 viii
Figure 3 Phenoli-free glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...81 Figure 4. Totl glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...82 Figure 5 Red-free glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...83 Figure 6. Phenoli-free glyoside onentrtion, expressed s µm glyosyl gluose in Pinot noir wines inoulted with one of six strins of Brettnomyes intermedius nd stored sur lie. LSD nlysis of tretment mens. Mens with the sme letters re not signifintly different t the P 0.05 level...84 Tle 3. Results of Duo-trio sensory evlutions of Pinot noir wines inoulted with different strins of Brettnomyes intermedius. * indites signifint differenes t the P 0.1 level when mesured y Duo-trio signifine testing...85 ix
Introdution Wine qulity is influened y grpe rom nd flvor ompounds whih re, in prt, ound to sugrs, known s glyosides. Glyosides re non-voltile nd my e hydrolyzed y id or enzyme tlysis during fruit mturtion, vinifition, nd ging. One of the produts of glyoside hydrolysis is the glyones. Aglyones onstitute lrge nd omplex ssortment of hemils whih hve different quntittive nd qulittive effets on rom, flvor, struture, nd olor. Beuse glyosides re flvor preursors whose quntifition my e n ojetive inditor of potentil qulity, reserh onerning vineyrd mngement in order to mximize glyoside development nd vinifition tehniques to inrese their hydrolysis is of urrent interest to the wine industry. Beuse the identifition nd quntifition of potentil voltiles would e very ostly nd timeonsuming, nother method ws developed to estimte the totl pool of seondry metolites. Hydrolysis of glyosides yields equimolr onentrtions of D-gluose (glyosyl-gluose) nd glyones. Therefore, y hydrolyzing glyosides nd then quntifying the glyosyl-gluose in juie or wine, n estimte n e inferred regrding the onentrtion of ound rom nd flvor omponents. Cnopy mirolimte is importnt in determining fruit nd wine omposition. Altertions in nopy mirolimte ffet rdition, temperture, humidity, wind speed, nd evportion within the grpevine nopy nd n influene fruit omposition through either diret or indiret hnges in vine physiology. Cnopy mngement tehniques used to improve nopy mirolimte re onduive to the prodution of high qulity grpes. Vitiulturl prties suh s shoot thinning nd sl lef removl tht inrese sunlight exposure to leves nd fruit re known to influene grpe omposition. However, not ll studies hve reported improvements in grpe omposition or wine qulity in response to shoot thinning or sl lef removl. It is generlly epted tht vineyrd yield nd wine qulity re inversely relted. However, there re mny prmeters whih n influene the qulity nd mturtion rtes of grpes.. Cluster thinning is performed on grpevines s mens of improving lef re:fruit weight rtio. The mount of lef re needed to support unit weight of fruit is influened y ultivr, lone, vine ge, trellising system, temperture, light ondition, dy length, soil, ville nutrients, nd generl ulturl prties. Cluster thinning of grpevines helps to properly lne supply nd demnd for growth metolites nd produes x
high qulity grpes with low ph nd high solule solids due to inresed rte of mturtion. Low fruit yields hve een shown to inrese potentilly voltile terpenes, intensify romtis, nd enhne vrietl hrter in some grpes nd wines. However, not ll studies hve reported improvements in grpe omposition or wine qulity in response to luster thinning. It hs reently een shown tht glyoonjugte hydrolysis y ntive yests my ontriute to enhned flvor nd/or ouquet properties in wine in ddition to the phenols nd ftty ids whih imprt Brettnomyes hrter to wines. Some strins of B. intermedius disply high β-gluosidse tivities, whih my ontriute to glyoside hydrolysis nd potentilly enhne wine qulity. Some winemkers onsider some Brettnomyes nd other non-shromyes hrter my e enefiil in ertin styles of wine y plying positive role in flvor nd ouquet omplexity s well s imprting ged hrters in some young red wines. Unfortuntely, these speultions remin lrgely unvlidted, s glyoonjugte hydrolysis y these orgnisms in onjuntion with fermenttion nd the influene of different strins on sensory ttriutes of wines remin lrgely unstudied. The ojetive of this reserh ws to determine the effets of different vineyrd mngement tehniques on the development nd onentrtion of glyosides in Cernet Suvignon (Vitis vinifer L.) nd to determine the influene of Brettnomyes intermedius on Pinot noir (Vitis vinifer L.) wine glyosides nd sensory differenes. xi
Chpter I: Review of the Literture A. Grpevine Mirolimte nd Vineyrd Mngement Cnopy mirolimte hs een defined s the limte within nd immeditely surrounding the nopy of grpevine, omprising the omponents of rdition, temperture, humidity, wind speed, nd evportion (Smrt, 1985). Vineyrd mngement tehniques for nopy mnipultion hve een shown to e effetive mens of improving grpe qulity (Bledsoe et l., 1988; Kliewer et l., 1988; Reynolds nd Wrdle, 1989; Hunter et l., 1991). Cnopy mngement tehniques mnipulte mirolimte y influening the numer nd position of shoots, leves, nd fruit whih lter the nopy mirolimte nd ffet vine physiology nd onsequently fruit omposition (Smrt, 1985; Smrt et l., 1990). Furthermore it hs een doumented tht nopy mnipultion n ffet the sensory ttriutes of wines (Winkler, 1930; Edson et l., 1993; Hunter et l., 1995; Reynolds et l., 1996,, ). It hs lso een shown tht vrying systems of vine trining nd vine sping in ddition to nopy mnipultion n ffet vine performne, erry omposition, nd nopy mirolimte (Dokoozlin nd Hirshfelt, 1995), whih n, in turn, influene wine s sensory ttriutes (Reynolds et l., 1996, ). The soure/sink reltionship in grpevines plys mjor role in grpe omposition nd qulity (Yoder, 1996). A sink orgn (grpe) in plnt is net importer of ssimilte, while soure orgn (leves) supplies ssimiltes (Ho, 1988). Vineyrd mngement tehniques suh s rop thinning influene sink size (Ross, 1999), while lef thinning nd shoot thinning ffet soure size (Hunter nd Visser, 1988). Smrt et l. (1990) doumented three nopy mngement priniples whih help to produe grpes of superior qulity. These inlude the mintenne of helthy, well-exposed leves, modertely to wellexposed fruit, nd tht fruit should e the mjor physiologil sink during ripening. Vrying prties of vineyrd mngement hve long een used in ttempts to produe grpes of superior qulity. The mount of lef re needed to support unit weight of fruit is influened y suh ftors s ultivr, lone, vine ge, trellising system, temperture, light onditions, dy length, plnt nutrients, nd generl ulturl prties (Kliewer nd Wever, 1971). The photosyntheti effiieny of lef is primrily determined y its lotion within the nopy (Bordmn, 1977). The qulity nd quntity of rdition reeived y the fruit zone folige ffets vine physiology (Smrt, 1987). Grpe omposition nd onsequent wine lier n e redued y exessive shding (Smrt et l., 1988) euse redutions
of photosyntheti tivity our with exessive folige growth (Smrt, 1985; Hunter nd Visser, 1988) nd dversely ffet yield, grpe omposition, nd wine qulity (Hunter et l., 1995). Lef removl nd shoot thinning re prties whih hve the potentil to diretly ffet fruit omposition through ltertions in the light onditions nd temperture, nd indiretly s result of inresed ir irultion through the nopy whih redues the inidene nd severity of Botrytis inere infetion (Guler et l., 1987; Smith et l., 1988; Zoeklein et l., 1992; Perivl et l., 1994) nd powdery mildew infetion (Chellemi nd Mrois, 1992). Bsl lef removl nd shoot thinning hve een shown to inrese the mount of ville light in the fruit zone s mesured y perentge sunlight, irlst spry penetrtion, point qudrnt nlysis, nd the use of light meter to mesure the mount of photosynthetilly tive rdition (PAR) (Kliewer nd Bledsoe, 1987; Bledsoe et l., 1988; Zoeklein et l., 1992; Perivl et l., 1994; Yoder, 1996). Proper pruning retes lne etween the vine s vegettive vigor nd rop level (Ross, 1999). Modifying the vine nopy to derese folige density my improve erry olor, rom, nd flvor (Smrt nd Roinson, 1991). Shoot removl nd sl lef removl hve een shown to inrese sugr onentrtion nd redue titrtle idity in Chrdonny grpes (Wolf, et l., 1986). Bledsoe et l. (1988) demonstrted improvements in fruit omposition of Suvignon ln y lef removl. Reynolds et l. (1995) found tht sl lef removl lowered TA, ph, nd potssium, while rising odor-tive free voltile terpenes (FVT s) nd potentilly voltile terpenes (PVT s) in some Vitis vinifer ultivrs. Kliewer (1970) nd Ptterson nd Zoeklein (1990) found tht erry weight ws the vrile most ffeted y defolition tretments. Hunter et l. (1995) demonstrted tht prtil nopy defolition used inreses in fruit yield nd ostensile inreses in wine onstituents (olor density, nthoynins, nd phenolis), ultivr hrter intensity, nd overll wine qulity in Cernet Suvignon. The prtie of rop redution y luster thinning hs een shown to yield higher qulity fruit (high solule solids nd low ph) thn do other methods of rop ontrol (Winkler, 1974; Smrt, 1985; Reynolds, 1989). The prtie of djusting rop lod, on vines whih re ilterlly trined, hs een shown to inrese the terpene nd glyoside ontent of erries (Miller et l., 1996, ). Terpenes re flvor nd rom ompounds found in some grpes nd their susequent wines. Mny exist s odor-tive FVT s or s glyosides nd odorless polyols, whih n e hydrolyzed nd therefore hve the potentil to eome voltile ompounds whih my ffet rom nd flvor. These onstitute more thn fifty perent of the 2
totl terpenes found in grpes, nd they re hydrolyzed to produe FVT s y id hydrolysis, must heting, enzyme hydrolysis, nd/or wine ging (Struss et l., 1986). Blsurhmnym et l. (1979) showed tht low yielding vineyrds produe fruit with greter rom nd flvor omponents, while Sinton et l. (1978) reported tht low yields resulted in wines with high onentrtions of ette esters, low seondry lohols, nd more intense roms. Vitiulturl prties hve een shown to e influentil to seondry metolites (MCrthy, 1986; Hrdie nd Mrtin, 1990; Smrt nd Roinson, 1991; Ilnd et l., 1993; Zoeklein et l., 1996,, 1998, ) nd to wine qulity (Winkler, 1930; Sinton et l., 1978; Brvdo et l., 1984; MCrthy et l., 1987; Hunter et l., 1995; Reynolds et l., 1996; Miller et l., 1996, ; Stff et l., 1997). It hs een reported tht seletive fruit zone lef removl ffets onentrtions of totl grpe glyosides nd seleted glyones of Riesling vines grown on different trining systems (Zoeklein et l., 1998). Miller et l. (1996) demonstrted tht mong Chmourin grpevines, the djustment of shoot numer nd rop lod n influene the ssimiltion of rohydrte mteril found in grpes. Redution in rop lod of Crignne grpes, within limits, inresed wine qulity (Brvdo et l., 1984). MCrthy et l. (1987) found tht PVT s were found in higher onentrtions in rop-thinned Riesling vines. Aott et l. (1991) oserved tht higher glyoside onentrtion ws found in Shirz wines produed from vineyrds reporting superior qulity grpes while lower glyoside onentrtion oinided with low qulity fruit. Ilnd et l. (1996) nd Yoder (1996) oserved tht rop thinning inresed sugr onentrtion ( Brix), totl glyosyl-gluose (GG), olor, nd red free GG. Reynolds et l. (1996, ) demonstrted tht reduing rop level inresed rom, olor, stringeny, nd finish intensity in Pinot noir wines. Dokoozlin nd Hirshfelt (1995) demonstrted tht rop thinning etween preloom nd up to four weeks fter erry set inresed rtes of erry mturtion, olor gglomertion, solule solids umultion, nd fruit yield in grpes. Cluster thinning is the esiest mens of reduing rop lod on highly produtive vines, llowing the remining fruit to develop nd mture normlly (Winkler, 1974). Beuse wine rom nd flvor ompounds originte in grpes, it is pertinte tht we exmine how to mximize rom nd flvor preursors in the vineyrd. 3
B. Fermenttion, Yest Hydrolysis, nd Arom nd Flvor Compounds Flvor nd rom ompounds in wine re of prmount onern to vintners euse they onstitute the desriptors to whih onsumers respond. Flvor is the result of omplex retions of gusttion nd olftory reeptors (Winkler, 1974). Arom is the olftory pereption of voltile ompounds while outside the mouth (Aree, 1993). Two generl tegories of grpe flvor ompounds exist in wines: free voltiles nd sugr-ound onjugtes. Free voltiles re responsile for the roms nd flvors ommonly ssoited with vrietl wines. Sugr-ound onjugtes onsist of free voltiles ound to sugr moleules, usully gluose nd other mono- nd dishrides (Aott et l., 1990). Wine flvors, whih n e sried diretly to fermenttions, involve omplex mixtures of these voltiles nd intertion produts of these ompounds with ethnol, suh s mixed esters nd etls. These mixtures give rise to fmilir yesty nd fruity flvors ssoited with wines (Fennem, 1996). The formtion of yest-produed voltile ompounds during fermenttion is losely relted to the prtiulr speies of yest involved (Rnkine, 1967). There hve een mny different speies of ntive yests isolted from grpevines, some of whih ply n importnt role t the eginning of fermenttion. However, Shromyes erevisie is usully the most influentil speies of yest to wine rom nd flvor y the onlusion of fermenttion (Frezier nd Duoudieu, 1992) euse fter lg phse they re le to grow nd produe enough lohol to inhiit the growth of other indigenous yests (Henik-Kling et l., 1998). Differenes etween yest-produed rom ompounds oserved in wines hving undergone similr fermenttions under the sme onditions n e ttriuted to the tions of different yest strins within speies or different speies ltogether (Mteo et l., 1992; Luers et l., 1994). The role tht yests ply in fermenttion is n extremely importnt one. They re responsile for providing the enzyme β-gluosidse whih my hydrolyze glyosidi onds, whih is one wy of voltilizing rom nd flvor ompounds, while they onvert the simple sugrs found in grpes into ethnol. The role yests ply hs een shown to e two-hnded, though. Although yests my ontriute to the mehnism of glyoside hydrolysis nd prodution of voltile rom nd flvor ompounds, they hve lso een shown to use redutions in rom nd flvor ompound onentrtions. Luers et l. (1994) demonstrted tht yest wlls ind to rom sustnes. One ound to yest wlls, these rom ompounds re no longer voltile nd re therefore not pereived y humns. 4
Wine qulity is dependent on rom nd flvor ompounds whih exist s free voltiles or s ound glyoonjugtes (Aott et l., 1993; Willims et l., 1995). Beuse trditionl methods of ssessing grpe omposition nd mturity, suh s sugr onentrtion, titrtle idity, nd ph, nnot urtely predit grpe nd wine qulity (Jkson nd Lomrd, 1993), Aott et l. (1993) nd Willims nd Frnis (1996) suggest tht the nlysis of glyosides e used s n ojetive mens of mesuring potentil wine hrter. Glyosyltion is elieved to e the terminl step in the iosyntheti pthwy, rendering fruit rom nd flvor omponents nonvoltile. Glyosides re formed during grpe mturtion nd re theorized to e the result of glyosyltrnsferses whih tlyze the relotion of rohydrtes from sugrrrying nuleotides to glyones (Willims et l., 1982). Glyosides re non-voltile, odorless ompounds until they re hydrolyzed y yest β-gluosidse, id (Willims et l., 1982; Gunt et l., 1985; Frnis et l., 1992, 1996), or petinolyti enzymes with high β-gluosidse tivity (Willims et l., 1987) during fermenttion. Thus, onjugted glyosides my represent rom nd flvor preursors (Willims et l., 1989). Although there re hundreds of struturlly different glyosides present in grpes, most ontin gluose (Frnis et l., 1998). Compounds tht ontriute to rom nd flvor, the glyones, re glyosidilly linked to gluose moleules. Hydrolysis of glyosides yields equimolr onentrtions of glyones nd D-gluose, lso known s glyosyl-gluose or GG (Willims et l., 1995). This ours nturlly to limited extent during fruit mturtion due to the presene of endogenous β-gluosidses (Cordonnier nd Byonove, 1974; Cordinnier et l., 1986). However, these enzymes nnot hydrolyze the totl romti potentil in grpes (Gunt, 1984; Cordinnier et l., 1986). Wilson et l. (1986) found tht glyosides re primrily loted in juie, rther thn in the skip or pulp frtions. By determining the onentrtions of GG present in juies, n estimte of the totl pool of glyosides present n e mde (Aott et l., 1993; Willims et l., 1995). With this estimtion, n inferene n e mde to the onentrtions of voltile rom nd flvor ompounds whih my e present in wine (Willims et l., 1995). Aott et l. (1993) found tht diret orreltion existed etween GG onentrtions nd voltiles relesed during enzymti tretment. They then suggested the use of GG onentrtions s n ojetive mesure of wine qulity nd suggested these mesurements ould e used to supplement or void the 5
sujetive ssessment of judging juie qulity y sensory evlution. However, the nlysis of GG nd GG frtions nnot e used s mesure of rom/flvor sine it is known tht lrge proportion of nonrom/flvor glyosides re inluded. Nonetheless, the nlysis of GG nd GG frtions n e used in onjuntion with trditionl mesures suh s solule solids, titrtle idity, nd ph in n effort to stisftorily predit wine qulity (Reynolds nd Wrdle, 1997). C. Brettnomyes intermedius Brettnomyes spp. hs historilly een held responsile for using spoilge in wines nd is redited with using losses of millions of dollrs nnully to the wine industry worldwide (Fugelsng, 1997). It n use wines to develop unplesnt odors suggesting mmoni, mouse droppings, urnt ens, nd the pungent sent of rnyrd nimls (Hok, 1990; Chtonnet et l., 1992). Reently, however, some hve questioned whether or not the presene of Brettnomyes spp. my hve some positive influenes on wines suh s ontriuting to wine omplexity or elerting the ging proess in young red wines. Fugelsng et l. (1993) found tht o-ulture of Brettnomyes nd Shromyes erevisie n use roms nd flvors similr to those ttriuted to mlolti fermenttion, suh s enhned omplexity, ugmented fruitiness, nd diminished vegettive odors. Most ses of suspeted Brettnomyes ontmintion in wines re not usully well hrterized in terms of whih speies is involved (Fugelsng et l., 1993). However, Brettnomyes intermedius is the most frequently identified (Sponholz, 1993). Assuming tht the mjority of these identified yests re B. intermedius, this suggests the existene of severl to mny strins tht my e involved in the winemking proess. Severl proedures for detetion of Brettnomyes sp. hve een suggested inluding n indiret ondutimetri tehnique (Dek nd Beuht, 1995), polymerse hin retion (PCR) proedure (Ies et l., 1996), ftty id profiling (Rozes et l., 1992; Mlfeito-Ferreir et l., 1997), sreening methods whih identify yests tht produe β-gluosidse (Blondin et l., 1983; Chroenhi et l., 1997), nd n enzyme-linked immunosorent ssy (ELISA) (Kuniyuki et l., 1984). Heresztyn (1986) demonstrted tht Brettnomyes n e responsile for the presene of 4-ethylphenol in wines. 4-Ethylphenol hs een 6
proposed for use s n inditor for present/pst growth of Brettnomyes in wine (Chtonnet et l., 1992, 1995; Zoeklein et l., 1995; Fugelsng, 1997). Brettnomyes intermedius is known to hve n influene on wine s flvor nd rom. Two of the ompounds whih influene wines sensory hrteristis re voltile phenols suh s 4-ethylguiol nd 4-ethylphenol (Duois nd Dekimpe, 1982; Heresztyn, 1986, ; Chtonnet nd Boidron, 1988; Chtonnet et l., 1992, 1995), nd medium-hin ftty ids inluding otnoi (C8), dodenoi (C12) (Rozes et l., 1992), isoutyri, isovleri, nd 2-methylutyri ids (Fugelsng, 1997). Other odor-tive ompounds responsile for the Brettnomyes flvor found in wines inlude 2-phenylethnol, isomyl lohol, is-2-nonenl, trns-2-nonenl, β-dmsenone, nd ethyldenote (Liker et l., 1999). It hs een shown tht Brettnomyes n synthesize exessive quntities of ethylphenols y fermenting very smll mounts of the residul sugrs gluose, frutose, gltose, nd trehlose (Chtonnet et l., 1995). The mehnism y whih ethylphenols re generted y Brettnomyes involves sequene of two enzyme tivities. First, roxylse deroxyltes phenoli id diretly into vinylphenol (innmte deroxylse). Next, n oxido-redutse onverts the vinyl into ethylphenol (vinylphenol redutse) (Chtonnet et l., 1992, 1995). Fugelsng (1997) reports tht 4-ethylguiol nd 4-ethylphenol rise y these mehnisms from feruli nd p-oumri ids, respetively. In ddition to their ontriutions to the wine s sensory profile, ftty ids produed y Brettnomyes hve een shown to e inhiitory to S. erevisie during fermenttive phse growth (Viegs et l., 1989; Fugelsng et l., 1993; Rsmussen et l., 1995; Mortimer et l., 1996;). The inhiition of S. erevisie y Brettnomyes is due to the lrge mounts of eti id Brettnomyes produes from growth on gluose whih inhiit nd eventully kill yest ultures (Fugelsng, 1997). It hs reently een shown tht glyoonjugte hydrolysis y ntive yests my ontriute to enhned flvor nd/or ouquet properties in wine (Zoeklein et l., 1997; Henik-Kling et l., 1998) in ddition to the phenols nd ftty ids whih imprt Brettnomyes hrter to wines. MMhon et l. (1999) found tht some strins of B. intermedius disply high β-gluosidse tivities, whih my ontriute to glyoside hydrolysis nd potentilly enhne wine qulity. Some winemkers onsider some Brettnomyes nd other non-shromyes hrter to e enefiil in ertin styles of wine y plying positive role in flvor nd ouquet omplexity s well s imprting ged hrters in some young red 7
wines (Fugelsng, 1997; Henik-Kling et l., 1998). Unfortuntely, these speultions remin lrgely unvlidted, s glyoonjugte hydrolysis y spoilge orgnisms in onjuntion with fermenttion remins lrgely unstudied. D. Sensory Evlution Sensory evlution is the sientifi disipline used to mesure, nlyze, nd interpret retions to those hrteristis of foods nd mterils s they re pereived y the senses of sight, smell, tste, touh, nd hering (Meilgrd et l., 1991). Beuse modifition of nopy mirolimte n influene fruit omposition, nd ultimtely wine qulity through either diret or indiret ltertions in vine physiology (Smrt, 1985), sensory evlution is neessry to qulify nd quntify how wine ttriutes re ffeted. Flvor nd rom re the two most sujetive ttriutes of wine whih n e influened y vineyrd mngement. Flvor is the result of omplex retions of gusttion nd olftory reeptors (Winkler, 1974), while rom is the olftory pereption of voltile ompounds while outside the mouth (Aree, 1993). When hoosing pnel to evlute food, there re some importnt ftors to onsider. Mny psyhologil ftors n influene pnelist s pereption of food. Some influening ftors inlude the time of dy, the order in whih foods re presented to the evlutor, the numer of smples given to pnelists et (Poppers, 1994). These ftors must e reognized nd ounted for y testers in order to ttin the most urte results tht re representtive of popultion. The Duo-trio test is simple nd esily understood test. To perform this test, sujet is presented with three smples, one of whih is leled s referene to whih the pnelist my refer t ny time during the test. The other two smples inlude two unknowns of whih one is the sme smple s the referene nd the other is different unknown smple. Both re oded with rndom three-digit numers so s not to influene the judges pereptions of the smples. The ojet of the test is for the judges to identify the unknown smple whih is the sme or different s the mrked referene, ording to the testers preferene (Meilgrd et l., 1991). Results n e nlyzed using nlysis of vrine (ANOVA) sttistil tests (Roessler et l., 1973). This test n e used to determine whether n overll sttistilly signifint differene exists etween the two unknown smples. 8
Vrious vineyrd mngement tehniques hve een shown to influene wine qulity. Some studies hve demonstrted tht lef removl tretments resulted in wines with higher levels of PVT s nd FVT s (Reynolds nd Wrdle, 1989; Muley nd Morris, 1993; Reynolds et l., 1996), while others found tht lef removl tretments resulted in wines with less vegettive, grssy roms (Arnold nd Bledsoe, 1990) nd more desirle must nd florl roms nd flvors (Reynolds et l., 1996) Severl studies hve shown tht rop lod ffets wine qulity s determined y sensory pnels in some grpe ultivrs (Sinton et l., 1978; Brvdo et l.., 1984; MCrthy et l.., 1987), ut not in others (Brvdo et l.., 1985, ). Frnis et l. (1992) found tht roms resulting from hydrolysis were distinguishle from neutrl wine due to enhned rom ttriutes. The effets of Brettnomyes on the sensory hrteristis of wines hve limited doumenttion. Egli et l. (1998) suggests tht non-shromyes yests re orrelted with more intense positive nd negtive flvor ttriutes. Heresztyn (1986) desried hrteristis of wines with Brettnomyes s eing strong spiy, smoke-like, mediinl, love-like, woody, ider-like, or phenoli, while Hok (1990) doumented unplesnt odors suggesting mmoni, mouse droppings, urnt ens, nd the pungent sent of rnyrd nimls. Sensory differenes mong strins hve yet to e studied. 9
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