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Transcript of Flavonoids in grapes - awri.com.au · • Flavonoids are important for the colour & taste of wine...
Flavonoids in grapes
PLANT INDUSTRY
Simon Robinson, Mandy Walker, Rachel Kilmister and Mark Downey
11 June 2014
Grapes to wine – a 2 metabolic zoo
Grapevines
Grapes
• Grape variety
• Soil, climate etc
• Vineyard management
• Seasonal influences
• Harvest timing
Wine
• Transport & storage
• Processing
• Fermentation
• Pressing
• Added tannin, oak etc
• Ageing
Consumer
Hundreds of different metabolites determine
‘Wine Quality’
Many of these compounds are synthesised in the grape berry
• Flavonoids are important for the colour & taste of wine • Tannins are the major flavonoid present in grapes and wine • Tannins plus anthocyanins form pigmented polymers
Grape Flavonoids
Anthocyanins
Red Grapes
0
1000
2000
3000
4000
Flavonols Tannins
mg/kg
Flavonoids in grapes
‘Moon’ series carnations : on sale since 1996
Flavonoids – a diverse range of compounds
Basic anthocyanin structure
• Complex metabolic pathway • 1,000’s of different flavonoids • Pathway can be manipulated
FLS Flavonols
Coumaroyl-CoA
Chalcones
Flavanones
Dihydroflavonols
DFR
CHI
F3H
LDOX
Leucoanthocyanidin
Anthocyanidin
CHS
UFGT
Anthocyanins
Catechin LAR
Flavonoid biosynthetic pathway in grape
Epicatechin ANR
Tannins
FGT F3’H
Dihydroflavonols
DFR
LDOX
Leucoanthocyanidin
Anthocyanidin
UFGT
Anthocyanins
F3’5’H
Malvidin-3-glucoside
?
Good knowledge of this
pathway in grapes
Colour & tannin during development
• Anthocyanins are synthesised after veraison
•Tannins and flavonols are made between flowering & veraison
• Tannin maturation occurs from veraison to harvest
Tannin synthesis and maturation
Regulatory Genes Control the Pathway
ANTHOCYANINS
Structural Pathway Genes
Enzymes
Regulatory Genes
(bHLH & MYB) TANNINS
FLAVONOLS
Regulatory ‘Master Genes’ control the pathway
Red White
grape seedlings
Predicts fruit colour
Test for DNA marker
MYBA controls colour in grapes
Chardonnay MYBA promoter
35S promoter
‘Black’ Chardonnay – gain of VvMYBA gene expression
Shiraz Pink White
‘Pink’ and ‘White’ Shiraz – reduction of VvMYBA gene expression
MYBA controls anthocyanin synthesis
Formation of tannin polymers
Leucocyanidin Catechin
Epicatechin Flavan-3-ols
LDOX
Leucocyanidin
Anthocyanidin
Catechin LAR
Epicatechin ANR
Tannins ?
Tannin Synthesis - the First Steps Formation of tannin polymers
Leucocyanidin Catechin
Epicatechin Flavan-3-ols
Monomers Dimer
Terminal
unit
Extension
unit
Terminal
unit
Extension
units
Trimer
Tannin Polymer
N=3-200
We need to know about tannin composition, as well as the total amount
OHO
OH
OR2
OH
OH
R1
OHO
OH
OR2
OH
OH
R1
OHO
OH
OR2
OH
OH
R1
OHO
OH
OH
OH
OR2
A C
B
n
Terminal Subunit
Extension Subunit
Interflavan Bond
2
345
6
78
3'
4'
5'
LAR
Leucocyanidin
Catechin / Epicatechin
Tannins ain’t tannins !
• Seed tannins mdp 5 units • Skin tannins mdp 25 units • Different polymer composition
Tannin composition
Epigallotatechin
Catechin
Epicatechin
Epicatechin gallate
Sunraysia Shiraz 2008
Epigallotatechin
Catechin
Epicatechin
Epicatechin gallate
Sunraysia Shiraz 2009
Epigallotatechin
Catechin
Epicatechin
Epicatechin gallate
Sunraysia Cabernet Sauvignon 2008
Epigallotatechin
Catechin
Epicatechin
Epicatechin gallate
Glenrowan Shiraz 2009
Role of ANR & LAR in tannin synthesis using transgenic grapevines
LDOX
Leucoanthocyanidin
Anthocyanidin
UFGT
Anthocyanins
Catechin LAR
Epicatechin ANR
Tannins ?
Silence the grape ANR gene
Silence the grape LAR gene
No obvious phenotype !
Silence the genes for ANR & LAR to decrease their enzyme activity and determine the effect on tannin synthesis and grape tannin composition
VvANR Si
leaves More cyanidin
roots normal
berries More cyanidin
leaves More flavonols
seeds Lots of flavonols
skins More flavonols
skins Less tannin
seeds Same tannin ?
leaves Less tannin
Anthocyanin
Flavonol
Tannin
Silencing tannin genes
• Less tannin in leaves & grape skins from ANR Si lines (10-30% Controls)
• Changes in composition – extension units have less epicatechin, more catechin & epigallocatechin
• Seed tannin slightly lower (70-85% Controls)
• Impact on wine?
Shiraz VvANR Si
How do you manage colour in the vineyard?
Where are you now? Where do you want to be?
• More grape colour ? • Better colour extraction & stability ? • Better anthocyanin composition ?
Based on grape or wine observations?
• Season & site are the biggest drivers of variation • Day and night temperatures • Practices that effect colour include:
– Yield / vigour / vine balance
– Rootstocks
– Irrigation
– Canopy management / bunch exposure
– Vineyard variability
How do you manage tannin in the vineyard?
Where are you now? Where do you want to be?
• More tannin
• Less tannin
• Different tannin
• Better tannin • Smooth tannin
• Soft tannin
• Persistent tannin
Based on grape or wine observations?
Wine Tannin = Grape Tannin x Extractability x Stability
• How much tannin is in the grapes ?
• How much of that tannin is extracted into must ?
• How stable is the tannin in the wine ?
If the tannin isn’t extracted during winemaking, there’s no point trying to get more into the grapes.
Grape tannin versus wine tannin
0.0
0.5
1.0
1.5
2.0
2.5
3.0
2003 2004 2005 2006 2007 2008 2009 2010
Tannin
(m
g/g
FW
T b
err
y)
Season effects Sunraysia Shiraz- whole berries at harvest
0.0
1.0
2.0
3.0
4.0
5.0
Carignane
(n=
5)
Barb
era
(n=
18)
Rubired
(n=
149)
Zin
fandel (n
=68)
Gre
nache (
n=
13)
Carn
elia
n (
n=
3)
Shiraz (
n=
73)
Ruby C
abern
et (n
=33)
Cabern
et S
auvig
non (
n=
77)
Merlot (n
=154)
Chard
onnay (
n=
3)
Durif(n
=8)
Tota
l T
annin
(m
g/g
fre
sh w
eig
ht w
hole
berr
y)
0.0
1.0
2.0
3.0
4.0
5.0
Carignane
(n=
5)
Barb
era
(n=
18)
Rubired
(n=
149)
Zin
fandel (n
=68)
Gre
nache (
n=
13)
Carn
elia
n (
n=
3)
Shiraz (
n=
73)
Ruby C
abern
et (n
=33)
Cabern
et S
auvig
non (
n=
77)
Merlot (n
=154)
Chard
onnay (
n=
3)
Durif(n
=8)
Tota
l T
annin
(m
g/g
fre
sh w
eig
ht w
hole
berr
y) ~600 grape samples
whole berry homogenate
Californian vineyards
Variation between varieties
0.0
1.0
2.0
3.0
4.0
5.0
6.0
SHIRAZ CABERNET SAUVIGNON
PINOT NOIR SHIRAZ CABERNET SAUVIGNON
PINOT NOIR SHIRAZ CABERNET SAUVIGNON
PINOT NOIR
Sunraysia 2003-2004 Yarra Valley 2003-2004 Mornington 2003-2004
Tan
nin
mg
/g f
wt
ber
ry
Varietal differences and site effects
Sunraysia Yarra Valley Mornington
Viticultural management
• Season is the biggest driver of variation
• Understanding other drivers creates management strategies
• Practices that effect tannins might include:
– Irrigation
– Canopy management
– Trellis design
– Nutrient status
– Pruning
– Harvest date
– Cover crop
– Mulching
– Rootstock
Yield
0.0
1.0
2.0
3.0
4.0
5.0
0.0 2.0 4.0 6.0 8.0 10.0
To
tal T
an
nin
(m
g/g
wh
ole
be
rry)
r2 = 0.2771
0.0
0.5
1.0
1.5
2.0
2.5
3.0
0.0 2.0 4.0 6.0 8.0 10.0
To
tal A
nth
ocya
nin
(m
g/g
who
le b
err
y)
r2 = 0.5724
California
Yield (kg/m2) Yield (kg/m2)
Anthocyanin Tannin
Sustained deficit irrigation
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
2004 2005 2006 2007
To
tal Tan
nin
(m
g/g
berr
y)
Cabernet Sauvignon 43% 52% 70% 100%
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
2004 2005 2006 2007
To
tal Tan
nin
(m
g/g
berr
y)
Shiraz 35% 45% 65% 100%
Sunraysia
• Large seasonal variation
• Irrigation has no major effect on tannin synthesis
• It may effect vigour, berry size and tannin extractability
Rootstocks
0
2
4
6
8
10
12
14
16
18
1103
PAULSEN
140
RUGGERI
DOGRIDGE FREEDOM MG5512 MG6262 MS1010 MS3769 RAMSEY
Ta
nn
in (
mg
/g f
wt
sk
in)
2008 2009
Sunraysia Shiraz
0.90 - 1.29
1.30 - 1.68
1.69 - 2.07
2.08 - 2.46
2.47 - 2.85
2.86 - 3.24
3.25 - 3.63
3.64 - 4.02
Tannins (mg/g berry CE)
0.35 - 0.39
0.40 - 0.43
0.44 - 0.47
0.48 - 0.51
0.52 - 0.55
0.56 - 0.59
0.60 - 0.63
0.64 - 0.67
Canopy Assessments (EAS)
Averaged Verasion to Harvest 2007
Canopy Assessments Tannins at Harvest 2007
0 50 100 150 20025Meters
Vine vigour Sunraysia Shiraz
Vine vigour
r² = 0.3741
r² = 0.4389
r² = 0.4284
0.00
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8.00
9.00
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7
Phe
no
lics
(mg
/g b
err
y)
Canopy Assessment (EAS)
Anthocyanin
tannin
PhenolicsFeReact
Vineyard variability Sum of normalised Tannins
Lowest
Low
Moderate
High
Highest
Sum of normalised anthocyanins
Lowest
Low
Moderate
High
Highest
Sunraysia Shiraz
2006
2009 2008
2007
Tannins at Harvest
0.90 - 1.29
1.30 - 1.68
1.69 - 2.07
2.08 - 2.46
2.47 - 2.85
2.86 - 3.24
3.25 - 3.63
3.64 - 4.02
Tannins (mg/g berry CE)
0 50 100 150 200 25 Meters
Vineyard variability
Summary Tannin synthesis to veraison
Tannin maturation post-veraison
Measurement
Tannin composition & extractability
Season & site Yield / vigour / vine balance Irrigation Rootstocks Vineyard variability
Simon Robinson CSIRO Plant Industry Flavonoids in grapes
t +61 8 8303 8600 e [email protected] w www.csiro.au
PLANT INDUSTRY
Thank you