BROWNI NG OF WHI T E WI NE S AND AN ACCELERATED TEST

FOR BROWNI NG CAPACI T Y

V. L. Si ngl et on and T. E. Kr aml i ng a
Depar t ment of Vi t i cul t ur e and Enol ogy, Uni ver si t y of Cal i forni a, Davis 95616. ~Pres-
ent address E. & J. Gallo Wi nery, Modest o, Cal i forni a 95353.
The Cal i forni a Wi ne Advi sor y Board is t hanked f or part of the suppor t f or thi s
wor k, and a separate port i on of the research was support ed by the KWV whi l e VLS was
on sabbati cal leave at t he Research I nst i t ut e f or Oenol ogy and Vi t i cul t ure, St el l enbosch,
South Afri ca.
Accept ed f or publ i cat i on Oct ober 4, 1976.
ABST RACT
A st andar di zed t est f or t he br owni ng capaci t y
of whi t e wi nes was developed. A port i on of t he
wi ne is t r eat ed wi t h 1 g of bent oni t e per 100 ml
under n i t r o g e n so t hat t he developed br own pig-
ment will st ay in solution. Por t i ons of t he t r eat ed
wi ne are i ncubat ed in sealed t ubes wi t h 25% or
mor e headspace well spar ged wi t h oxygen and
separ at el y wi t h ni t r ogen for 5 days at 55C. The
absor bance at 420 nm r emai ns near l y unchanged
f or t he N2-sparged sampl es and under oxygen
measur es wi nes' rel at i ve capaci t y to br own oxi-
datively. Evi dence is pr esent ed t hat a wi ne well
prot ect ed f r om oxi dat i on is l argel y pr event ed f r om
br owni ng wher eas a wi ne t r eat ed by addi ng d-care-
chin is consi derabl y i ncreased in its capaci t y to
br own by t hi s test.
Ther e have been a sizable number of st udi es
upon t he br owni ng of whi t e wines. Most have used
ar bi t r ar i l y chosen condi t i ons to produce and mea-
sur e t he brown, wi t hout st udyi ng t he vari abl es.
Ber g (1) used 200 ml of wi ne in a scr ew- capped
bot t l e l ar ge enough to provi de ai r equi val ent to 600
ml 0.,/1. Sampl es wer e held 7 days at 49C bef or e
f i l t er i ng (i f necessar y) and compar i ng absor bance
at 425 nm wi t h t he ori gi nal value. Wr i ght (2)
used absor bance at 470 nm af t er st or age of ver-
mout h sampl es f or 4 mont hs at 2, 20, or 30C to
est i mat e t he rel at i ve br owni ng of di f f er ent wines.
De Vi l l i ers (3) used a por t i on of each wi ne in a
sealed full f l ask at 4C as t he cont rol and measur ed
br owni ng at 425 nm at 10-day i nt er val s af t er st or-
i ng 150 ml of wi ne wi t h 200 ml of ai r at 49C.
Ful l er and Ber g (4) heat ed 20 ml of wi ne plus 12
ml of ai r in closed gl ass t ubes unt i l i ncr ement of
absor bance at 440 nm r eached 0.150 in a 12-mm
col ori met er tube. Caput i and Pet er son (5) com-
oar ed ni t r ogen- st r i pped and oxygen- sat ur at ed bot-
tled wi ne af t er 10 days at 40C or 100 days at room
t emper at ur e. Du Pl essi s and Uys (7) heat ed sampl es
in 30-ml scr ew- capped bottles, l eavi ng 2 ml ai r space
and cent r i f ugi ng if necessary, and de t e r mi ne d ab-
sorbance peri odi cal l y in l - cm cells at 425 nm. Sapi s
and Ri b6r eau- Gayon (8) st ored wi nes wi t h excess
oxygen in sealed bot t l es f or 7 and 21 days at 40C
bef or e meas ur i ng br owni ng at 440 nm. Peri et al.
(9) est i mat ed (450 nm) enzymat i c br owni ng at
5 C and nonenzymi c br owni ng at 50 C in 100 ml
of wi ne in 500-ml conical fl asks f or up to 15 days.
Ber g and Aki yoshi (10) st udi ed t he par amet er s
of br owni ng of whi t e wi nes in some detail. They
showed t hat i ncreases in oxygen up to but not
beyond 60 ml/1 of wi ne gave gr eat er br owni ng, as
did also hi gher pH or t emper at ur e. They concluded
t hat enzymi c oxi dat i on was not si gni f i cant in t he
br owni ng of whi t e wine.
It appear ed to us desi rabl e to i nvest i gat e f ur t he r
t he condi t i ons f or a reproduci bl e accel erat ed t est to
det er mi ne t he capaci t y of a wi ne to brown. If such
a t est could quant i t at e di f f er ences among wi nes and
cl ari fy br owni ng r eact i on conditions, it should be
ver y useful. St andar di zat i on of t est condi t i ons
woul d also be ver y hel pful in compar i ng var i ous
wor ker ' s resul t s.
MAT ERI AL S AND ME T HODS
Whi t e wi nes wi t h alcohol levels of t abl e wi nes
wer e pr epar ed by s t andar d met hods ei t her at Davi s
or St el l enbosch and used in t hese exper i ment s. Ab-
sorbance was det er mi ned at 420 nm wi t h ei t her a
Spect roni c 20 (Davi s) or a Spekker phot omet er
(St el l enbosch) in 18-mm-ID t est tubes. The sampl es
Am. J. Enol . Vi t i c. , Vol . 27, No. 4, 1976
157
158--BROWNING OF WHITE WINES
to be tested for br owni ng were placed in the t est
tubes (7-10 ml) sparged wi t h N2 or 02 gas, f i r ml y
sealed wi t h ei t her a r ubber st opper held in place
by electrician' s plastic t ape under t ensi on or by
rubber-l i ned screwcaps, and heat ed in racks in a
regul at ed oven. The samples were removed period-
ically, cooled to room t emper at ur e, absorbance
measured, and r et ur ned to t he oven.
RESULTS AND DISCUSSION
Oxygen content: For oxidative br owni ng to pro-
ceed as rapi dl y as possible, excess oxygen must be
available t hr oughout the react i on period. For this
reason 02 was used in the headspace r at her t han air.
To be sure t hat 02 t r ansf er into the solution was not
l i mi t i ng t he react i on rate, wine sampl es were
browned wi t h and wi t hout agi t at i on. Agi t at i on was
in a t i ssue-cul t ure rotor, a 2-foot -di amet er wheel
t ur ni ng at 1 r pm at about 60 f r om vertical, versus
the st at i c samples vert i cal in a rack. Exper i ment s
were conducted also upon the mi ni mum headspace
of 02 whi ch gave maxi mum br owni ng rat e and pro-
duced maxi mum br own pi gment levels af t er pro-
longed t r eat ment . Typical dat a are shown in Table
1; this and ot her tests gave no si gni fi cant di ffer-
ences in brown color level in rel at i on to agi t at i on
or 02 headspace as large as or l arger t han 5-ml / 22-
ml -capaci t y tube ( 23%) . Thus, the oxygen con-
t ent and t r ansf er rat e into the wine do not appear
to be in any danger of l i mi t i ng the amount or rat e
of the react i on under the conditions in use here.
Wavelength: The absorbance was measured at
420 nm. This is commonl y measured in red wine,
along wi t h 520 nm absorbance, to est i mat e brown
versus red shades of color and is t hus a usual value
for an enologist. Some browned wines exhibit a
shoulder or broad maxi mum near 445 nm, t hough
not all do so, and such hi gher wavel engt hs would be
more subj ect to i nt erference f r om possible pi nki ng
of l eucoant hocyani di ns. Owi ng to the general fact
t hat browns are "spillover" visible color f r om more
i nt ense absorbance in t he ultraviolet, many wave-
l engt hs in the 400-480-nm range have been used
to measure br owni ng in vari ous products. Never-
theless, 420 nm is pr ef er r ed for wine for the reasons
given and to provide st andardi zat i on.
Sugar, temperature, and amino acids: The effects
of sugar and di fferent t emper at ur es upon br owni ng
of a Char donnay wine are shown in Table 2. Near-
ly identical dat a were obtained wi t h a Thompson
Seedless wine. These dat a showed t hat at 52C or
below, glucose cont ent was not an appreci abl e fac-
t or in accelerated browni ng, whereas at 80C ext ra
br owni ng was caused by the presence of sugar.
Fur t her t est i ng wi t h i nt ermedi at e t emper at ur es
gave oxi dat i ve br owni ng absorbance at 14 days in
Table 2. Effect of temperature, oxygen, and glucose on
browning of a Chardonnay wine.
Absorbance (420 nm) at i n d i c a t e d days
No glucose 10% glucose
C 0 2 8 14 0 2 8 14
Nitrogen-sparged
31 . 180 . 128 . 133 .165
52 .181 . 146 . 160 .210
80 . 183 . 420 . 375 .520
Oxygen-sparged
31 . 180 . 185 . 212 .330
52 . 178 . 230 . 380 .530
80 . 180 . 430 . 690 .900
.180 . 127 . 135 .190
.176 . 138 . 154 .210
.181 .255 . 900 1.50
.180 . 182 . 205 .310
.171 .224 . 360 .530
.183 . 420 1. 50 2.10
Table 3. Effect of amino acid addition on the ability of a
w h i t e wine to brown under nitrogen at 52C.
Table 1. Effect of headspace oxygen and roller tube
a g i t a t i o n on browning of a whi te table wine (18 x 150-mm
t e s t tubes, 52C).
Amino acid added
Absorbance
Amount at 420 nm a
added
(mg/I) 26 days 44 days
Headspace
Absorbance at 420 nm
Agitated 0 days 3 days 7 days
5 ml no .133 .270 .460
10 ml no .133 .310 .450
15 ml no .133 .280 .460
5 ml yes .133 .290 .460
10 ml yes .133 .300 .480
15 ml yes .133 .300 .480
Control - - .278 .355
L-arginine monohydrochloride 250 .295 .360
L-glutamic acid 250 .292 .362
Glycine 250 .270 .368
L-histidine monohydrochl ori de 250 .290 .390
monohydrate
L-proline 500 .310 .380
L-hydroxyproline 500 .278 .365
L-threonine 250 .330 .365
Absorbance at 0 days for all samples was .175.
A m . J. Enol . V i t i c . , V o l . 27, No. 4, 1976
BROWNI NG OF WHI TE WI NES- - 159
Char donnay plus 10% glucose t hat at 31C was
1.06 t i mes t he absor bance of t he same wi ne wi t hout
glucose, 1.00 t i mes at 52C, 1.24 t i mes at 65C,
1.39 t i mes at 72C, and 2.88 t i mes at 80C. It t hus
appear s t hat s ugar is of negl i gi bl e ef f ect at 55C
or below but is i ncr easi ngl y i mpor t ant as t emper a-
t ur e rises, par t i cul ar l y at 80C or hi gher. Thi s
car amel i zat i on ef f ect is evi dent also under ni t r ogen
(Tabl e 2). The addi t i on of ami no aci ds under
oxi dat i ve condi t i ons or under ni t r ogen (Tabl e 3)
gave little or no i ncr ease in br owni ng in a dr y
t abl e wi ne over t hat in t he controls.
Be nt oni t e t reat ment : Pr ot ei ns or pol ypept i des
evi dent l y i nt er f er e in br owni ng meas ur ement s by
r eact i ng wi t h and pr eci pi t at i ng t he br own pi gment .
That is pr event ed by addi ng a rel at i vel y heavy dose
of bent oni t e. To avoi d di l ut i on probl ems t he ben-
t oni t e was added dr y to t he wi ne sample, mi xed
in unt i l t hor oughl y wet t ed, and t he wi ne held under
N2 at room t e mpe r a t ur e f or about 2 hour s wi t h
occasional shaki ng. The wi ne was t hen f r eed of
bent oni t e by cent r i f ugat i on and f i l t r at i on t hr ough
a membr ane (0. 45-mi cron) fi l t er. Fig. 1 shows
t hat wi t hout bent oni t e t he ma xi mum br own color
is less and i ncreases r api dl y as t he bent oni t e level
is i ncr eased to 0.384 g/ 100 ml (32 l b/ 1000 gal ) but
ver y little beyond. Levels as hi gh as 3.84, 7.68, and
15.36 g of bent oni t e/ 100 ml of wi ne gave little or
no f ur t he r i ncr ease in br owni ng r at e or ext ent in
any wi ne t est ed except as t he pH began to be in-
creased sl i ght l y at t hese ext r eme levels. The ben-
0. 8
E
C
o 0. 6
N
w
o
z
= 0 . 4
n-
O
o~
I I I I I I I
I . . ~ - - " - - - - - - ~ F
s o
I i
/
J A
J
.f
A 1
J
J
0 . 2
[ I I I I I I I
" 0 e ,2 ,e 2 4
DAYS AT 52" C
Fig. 1. The ef f ect on br owni ng caused by p r i o r t r e a t m e n t
o f a w h i t e wi ne wi t h d i f f e r e n t l e v e l s o f b e n t o n i t e , a -- 0 ben-
t oni t e, b = 8 I b/ 1000 gal (0.1 g/ 100 ml ), c = 16 I b/ 1000 gal
(0.19 g/ 100 m l ) , d = 32 I b/ 1000 g a l ( 0 . 3 8 g / 1 0 0 m l ) , e = 6 4
I b/ 1000 gal (0.77 g/ 100 ml ), f = 128 I b/ 1000 gal (1.54 g/100
ml ).
tonite t r e a t m e n t s did n o t a f f e c t the total p h e n o l
cont ent appreci abl y, e.g., an unt r eat ed wi ne wi t h
163 mg GAE/ 1 t ot al phenol assayed 160 mg GAE/ 1
af t er 3.84 g/ 100 ml of bent oni t e and 168 mg
GAE/ 1 af t er 7.68 g. Fur t he r t est s i ndi cat ed t hat
3.84 g of bent oni t e per 100 ml may l ower whi t e
wi ne t ot al phenol about 5 %, and t hat 7.68 g/ 100 ml
usual l y l owered it about 10%. Consi der i ng t hat
di f f er ent lots of bent oni t e may di f f er consi derabl y,
t he t r e a t me nt should be as hi gh as possible to allow
f or less effect i ve bent oni t e. Fig. 1 shows t hat levels
of 0.38, 0.77, and 1.54 g/ 100 ml give si mi l ar resul t s,
and t he ot her t est s showed t hat t hese should have
little or no effect on t ot al phenol cont ent or pH.
For r out i ne use, 1 g of bent oni t e per 100 ml of
wi ne is recommended.
R e p r o d u c i b i l i t y : If no bent oni t e is used, some
wi nes produce pr eci pi t at es of br owned mat er i al and
t he devel opment of br own in solution is decreased
(Fi g. 1). If heavi l y bent oni t ed, t hese wi nes pro-
duce br own pi gment at a uni f or m r api d r at e (Fi g.
1) but event ual l y r each a maxi mum of soluble br own
pi gment . In some sampl es t he br own pi gment t hen
begi ns to decrease by pol ymer i zat i on and pr eci pi t a-
tion. The maxi mum br own is a reproduci bl e val ue
if ei t her no pr eci pi t at i on occurs or t he br own pre-
ci pi t at e is dissolved by accur at e dilution wi t h 60%
aqueous et hanol at pH 3.3. Tabl e 4 shows t hat re-
produci bl e val ues wer e obt ai ned in t r i pl i cat e t r i al s
wi t h t wo di f f er ent wines.
Wi t h wi nes havi ng consi derabl y di f f er ent br own-
i ng r at es t he t i me to r each ma xi mum br own in t hese
t est s can be di f f er ent (Fi g. 2) besides r equi r i ng
t wo weeks or mor e for a test. Dur i ng t he f i r st day
or t wo of heat i ng of a heavi l y bent oni t ed whi t e
wi ne in t he presence of oxygen t he wi ne may even
decrease in brown, may r emai n unchanged, or, mor e
commonly, may brown, t hough at a di f f er ent r at e
t han l at er (Fi g. 2). Fr om about 3 to 11 days, how-
ever, t he r at e of br own absor bance i ncrease is
T a b l e 4. R e p r o d u c i b i l i t y o f t h e a c c e l e r a t e d m e t h o d u s e d t o
i nduce br owni ng in whi t e wi nes (0.77 g bent oni t e/ 100 ml ,
52C).
Absor bance at 420 nm a
S a m p l e 0 days 21 days 27 days
W h i t e Ri esl i ng .167 1.30 1.32
W h i t e Ri esl i ng .165 1.30 1.31
W h i t e Ri esl i ng .171 1.28 1.31
Ch a rd on nay .240 1.19 1.18
Cha rdon nay .241 1.20 1.20
Cha rdon nay .228 1.21 1.20
a For 21 and 27 days t h e a b s o r b a n c e v a l u e i s c a l c u l a t e d
f r o m t h e a b s o r b a n c e r e a d i n g o f o n e p a r t s a m p l e d i l u t e d wi t h
t h r e e p a r t s o f 60% aqueous et hanol adj ust ed t o pH 3.3.
A m . J . E n o l . V i t i c . , V o l . 2 7 , N o . 4 , 1 9 7 6
160--BROWNI NG OF WHI TE WI NES
E
0
OJ
bJ
L)
z 3
< "
m
n -
O
0 3
r n
<
.1
. , J ~"
~ X I ~ Y ' 1 ~ ' "
I I I i I I I I i I I I I I I 1,5
5 I0
DAYS OX I DA T I ON ( 5 5C. )
Fig. 2. Br owni ng of t hree di f f er ent wi nes wi t h t i me under
oxygen.
essentially linear (Fig. 2) wi t h time for each wine.
Thus the browni ng capacity of di fferent wines can
be estimated by comparing t hei r relative brownness
at a constant time. We chose five days.
B r o w n i n g t est : The chosen procedure, then, for
comparing browni ng capacity is" mix 100 ml of
the wine thoroughly with 1.0 g of dry Wyoming
bentonite, stopper it under nitrogen, and wai t (with
occasional shaking) for at least 2 hours. Settle and
centrifuge, then filter t hrough a 0.45-micron mem-
brane filter enough to prepare four replicate test
tubes. We have used 7 ml in 18 x 150-mm test
tubes. Sparge two of the tubes thoroughly with N2
free of 02, and the other two with 0~. Seal each
tube for the duration of the experiment with taped-
down rubber stoppers or t i ght screw caps as the
spargi ng tip is removed. Hold at 55C for 5 days
and read the brown absorbance developed in each
tube at 420 nm. If desired, the sample can be held
until maxi mum brown is reached, and t hat de-
termined separately.
M i n i ma l b r o w n i n g u n d e r ni t r ogen" The nitrogen-
sparged samples serve as controls indicating any
nonoxidative browning. Duplicates are recom-
mended because samples t hat are completely de-
pleted of oxygen by nitrogen generally do not
brown. If more t han one sample is N2-sparged, an
incorrectly high sample owing to incomplete re-
moval of 02 or loosening of a stopper can usually
be detected. The 02-treated samples usually dupli-
cate each other very closely, but, again, can indicate
leaks. Under the conditions indicated, the browni ng
of dry white wines is almost completely oxidative.
This is illustrated by the fact t hat a series of 5 dif-
ferent wines heated under N2 at 60C gave original
and 8-day absorbances at 420 nm of, respectively,
0.058-0.057, 0.107-0.123, 0.086-0.097, 0.074-0.084,
and 0.094-0.100. The same wines under 02 for the
same t reat ment gave absorbances of 0.414-0.712. Fur-
thermore, if a sample browni ng under 02 is then
sparged wi t h N2 and reheated, the increase in
W
0
Z
< 0. 5
O
( D
m
< C
b
m
O ~ " =
o
I I
B ...._-..o
v
5 I 0 15
D A Y S AT 5 2 C
Fig. 3. Increase in br owni ng caused by the addi t i on of
d-cat echi n to a whi t e wi ne. a - Chardonnay wi ne alone, b -
200 ppm d-cat echi n, c - Char donnay wi ne plus 200 ppm d-
cat chi n.
brown ceases and, in fact, the absorbance may de-
crease slightly.
Pr a c t i c a l a p p l i c a t i o n s : Caputi and Peterson (5)
did not find good correlation between their acceler-
ated browni ng test and wine browning on the shelf.
That may very well be t rue of the test procedure
suggested here. Nevertheless, the test does distin-
guish among wines with differing capacity to brown.
As an example, Fig. 3 shows the browni ng of a
Chardonnay wine and the same wine with 200 ppm
of d-catechin added, as mi ght be the case with in-
creased pressing for example.
L I T E R A T U R E C I T E D
1. Berg, H. W. Vari etal suscept i bi l i t y of whi t e wi nes to
br owni ng. I1. Accel erat ed st orage tests. Food Research 18:
407-10 (1953).
2. Wr i ght , D. Factors af f ect i ng the col or of dry ver mout h.
Am. J. Enol. Vi ti c. 11:30-34 (1960).
3. de Vi l l i ers, J. P. The cont rol of br owni ng of whi t e t abl e
wi nes. Am. J. Enol. Vi t i c. 12:25-30 (1961).
4. Fuller, W. L., and H. W. Berg. Tr eat ment of whi t e wi ne
wi t h Nyl on 66. Am. J. Enol. Vi ti c. 16:212-18 (1965).
5. Caputi , A., Jr., and R. G. Peterson. The br owni ng prob-
lem in wi nes. Am. J. Enol. Vi ti c. 16:9-13 (1965).
6. Peterson, R. G., and A. Caputi , Jr. The br owni ng prob-
lem in wi nes. I1. Ion exchange effects. Am. J. Enol. Vi t i c.
18:105-12 (1967).
7. du Plessi's, C. S., and A. L. Uys. Br owni ng in whi t e
wi nes. I1. The ef f ect of cul t i var, f er ment at i on, husk, seed, and
stem cont act upon br owni ng. S. Af r. J. Agri c. Sci. 11:637-48
(1968).
8. Sapis, J. C., and P. Ri b~reau-Gayon. Etude de bruni sse-
ment des vi ns blancs. I. Tr ansf or mat i on des compos~s ph~no-
l i ques au cours du bruni ssement . Connai ssance Vi gne Vi n
2:323-48 (1968).
9. Peri, C., C. Pompei , G. Mont edor o, and C. Cantarel l i .
Maderi zat i on of whi t e wi nes. I. I nf l uence of pressi ng of t he
grapes on the suscept i bi l i t y to oxi dat i ve br owni ng. J. Sci.
Food Agri c. 22:24-8 (1971).
10. Berg, H. W., and M. Aki yoshi . Some f act ors i nvol ved in
br owni ng of whi t e wi nes. Am. J. Enol. 7:1-7 (1956).
Am. J. Enol . Vi t i c. , Vol . 27, No. 4, 1976