Download Maillard Browning in Ethanolic Solution

JFS:
Food Chemistry and Toxicology
Maillard Browning in Ethanolic Solution
ABSTRACT: Maillard browning in ethanolic solutions of a 0.2 M glucose–0.2 M glycine (G-G) system in a heat
treatment becomes more pronounced with an increase in ethanol concentration (0% to 50%, v/v). Extents of
browning, as measured by absorbance at 420 nm, of a fructose-glycine system in aqueous and 50% ethanolic
solutions buffered at pH 4.3 are very close after the treatment, whereas that of the G-G system in 50% ethanolic
solution is much enhanced. A higher hydroxymethylfurfural (HMF) content is present in the ethanolic G-G
system, suggesting the involvement of ethanol in the G-G reaction to form HMF. Evidence also shows that
ethanol inhibits the formation of HMF in an ethanolic solution containing glucose alone. Much more HMF is
formed in the pH 4.3 buffered G-G system in an ethanolic solution at a water activity (aw) above 0.81 or at an
ethanol concentration below 30% than in a glycerolic solution at the same aw. We propose that the reduction of aw
is not the sole major mechanism for ethanol to accelerate HMF formation and Maillard browning in G-G solution.
Keywords: Maillard reaction, browning, ethanol, hydroxymethylfurfural, water activity
Introduction
T
he Maillard reaction is a class of nonenzymatic browning that
involves the interaction of reducing sugars and free amino acids or a free amino group of an amino acid that is a part of the protein chain (BeMiller and Whistler 1996). The factors that influence
the rate of the Maillard reaction in the processing and storage of
food include the composition of food, the time-temperature condition, pH, water activity (aw), oxygen tension, the presence of promoters, and inhibitors, and so forth (Ames 1990, 1998). Conventionally, research on the Maillard reaction has used water as the
solvent. Studies in nonaqueous systems are rare.
In aqueous systems, the key chemical components that directly
influence the progress of the Maillard reaction are sugars and amino acids. Del Pilar Buera and others (1987) compared the activities
of various monosaccharides and disaccharides below pH 6 and
found that they are xylose, fructose, glucose, lactose, maltose, and
sucrose in descending order. Brands and others (2000) investigated
the Maillard reaction in sugar-casein system and found that those
sugars having a higher proportion of acyclic forms in the solution
brown more rapidly. They also found that aldoses brown slower
than their corresponding ketoses and that nonreducing disaccharides brown even slower because they need to be hydrolyzed to
reducing sugars first. Ashoor and Zent (1984) compared the rate of
Maillard browning among common amino acids and grouped them
into high, intermediate, and low browning producing groups. Higher temperature and longer time result in more serious browning
(Del Pilar Buera and others 1987; Lee and Nagy 1988). In alkaline
condition, both of the browning rates in fructose-glycine and glucose-glycine systems reach maximum at pH 10 (Ashoor and Zent
1984). In the pH range between 3.4 and 7.7, the browning in starchglucose-lysine mixture increases with the increase in pH (Bates and
others 1994). Renn and Sathe (1997) showed that the browning rate
between L-leucine and D-glucose is positively correlated with pH.
MS 20030383 Submitted 7/6/03, Revised 9/24/03, Accepted 1/29/04. Authors
Shen and Wu are with the Inst. of Food Science and Technology, Natl. Taiwan
Univ., P.O. Box 23-14, Taipei 106, Taiwan. Direct inquiries to author Wu (Email: [email protected]).
© 2004 Institute of Food Technologists
Further reproduction without permission is prohibited
Research in Maillard browning regarding the effect of aw (Toribio
and Lozano 1984; Cuzzoni and others 1988), organic acids (Akhavan
and Wrolstad 1980), metal ions (Petriella and others 1988), and high
pressure (Hill and others 1996) have also been reported. These
reviewed studies were all done in aqueous systems to stand for
common foods. However, some foods, such as beer, sake, rice wine,
many fortified wines, and liqueurs are types of ethanolic solutions
that contain noticeable amounts of Maillard reaction substrates.
The investigation of Maillard browning in ethanolic solution shall
be worthwhile.
Materials and Methods
Reagents
The sugar species tested in the present study covered pentose
(xylose), hexoses (glucose and fructose), monosaccharides (xylose,
glucose, fructose, and galactose), disaccharides (sucrose, lactose,
and lactulose), aldoses (glucose and lactose), and ketoses (fructose
and lactulose). All the sugars, amino acids, and hydroxylmethylfurfural (HMF) are the products of Sigma Co. (St. Louis, Mo., U.S.A.).
Ethanol (95% v/v) is purchased from Taiwan Tobacco & Liquor Co.
(Taipei, Taiwan).
Preparation of reaction mixtures
Among all the model systems for Maillard reaction, hexose- and
pentose-glycine systems are the most frequently used (Wedzicha
and Kaputo 1992). The present study uses glucose (0.2 M)-glycine
(0.2 M) (G-G) model solutions to investigate the influence of ethanol
concentration and pH on browning. The G-G solutions were prepared
by dissolving glucose and glycine in 0.05 M succinic acid–sodium
hydroxide buffered 0% to 50% ethanolic solutions. The pH value was
adjusted to 4.3 to represent sake or to 5.5 to represent a liqueur made
from spirit and a common low acid food (Leong and Wedzicha 2000).
Aliquots (4 mL) of the model solutions were transferred to 4.5-mL glass
vials, hermetically sealed, and heated in boiling water for 0 to 6 h as
an accelerated storage test to simulate the storage at room temperature for approximately 8 mo. Glucose and glycine were also dissolved in ethanolic solutions without buffering for comparison.
Vol. 69, Nr. 4, 2004—JOURNAL OF FOOD SCIENCE FCT273
Published on Web 4/15/2004
Food Chemistry and Toxicology
S.-C. SHEN AND J.S.-B. WU
Maillard browning in ethanolic solution . . .
Food Chemistry and Toxicology
Xylose, fructose, galactose, sucrose, lactose, or lactulose was
then substituted for glucose in the preparation of solutions containing 0%, 15%, and 50% ethanol to investigate the browning potential of different sugars in an ethanolic system. Similarly, lysine,
threonine, cysteine, serine, or alanine was substituted for glycine
in the preparation of ethanolic solutions for investigating the effect
of amino acid species on Maillard browning. The 0%, 15%, and 50%
ethanol concentrations were chosen to represent the ethanol concentrations in aqueous solution, sake, and liqueur, respectively.
Water used in the present study was obtained from a Milli-Q system
(Millipore, Osaka, Japan).
In comparison, glycerolic solutions were prepared by dissolving
either 0.2 M glucose only or 0.2 M glucose-0.2 M glycine in aqueous
solutions of glycerol (0% to 54.3%, w/w) buffered with 0.05 M succinic acid-sodium hydroxide at pH 4.3.
Evaluation of aw
The aw of all the ethanolic and glycerolic solutions were evaluated by the method described by Ferro Fontan and Chirife (1981)
and Miracco and others (1981), with minor modifications. The freezing point of each solution was measured using a differential scanning calorimeter (DSC) (Model DSC 822e, Mettler-Toledo GmbH,
MatChar, Switzerland). Purified nitrogen (99.999%) flowed at 80
mL/min and 200 mL/min were the purge gas and the dry gas, respectively. Samples of 14 to 18 mg were weighed into aluminum
pans, and covers were hermetically sealed into place. An empty,
hermetically sealed aluminum pan was used as the reference. Before the analysis of samples, the baseline was obtained with the
reference pan. After both the sample pan and the reference pan
were placed in the chamber at 25 °C, the cell block of the DSC was
cooled to –50 °C at 10 °C/min and held there for 2 min, followed by
heating at a rate of 5 °C/min to 25 °C. The heating thermograms of
samples showed a single melting peak. The peak temperature and
the enthalpy was obtained by analyzing the thermogram with
Mettler Toledo Stare Software V6, 1.0. The freezing point is defined
as the intersection of baseline and the tangent line of melting
peak.
The aw of a sample was calculated using the following equation:
–ln aw = 9.6934 × 10–3 × ␶F + 4.761 × 10–6 × ␶F2
where ␶F (freezing point depression) = To – TF; To = freezing point of
pure water; TF = freezing point of sample solution.
system to identify the corresponding retention time and elution
profile in the chromatogram of the browning sample solutions. The
content of HMF was analyzed adapting the method reported by
Lee and Nagy (1988) by the constructed calibration curve of HMF
(0.05 to 10 mg/mL) (y = 42849x + 1510.8; R2 = 0.999). Triplicate samples were prepared, and each sample was injected 3 times. Each
experiment was repeated 4 times. The comparison among the values of optical absorbance or HMF content in different treatments
was done by analysis of variance and Duncan’s multiple range test
(P ⱕ 0.05).
Results and Discussion
Effect of ethanol concentration on pH and browning
in nonbuffered G-G solutions
The effect of ethanol concentration on the pH of nonbuffered GG solutions is shown in Figure 1. In the range of 0% to 50% ethanol
content, a higher ethanol concentration corresponds to a higher
initial pH value, in a well-adjusted quadratic relation (R2 = 0.985).
Gutiérrez (2003) reported similar results in reconstituted wines
containing 0% to 22% ethanol.
The pH value in G-G solutions decreases with an increase in
heating time (Figure 2a), similar to the findings in aqueous glucose-glycine solutions reported by Morales and Jimenez-Perez
(2001). Among all the samples heated for the same duration, the
one containing no ethanol shows the lowest pH value. The decrease in pH observed during the Maillard reaction can be attributed to the degradation of sugar into acid (Beck and others 1990) or
the condensation between the free amino group of amino acid and
the carbonyl group of glucose (Trifiro and others 1990).
The effect of ethanol concentration on browning (A420nm) of the
nonbuffered G-G solutions during incubation is shown in Figure
2b. It is noteworthy that browning is more severe in G-G ethanolic
solutions containing a higher level of ethanol, revealing the involvement of ethanol in the glucose-glycine browning reaction,
which has never been reported before.
Effect of pH on browning in ethanolic solutions
The effect of ethanol concentration on the absorbance at 420 nm
in G-G ethanolic solutions buffered at pH 4.3 and pH 5.5 by 0.05 M
succinic acid–sodium hydroxide is shown in Figure 3. At pH 4.3, the
extent of browning increases with the increase in ethanol concentration. After heating in boiling water for 6 h, the absorbance at 420
Absorbance spectra of the solutions
The absorbance spectra of solutions were read using a UV/VIS
8500 double-beam spectrophotometer (Lab Alliance, State College,
Pa., U.S.A.). The absorbance at 420 nm was taken as a measure of
browning (Baxter 1995; Ajandauz and Puigserver 1999).
Analysis for HMF content
The solutions were heated in boiling water for 6 h, followed by
filtration through a Millex-HN nylon clarification kit with a 0.45-␮m
pore size (Millipore, Bedford, Mass., U.S.A.). The filtrate was injected into a high-performance liquid chromatographic (HPLC) system
that consisted of a Luna 5␮ C18 (2) Column (Phenomenex, Torrance, Calif., U.S.A.), a Series III Pump (Lab Alliance), a UV-970
Intelligent UV/VIS Detector (Jasco Co., Tokyo, Japan), and a Chem
Lab Data Station Integrator (Scientific Information Service Co.,
Taipei, Taiwan). The quantity of injection was 20 ␮L. The mobile
phase was acetonitrile-water-acetic acid in a 10:89.5:0.5 volume
ratio at a flow rate of 1.0 mL/min. The detected wavelength was 280
nm. Standard HMF (Sigma) was injected into the chromatographic
FCT274
JOURNAL OF FOOD SCIENCE—Vol. 69, Nr. 4, 2004
Figure 1—Plot of pH compared with ethanol content for
glucose (0.2 M)/glycine (0.2 M) solutions
URLs and E-mail addresses are active links at www.ift.org
Maillard browning in ethanolic solution . . .
Table 1—Absorbance at 420 nm of the ethanolic solutions of glycine (0.2 M)/sugar (0.2 M) buffered at pH 4.3 with 0.05
M succinic acid–sodium hydroxide and heated in boiling water for 6 ha,b
Ethanol
conc. (v/v)
0%
15%
50%
Glucose
Fructose
Lactose
Lactulose
Sucrose
Galactose
Xylose
0.26 ± 0.01ey
0.25 ± 0.01dy
1.29 ± 0.02bx
1.14 ± 0.01bx
0.86 ± 0.01by
1.14 ± 0.01bx
0.23 ± 0.01ex
0.17 ± 0.01dy
—c
0.56 ± 0.01dx
0.47 ± 0.01cy
0.49 ± 0.02cy
0.25 ± 0.03ey
0.20 ± 0.01dz
0.43 ± 0.02cx
0.75 ± 0.02cy
0.77 ± 0.01by
0.97 ± 0.07bx
6.28 ± 0.08az
10.94 ± 0.28ay
33.51 ± 2.04ax
a Values shown are the average of 3 replicates.
b Different letters (a–e) within the same row are significantly different ( P < 0.05) in Duncan’s multiple range test. Different letters (x–z) within the same column
nm of G-G system in 50% ethanolic solution reaches 1.03, approximately 3.8 to 3.6 times the absorbance value of 0.27 to 0.29 in the
0% to 20% ethanolic solutions. For samples with the same ethanol
level, more severe browning occurs at pH 5.5 than at pH 4.3. This
suggests that a high pH is favorable for the browning reaction in
ethanolic solution, as has been reported in aqueous solution (Ames
1998).
Table 1 presents the absorbance at 420 nm in 0%, 15%, and 50%
ethanolic solutions of 0.2 M glycine-0.2 M sugar buffered at pH
4.3 with 0.05 M succinic acid–sodium hydroxide after the heat
treatment. Regardless of the species of sugar, the extent of browning is more pronounced in ethanolic solution than in aqueous
solution. The data for lactose are not available because of the low
solubility of this sugar in the solution. Xylose, the most potent
browning-inducing sugar species in the glycine-sugar aqueous
system (Del Pilar Buera and others 1987), is also proven to be
most active among the sugars tested in 50% ethanolic solution
with an absorbance reading at 420 nm of 33.5, about 5 times of
that in aqueous solution (Table 1). This shows that ethanol enhances browning, as also indicated by Figure 2 and 3. Interestingly, the readings of absorbance at 420 nm for glucose in aqueous
and 50% ethanolic solution are about 0.26 and 1.29, respectively,
whereas those of fructose in both systems had the same value of
1.14 after heating (Table 1). Restated, glucose as an aldose browns
less severely than its ketose isomer, fructose, in aqueous solution
and in 15% ethanol, but more severely in 50% ethanol. It appears
that the concentration of ethanol may influence the reactivity of
some sugar species in the formation of Maillard reaction intermediates.
Figure 2—Effect of ethanol concentration on (a) the pH and
(b) the absorbance at 420 nm in nonbuffered glucose (0.2
M)/glycine (0.2 M) solutions during heating in boiling water
Figure 3—Effect of ethanol concentration on the absorbance at 420 nm in glucose (0.2 M)/glycine (0.2 M) solutions buffered at pH 4.3 and 5.5 by 0.05 M succinic acid–
sodium hydroxide during heating in boiling water
Effect of sugar species on browning in ethanolic
solutions
URLs and E-mail addresses are active links at www.ift.org
Vol. 69, Nr. 4, 2004—JOURNAL OF FOOD SCIENCE
FCT275
Food Chemistry and Toxicology
are significantly different ( P < 0.05) in Duncan’s multiple range test.
c Insoluble in 50% (v/v) ethanol.
Maillard browning in ethanolic solution . . .
Table 2—Absorbance at 420 nm of the ethanolic solutions of glucose (0.2 M)/amino acid (0.2 M) buffered at pH 4.3
with 0.05 M succinic acid–sodium hydroxide and heated in boiling water for 6 ha,b
Ethanol
conc. (v/v)
0%
15%
50%
Alanine
Cysteine
0.10 ± 0.01cy
0.07 ± 0.01cy
0.32 ± 0.04cx
0.08 ± 0.01cz
0.15 ± 0.02cy
0.59 ± 0.05cx
Glycine
Serine
0.24 ± 0.02cy
0.22 ± 0.02bcy
1.00 ± 0.02cx
0.28 ± 0.04cy
0.27 ± 0.01bcy
1.03 ± 0.05cx
Threonine
Lysine
0.64 ± 0.02bz
0.79 ± 0.02by
2.52 ± 0.09bx
3.23 ± 0.40az
5.73 ± 0.79ay
15.94 ± 0.42ax
a Values shown are the average of 3 replicates.
b Different letters (a–e) within the same row are significantly different ( P < 0.05) in Duncan’s multiple range test. Different letters (x–z) within the same column
are significantly different (P < 0.05) in Duncan’s multiple range test.
Food Chemistry and Toxicology
Effect of amino acid species on browning in ethanolic
solutions
The amino acids tested beside glycine were lysine, alanine,
serine, threonine, and cysteine. Table 2 shows the browning in ethanolic solutions of 0.2 M glucose and 0.2 M amino acid buffered at
pH 4.3. Among all the samples, the most severe browning occurred
in glucose-lysine solutions. In 15% and 50% ethanolic solutions of
glucose-lysine, the readings of absorbance at 420 nm are 5.73 and
15.94, respectively, which are about 1.8 and 4.9 times the value of
3.23 in aqueous solution. Similar browning-enhancing effects are
observed when other amino acids react with glycine in ethanolic
solutions at 50% ethanol concentration. This phenomenon is consistent with the tendency of discoloration in liqueur. It is well recognized that the carbonyl group of glucose condenses most easily
with the ⑀-amino group of lysine in aqueous solutions to form Nsubstituted glycosylamine in the initiation stage of Maillard browning (Van Martins and others 2001). The same thing would be true in
ethanolic solutions, with more pronounced browning reaction at
high ethanol concentrations (Table 1 and 2).
Mechanisms are not clear. However, fructose browns faster than glucose in aqueous system because of the higher proportion of the acyclic form (Davies and others 1998). It is possible that ethanol increases the concentration of the acyclic form of glucose.
Reduction of aw by ethanol and its effect on Maillard
browning
Ethanol, a highly volatile solvent, would interfere with any direct
instrumental measurement for aw. Therefore, we measured the
freezing point depression and calculated aw. Eichner and Karel
HMF content in ethanolic solutions
Figure 4 provides evidence for the involvement of ethanol in the
browning of ethanolic solutions containing either 0.2 M glucose
alone or G-G system. An absorption peak occurs at 280 to 290 nm.
The height of the absorption peak in the heated and diluted G-G
solutions decreases with the decreasing ethanol concentration,
from about 2.42 at 288 nm in 50% ethanol to about 0.62 in deionized
water (Figure 4a). In comparison, the height of the absorption peak
at 286 nm in the solution containing 0.2 M glucose alone decreases
with the increasing ethanol concentration, from about 0.57 in aqueous solution to 0.27 in 50% ethanolic solution (Figure 4b). In the
system containing glucose alone, ethanol suppresses the absorbance at 280 to 290 nm (Figure 4b). However, in G-G ethanolic solutions, higher absorbance readings correspond to higher ethanol
concentrations (Figure 4a). At pH 4.3, hexoses may react with amino
acids to form Amadori rearrangement products, which then undergo 1,2-enolisation to form HMF as a key intermediate in the Maillard browning (Tress and others 1995). The maximum absorbance
in the spectrum of HMF occurs at approximately 280 nm (Lee and
Nagy 1988). Accordingly, we recognize the occurrence of a dominant absorption peak at 280 to 290 nm in HPLC analysis as evidence for HMF formation (Figure 5). HMF is the dominant peak in
the chromatogram for all the samples tested (Figure 5), revealing
this compound as the main intermediate in ethanolic solutions in
the present browning reaction.
An increase in ethanol concentration favors the formation of HMF
in the G-G system more than that in the fructose-glycine system.
The content of HMF in the G-G system starts to surpass that in the
fructose-glycine system before the ethanol concentration reaches
50% (Figure 6). Changes in browning with increasing level of ethanol
as shown in Figure 3a and Table 1 are consistent with these findings.
FCT276
JOURNAL OF FOOD SCIENCE—Vol. 69, Nr. 4, 2004
Figure 4—Absorbance spectra of (a) glucose (0.2 M)/glycine (0.2 M) and (b) glucose (0.2 M) solutions buffered at
pH 4.3 with 0.05 M succinic acid–sodium hydroxide, heated
in boiling water for 6 h and then diluted 8-fold
URLs and E-mail addresses are active links at www.ift.org
Maillard browning in ethanolic solution . . .
Figure 5—High-performance liquid chromatographic patterns of glucose (0.2 M)/glycine (0.2 M) and fructose (0.2
M)/glycine (0.2 M) in pH 4.3, 0.05 M succinic acid–sodium
hydroxide buffered solutions after heating in boiling water
for 6 h
URLs and E-mail addresses are active links at www.ift.org
Table 3—Water activity (aw)a values of pH 4.3, 0.05 M succinic acid–sodium hydroxide buffered ethanolic or
glycerolic solutions of glucose (0.2 M) alone or glucose (0.2
M)/glycine (0.2 M)
Ethanol
conc.
glucose
(v/v)
only
0%
10%
15%
20%
30%
40%
50%
0.98
0.96
0.93
0.90
0.86
0.83
0.76
aw
glucoseglycine
0.97
0.91
0.89
0.87
0.81
0.77
0.70
aw
Glycerol
conc.
(w/w)
glucose
only
glucose
glycine
0%
13.9%
20.9%
27.9%
41.3%
54.3%
0.98
0.91
0.87
0.83
0.76
0.73
0.97
0.89
0.86
0.82
0.73
0.67
a Calculated from Ferro Fontan and Chirife (1981) equation.
in both the glycerolic solutions of glucose alone (Figure 7a) and GG (Figure 7b). However, a lower aw value may correspond to either
a lower HMF content in the ethanolic solution of glucose alone (Figure 7a) or a higher HMF content in the ethanolic solution of G-G
(Figure 7b). Restated, ethanol has a net inhibitory effect on HMF
formation in the system containing glucose alone. The same phenomenon is also shown in Figure 4b. The mechanism of inhibition
may be as follows: the hydroxyl group of an ethanol molecule reacts
with the carbonyl group of a glucose molecule, loses 1 molecule of
water, and forms 1 chemically stable molecule of ethyl ␤-D-glucopyranoside (BeMiller and Whistler 1996), thus avoiding the 3-deoxyglucosone pathway and blocking the formation of HMF.
The trends in the formation of HMF in both glycerolic solution
and ethanolic solution of the G-G system are presented in Figure
7b. The HMF content in glycerol solution changes from 4.38 mg/100
mL at aw 0.82 to 8.95 mg/100 mL at aw 0.67, having an increment of
104%. Interestingly, in a similar change of aw, from 0.81 in 30% ethanol to 0.70 in 50% ethanol, the HMF content shifts from 6.05 mg/
100 mL to 19.41 mg/100 mL, having an increment of 221%. At an aw
of approximately 0.70, the HMF content in the ethanolic solution is
more than double that in the glycerolic solution. It is evident that
Figure 6—The relationship between ethanol concentration
and HMF content in pH 4.3, 0.05 M succinic acid–sodium
hydroxide buffered ethanolic solutions of fructose (0.2 M)/
glycine (0.2 M) and glucose (0.2 M)/glycine (0.2 M) after
heating in boiling water for 6 h
Vol. 69, Nr. 4, 2004—JOURNAL OF FOOD SCIENCE
FCT277
Food Chemistry and Toxicology
(1972) added glycerol to an aqueous solution to investigate the
effect of aw reduction on Maillard browning. We substituted ethanol with glycerol in the solutions containing either 0.2 M glucose
alone or the G-G system and then evaluated the aw for the same
reason. The aw values of aqueous, ethanolic, and glycerolic solutions are shown in Table 3. It appears that the presence of ethanol
reduces aw noticeably. For example, the aw values of aqueous solutions of 0.2 M glucose alone and G-G are 0.98 and 0.97, respectively,
whereas those of 50% ethanolic solutions containing the same solutes are reduced to 0.76 and 0.70. The ranges of aw values in the
glycerolic solutions containing 0.2 M glucose alone and G-G are 0.91
to 0.76 and 0.89 to 0.67, respectively. Each covers the range of aw
values of the ethanolic solution.
Figure 7 shows the relationship between aw and HMF content in
pH 4.3, 0.05 M succinic acid–sodium hydroxide buffered ethanolic
and glycerolic solutions of 0.2 M glucose alone and the G-G system
after the heat treatment. An increase in the concentration of ethanol causes a reduction of aw value in the ethanolic model solutions.
As expected, a lower aw value corresponds to a higher HMF content
Maillard browning in ethanolic solution . . .
Food Chemistry and Toxicology
the reduction of aw is not the sole major mechanism responsible for
the acceleration in the Maillard reaction by ethanol. This may also
explain the marked browning in ethanolic solutions at low aw, such
as a liqueur, during storage.
Figure 8 shows the relationship between aw and the extent of
browning in the 0.2 M glucose solution and the G-G solution. The
readings of absorbance at 420 nm of the samples containing 0.2 M
glucose alone are very low (Figure 8a). The contribution to browning
from caramelization should be negligible in the present study. Generally, severe heating and extreme pH are necessary for the caramelization of sugar in aqueous solutions (Morales and Van Boekel 1998).
The same thing could also be true in ethanolic solutions.
Both ethanol and glycerol accelerate the Maillard browning in GG solution through the formation of more HMF (Figure 7b and 8b).
Hofmann (1998) heated D-xylose, alanine, and furane-2-carboxaldehyde mixture in aqueous and methanolic solutions and reported
the formation of 3 additional color-active compounds in the presence of methanol. He concluded that methanol shifted the pathway for Maillard browning. Similar mechanism may present in ethanolic solutions as well.
Conclusions
T
he present study is concerned with the nonenzymatic browning in ethanolic solution. Maillard browning and HMF in etha-
Figure
7—The
relationship
between
aw
and
hydroxymethylfurfural (HMF) content in pH 4.3, 0.05 M
succinic acid–sodium hydroxide buffered ethanolic and
glycerolic solutions of (a) glucose (0.2 M) alone and (b) glucose (0.2 M)/glycine (0.2 M) after heating in boiling water
for 6 h. Different letters (a–j) within the same graph indicate significant difference (P < 0.05) in HMF content in
Duncan’s multiple range test.
FCT278
JOURNAL OF FOOD SCIENCE—Vol. 69, Nr. 4, 2004
nolic solutions of the G-G system are found to increase with an increase in ethanol concentration between 0% to 50%, suggesting the
enhancement of ethanol on the browning reaction. The results also
indicate that the mechanisms of the Maillard reaction in ethanolic solution and aqueous solution are not the same. Interestingly,
HMF content decreases with the increasing ethanol level in a system containing glucose alone. Experiments using glycerol as a control concluded that the reduction of aw is not the sole major mechanism for ethanol to accelerate HMF formation and Maillard
browning in the G-G solution.
The mechanism of the Maillard reaction in ethanolic solutions is
not known in detail. Further studies including the purification and
identification of the browning products are being done in our laboratory for a better understanding of this subject.
References
Akhavan I, Wrolstad RE. 1980. Variation of sugars and acids during ripening of
pears and in the production and storage of pear concentrate. J Food Sci 45:499–
501.
Ajandouz EH, Puigserver A. 1999. Nonenzymatic browning reaction of essential
amino acids: Effect of pH on caramelization and Malliard reaction kinetics. J
Agric Food Chem 47:1786-93.
Ames JM. 1990. Control of the Maillard reaction in food systems. Trends Food Sci
Technol 1:150–4.
Ames JM. 1998. Applications of the Maillard reaction in the food industry. Food
Chem 62:431–9.
Ashoor SH, Zent JB. 1984. Maillard browning of common amino acids and sugars. J Food Sci 49:1206–7.
Baxter JH. 1995. Free amino acid stability in reducing sugar systems. J Food Sci
Figure 8—The relationship between aw and absorbance at
420 nm in pH 4.3, 0.05 M succinic acid–sodium hydroxide
buffered ethanolic and glycerolic solutions of (a) glucose
(0.2 M) alone and (b) glucose (0.2 M)/glycine (0.2 M) after
heating in boiling water for 6 h. Different letters (a–j) within
the same graph indicate significant difference (P < 0.05)
in absorbance at 420 nm in Duncan’s multiple range test.
URLs and E-mail addresses are active links at www.ift.org
60:405–8.
Beck J, Ledl F, Sengl M, Severin T. 1990. Formation of acids, lactones and esters
through the Maillard reaction. Z Lebensm U Forsch 190:212–6.
BeMiller JN, Whistler RL. 1996. Carbohydrates. In: Fennema OR, editor. Food
chemistry. 3rd ed. New York: Marcel Dekker. p 157–223.
Brands CMJ, Alink GM, van Boekel MAJS, Jongen WMF. 2000. Mutagenicity of
heated sugar-casein systems: effect of the Maillard reaction. J Agric Food Chem
48:2271–5.
Cuzzoni MT, Stoppini G, Gazzani G, Mazza P. 1988. Influence of water activity and
reaction temperature of ribose-lysine and glucose-lysine Maillard systems
on mutagenicity, absorbance and content of furfurals. Food Chem Toxic 26:815–
22.
Davies CGA, Kaanane A, Labuza TP, Moscowitz AJ, Guillaume F. 1998. Evaluation
of the acyclic state and the effect of solvent type on mutarotion kinetics and on
Maillard browning rate of glucose and fructose. In: O’Brien J, Nursten H, Crabbe
MJ, Ames J, editors. The Maillard reaction in foods and medicine. London:
Royal Society of Chemistry. p 166–71.
Del Pilar Buera M, Chirife J, Resnik SL, Wetzler G. 1987. Nonenzymatic browning
in liquid model systems of high water activity: kinetics of color changes due to
Maillard’s reaction between different single sugars and glycine and comparison with caramelization browning. J Food Sci 52:1063–7.
Eichner K, Karel M. 1972. The influence of water content and water activity on
the sugar-amino browning reaction in model systems under various conditions. J Agric Food Chem 20:218–23.
Ferro Fontan C, Chirife J. 1981. The evaluation of water activity in aqueous solutions from freezing point depression. J Food Technol 16:21–30.
Gutiérrez IH. 2003. Influence of ethanol content on the extent of copigmentation in a Cencibel young red wine. J Agric Food Chem 51:4079–83.
Hill VM, Ledward DA, Ames JM. 1996. Influence of high hydrostatic pressure and
pH on the rate of Maillard browning in a glucose-lysine system. J Agric Food
Chem 44:594–8.
Hofmann T. 1998. Studies on the influence of the solvent on the contribution of
single Maillard reaction products to the total color on browned pentose/ala-
URLs and E-mail addresses are active links at www.ift.org
nine solutions—a quantitative correlation using the color activity concept. J
Agric Food Chem 46:3912–7.
Lee HS, Nagy S. 1988. Quality changes and nonenzymic browning intermediates in grapefruit juice during storage. J Food Sci 53:168–72.
Leong LP, Wedzicha BL. 2000. A critical appraisal of the kinetic model for the
Maillard browning of glucose with glycine. Food Chem 68:21–8.
Miracco JL, Alzamora M, Chirife J, Ferro Fontan C. 1981. On the water activity of
lactose solutions. J Food Sci 46:1612–3.
Morales FJ, Jimenez-Perez S. 2001. Free radical scavenging capacity of Maillard
reaction products as related to color and fluorescence. Food Chem 72:119–25.
Morales FJ, Van Boekel MAJS. 1998. A study on advanced Maillard reaction in
heated casein/sugar solutions: color formation. Int Dairy J 8:907–15.
Petriella C, Resnik SL, Lozano RD, Chirife J. 1985. Kinetics of deteriorative reactions in model food systems of high water activity: color changes due to nonenzymatic browning. J Food Sci 50:622–6.
Renn PT, Sathe SK. 1997. Effects of pH, temperature, and reactant molar ratio on
L -leucine and D-glucose Maillard browning reaction in an aqueous system. J
Agric Food Chem 45:3782–7.
Toribio JL, Lozano JE. 1984. Nonenzymatic browning in apple juice concentrate
during storage. J Food Sci 49:889–92.
Tress R, Nittka C, Kersten E, Rewicki D. 1995. Formation of isoleucine-specific
Maillard products from[1-13C]- D-glucose and [1-13C]- D-fructose. J Agric Food
Chem 43:1163–9.
Trifiro A, Gherardi S, Belloli S, Saccani G, Aldini R. 1990. Effetti della tecnologia
di lavorazione e delle condizioni di magazzinaggio sulle reazioni
d’imbrunimento non enzimatico in derivati del pomodoro. Industr Conserv
65:210–5.
Van Martins SIFS, Jongen WMF, Boejel MAJS. 2001. A review of Maillard reaction
in food and implications to kinetic modeling. Trends Food Sci Technol 11:364–
73.
Wedzicha BL, Kaputo MT. 1992. Melanoidins from glucose and glycine: composition, characteristics and reactivity toward sulfite ion. Food Chem 43:359–67.
Vol. 69, Nr. 4, 2004—JOURNAL OF FOOD SCIENCE
FCT279
Food Chemistry and Toxicology
Maillard browning in ethanolic solution . . .