Yilmaz 2005
Yilmaz 2005
Yilmaz 2005
Chemistry
Food Chemistry 93 (2005) 273–278
www.elsevier.com/locate/foodchem
Received 10 May 2004; received in revised form 22 September 2004; accepted 22 September 2004
Abstract
Maillard reaction products (MRP) results from a condensation reaction between amino acids (or proteins) and reducing sugars
or lipid oxidation products, and MRP exhibit in vitro antioxidant activities. The objective of this study was to determine antioxidant
activities of water-soluble MRP from the reaction between histidine (His) and glucose (Glu) by using the oxygen radical absorbing
capacity (ORACPE) assay with phycoerythrine. Heating His–Glu mixture at 100 C up to 30 min did not generate MRP with anti-
oxidant activity. However, significant formation of MRP with ORACPE values of 0.25, 0.43, and 0.44 lmol Trolox equivalent/mg
reaction mixture was observed when the mixture was heated at 120 C for 10, 20, and 30 min, respectively. Heating the mixture at
120 C over 30 min reduced the peroxyl radical scavenging activity of the MRP, possibly due to the degradation of antioxidant MRP
formed in the earlier stages of the reaction. In conclusion, MRP obtained from His–Glu mixture possesses peroxyl radical scaveng-
ing activity, and this activity can be quantified by the ORACPE assay.
2004 Published by Elsevier Ltd.
Keywords: Maillard reaction; ORAC; Antioxidant activity; Peroxyl radical scavenging activity; Glucose; Histidine
of reactants (Wijewickreme, Kitts, & Durance, 1997), & Prior, 1999). Antioxidant activities of foods are calcu-
temperature, pH (Monti, Bailey, & Ames, 1998), water lated in terms of Trolox equivalents. This indication pro-
activity, intermediate products (Vasiliauskaite & Wedzi- vides the means of comparison among the antioxidant
cha, 1997) and availability of oxygen can strongly affect activities of numerous foods.
the formation and properties of the final reaction prod- The objective of this study was to determine the anti-
ucts, model systems have been studied more often than oxidant activity of water-soluble MRP from glucose–
the actual food. histidine mixture by means of ORACPE assay.
MRP, especially melanoidins, have been reported to
have antioxidant activity through scavenging oxygen
radicals or chelating metals (Table 1). MRP from his- 2. Materials and methods
tidine had the highest antioxidant activity determined
by conjugated diene formation from peroxidation of 2.1. Production of MRP
linoleic acid among MRP from either dipeptides of
histidine–phenylalanine or lysine–alanine, amino acids L -Histidine hydrochloride monohydrate and D -(+)-
histidine, lysine, or ascorbic acid when glucose was glucose were purchased from Sigma Chemical Co. (St.
used as a reducing sugar (Reische, 1994). Compounds Louis, MO). Histidine and glucose were mixed (1:2 ra-
in the MRP with amino reductone structures may tio), and approximately 30% (w/v) deionized water was
have both antioxidant and pro-oxidant activities (Pis- added to the mixture in screw-capped tubes. The screw
chetsrieder, Rinaldi, Gross, & Severin, 1998) depend- cap was kept loose to expose the reaction mixture to
ing on the reaction conditions. MRP obtained from air while heating in a vegetable oil bath at either
heated histidine and glucose exhibit copper ion bind- 100 C for 10, 20, 30, and 60 min or 120 C for 5, 10,
ing ability in oil/water mixtures (Bersuder, Hole, & 20, 30 and 60 min simulating conditions on baking.
Smith, 2001). Then the screw cap was tightened, and tubes were kept
The oxygen radical absorbance capacity assay can be at 18 C until extraction. pH was not controlled during
used to quantify the antioxidant capacity of foods by the production of MRP. Triplicates of the experiment
measuring peroxyl radical scavenging activity of the were run.
compounds present (Cao & Prior, 1999). This assay is
based on the chemical damage to b-PE caused by a per- 2.2. Extraction of antioxidant MRP
oxyl radical producing compound (i.e. 2,2 0 -azobis(2-ami-
dinopropane) dihydrochloride (AAPH) in this assay), Deionized water (40 ml/g mixture) was added to the
reducing the fluorescence emission of b-PE. The presence mixture and vortexed until MRP dissolved. The mixture
of antioxidants in the medium prevents the damage and was centrifuged at 26,000g for 30 min at 4 C. Superna-
prolongs the fluorescence emission. Antioxidant capacity tants were collected and subjected to ORACPE assay. At
of foods can be quantified by measuring the area under least two ORACPE assays were done on each replicate of
the relative fluorescence intensity vs. time curves (Cao the treatments.
Table 1
Antioxidative properties of Maillard reaction products obtained from model systems reported in the literature
Model system Mode of antioxidative property Reference
Sugar–amino acid
Glu–His Copper chelator Bersuder et al. (2001)
Oxygen radical scavenger Lingnert et al. (1983)
Peroxyl radical scavenger Lingnert and Eriksson (1980a,b)
Glu–Lys Copper chelator Wijewickreme et al. (1997), Dittrich et al. (2003), Wijewickreme and Kitts (1998)
DPPH radical scavenger Morales and Jimenez-Perez (2001)
Peroxyl radical scavenger Bressa et al. (1996)
Hydroxyl radical scavenger Wijewickreme et al. (1999)
Glu–Gly Copper chelator Dittrich et al. (2003)
Peroxyl radical scavenger Wagner et al. (2002)
Hydroxyl radical scavenger Yoshimura et al. (1997)
Fe2+ chelator Yoshimura et al. (1997)
Fru–Lys Copper chelator Wijewickreme et al. (1997),Wijewickreme and Kitts (1998)
Hydroxyl radical scavenger Wijewickreme et al. (1999)
Lac–Lys Peroxyl radical scavenger Monti et al. (1999)
Y. Yilmaz, R. Toledo / Food Chemistry 93 (2005) 273–278 275
at 420nm 12 at 420nm
40
(a) (b)
Corrected Absorbance
Corrected Absorbances
35 10
30
8
25
20 6
15
4
10
5 2
0 0
0 20 40 60 80 0 20 40 60
Time (min) Time (min)
Fig. 1. Corrected absorbances (absorbance · dilution factor) and standard deviations of melanoidin formation in MRP solutions at 420 nm, which
were obtained from a His–Glu model system heated for up to 1 h at 120 C (a) and at 100 C (b). Bars represent standard deviations.
Blank
1.00 MRP at 100˚C for 30min (diluted)
MRP at 120˚C for 30min (diluted)
Trolox (1umol)
0.80
Relative Fluorescence Intensity
0.60
0.40
0.20
0.00
0 10 20 30 40
Time (min)
Fig. 2. Relative fluorescence intensity reduction of b-PE in phosphate buffer and in the presence of MPR and Trolox.
time nor temperature significantly affected ORACPE val- histidine. At 120 C, ORACPE values of MRP increased
ues of His (P > 0.05). Glucose was also subjected to the with heating time up to 30 min, then dropped after 1 h
ORACPE assay, but it did not have antioxidant activity of heating. At 120 C 30 min of heating approximately
in ORACPE assay. doubled the ORACPE value of the MRP at 10 min.
ORACPE values of MRP are shown in Fig. 3. OR- Neither glucose nor histidine by themselves showed
ACPE values of MRP solutions per gram of mixtures browning upon heating at either 100 or 120 C for up
did not significantly change when the mixtures were to 60 min. Therefore, the brown color (from light to
heated at 100 C for 10, 20 or 30 min (P > 0.05). dark brown) that appeared upon heating glucose and
Although using DPPH (2,2-diphenyl-1-picrylhydrazyl), histidine mixture was a result from the Maillard reac-
Murakami et al. (2002) revealed radical scavenging tion. A study on the effect of caramelization on the
activities of bright color pigments from histidine and xy- Maillard browning reaction revealed that heating glu-
lose heated at 30 C up to 120 h, heating histidine–glu- cose at 100 C at pH 6 or 8 showed little or no browning
cose mixture at 100 C for 10, 20 or 30 min was while the browning of glucose and lysine mixture in-
insufficient to generate MRP with peroxyl radical scav- creased at pH 8 (Ajandouz & Puigserver, 1999).
enging activities. Heating the reaction mixture at 100 Yoshimura, Iijima, Watanabe, and Nakazawa (1997)
C for 1 h, however, increased the ORACPE significantly reported that heating glucose and glycine at around 100
(P < 0.05) to about 0.3 mmol/g mixture. Statistical anal- C for 1–6 h produced compounds that are able to scav-
yses showed that peroxyl radical scavenging activity of enge hydroxyl radicals and superoxide anion (O2 ) and
the mixture prepared at 120 C for 5 min was similar to chelate with Fe2+. Our results showed that glucose
to that of the mixture heated at 100 C for either 10, and histidine also produce MRP with oxygen radical
20 or 30 min (P > 0.05). Furthermore, the ORACPE val- scavenging activity. When tested with ORACPE assay,
ues of MRP heated <1 h at 100 C was primarily from antioxidant activity of MRP from this model system
Y. Yilmaz, R. Toledo / Food Chemistry 93 (2005) 273–278 277
200
150
100
50
0
0 10 20 30 40 50 60 70
Time (min)
Fig. 3. Peroxyl radical scavenging activities of MRP per ml of solution in the graph and per g of initial mixture in the table below. Bars represent
standard deviations.
was >400 lmol TE/g mixture upon heating at 120 C for MRP formed during baking can show antioxidant
up to 30 min. activity that inhibits the lipid oxidation. Lingnert
Wagner, Derkits, Herr, Schuh, and Elmadfa (2002) (1980) successfully showed that addition of histidine
reported that MRP formed by heating glucose and gly- and glucose to cookie dough inhibited the oxidation of
cine at 120 C exhibit higher antioxidant activity in lipids in cookies. In our study, MRP from heated glu-
hydrophilic matrices than lipophilic matrices. The cose and histidine are able to scavenge peroxyl radicals
authors also found that addition of those MRP to coco- produced by AAPH. On the other hand, undesired com-
nut fat prior to frying at 200 C was ineffective for pounds like carcinogens or mutagens (Gazzani, Vagnar-
extending the shelf life. In our research, results indicated elli, Cuzzoni, & Mazza, 1987) can form at high
that water soluble MRP from a model system of glucose temperatures from mixtures of amino acid or peptides
and histidine also exhibit antioxidant activity, and fur- and sugars in overheated products, and could be a prob-
ther heating reduces the antioxidant activity of MRP. lem for human health. Moreover, acrylamide, a carcin-
Prior et al. (1998) reported that the estimated antiox- ogen and neurotoxin for humans, can form when
idant intake in terms of ORACPE ranges from 1.2 to 1.7 foods high in carbohydrate are fried, baked, or roasted
mmol TE/day in the US. Strawberry, plum, red grape at high temperatures (EC, 2002). Therefore, further re-
(whole), and orange has ORACPE values of 154, 80, search is needed in order to counteract the detrimental
36 and 52 lmol TE/g dry matter, respectively (Wang, effects of excessively heated MRP on health while taking
Cao, & Prior, 1996). Red raspberries have ORACPE val- advantage of their in vitro antioxidant activities.
ues ranged from 91 to 115 lmol TE/g dry matter (Wang
& Lin, 2000). ORACPE values of blueberries depend on
variety but the range was reported to be from 63 to 282 4. Conclusions
lmol TE/g dry matter (Prior et al., 1998). ORACPE val-
ues of MRP from histidine–glucose model system ran- In this study, we showed that MRP from heated his-
ged from 250 to 450 mmol/g His–Glu mixture; tidine and glucose have peroxyl radical scavenging
therefore, MRP formed during the thermal processing activity, an indicator of antioxidative activity in vitro.
of foods may have a beneficial effect on the total dietary ORACPE assay makes the quantification of antioxidant
antioxidant intake for humans. In our study, we demon- activity of MRP in food products possible. This advan-
strated that antioxidant activity of MRP could be quan- tage of ORACPE assay could be useful for future deter-
tified by a relatively new ORACPE assay compared with minations of the contribution of non-enzymatic
other methods. ORACPE assay can be successfully used browning reaction products with antioxidant activity
to determine the effect of dietary MRP on total antiox- to total antioxidant intake in the human diet. However,
idant intake in the human diet. precaution is needed since there is a possibility of the
278 Y. Yilmaz, R. Toledo / Food Chemistry 93 (2005) 273–278
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