Food Chemistry 164 (2014) 510–517

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Anthocyanin, phenolics and antioxidant activity changes in purple waxy

corn as affected by traditional cooking
Bhornchai Harakotr a,b, Bhalang Suriharn a,b, Ratchada Tangwongchai c, Marvin Paul Scott d,
Kamol Lertrat a,b,⇑
a
Department of Plant Science and Agricultural Resources, Khon Kaen University, Khon Kaen 40002, Thailand
b
Plant Breeding Research Center for Sustainable Agriculture, Khon Kaen University, Khon Kaen 40002, Thailand
c
Department of Food Technology, Khon Kaen University, Khon Kaen 40002, Thailand
d
USDA-ARS, Agronomy Hall, Iowa State University, Ames 50011, USA

a r t i c l e i n f o a b s t r a c t

Article history: Antioxidant components, including anthocyanins and phenolic compounds, antioxidant activity, and
Received 16 July 2013 their changes during traditional cooking of fresh purple waxy corn were investigated. As compared to
Received in revised form 10 April 2014 the raw corn, thermal treatment caused significant (p 6 0.05) decreases in each antioxidant compound
Accepted 14 May 2014
and antioxidant activity. Steam cooking preserved more antioxidant compounds than boiling. Boiling
Available online 21 May 2014
caused a significant loss of anthocyanin and phenolic compounds into the cooking water. This cooking
water is a valuable co-product because it is a good source of purple pigment. By comparing levels of anti-
Keywords:
oxidant compounds in raw and cooked corn, we determined that degradation results in greater loss than
Purple waxy maize
Bioactive compounds
leaching or diffusion into cooking water. Additionally, separation of kernels from the cob prior to cooking
Antioxidant capacity caused increased loss of antioxidant compounds.
Domestic cooking Ó 2014 Elsevier Ltd. All rights reserved.

1. Introduction and is harvested while immature and consumed on the cob as fresh
food similar to sweet corn. Normally, this type of corn is cooked by
Free radicals are known to be a major contributor to degenera- boiling or steaming. It is known that cooking induces changes in
tive diseases of aging (Atoui, Mansouri, Boskou, & Kefalas, 2005). physiological and chemical composition, influencing the concen-
Dietary antioxidants might confer health-protective benefits by tration and bioavailability of bioactive compounds in food
alleviating oxidative stress by preventing free radicals from damag- (Turkmen, Sari, & Velioglu, 2005). However, there are conflicting
ing proteins, DNA and lipids (Huang, Ou, & Prior, 2005). Corn is a results on the effects of conventional cooking methods on dietary
good source of natural antioxidants such as vitamins, carotenoids, antioxidant levels obtained by consumers. To understand better
flavonoids, and other phenolic compounds (Lopez-Martinez, the effects of cooking on antioxidant levels in food, it is necessary
Oliart-Ros, Valerio-Alfaro, Lee, & Parkin, 2009; Montilla, to test real cooking conditions, because the behavior of any food
Hillebrand, Antezana, & Winterhalter, 2011). Accumulated evidence cannot be predicted (Oliveira, Amaro, Pinho, & Ferreira, 2010).
suggests that anthocyanin pigments in corn are responsible for its The potential changes in anthocyanins, phenolic acids and antiox-
high antioxidant activities and have been shown to potentially idant activity of waxy corn during thermal treatment have not
reduce the risk of colon cancer (Hagiwara et al., 2001), prevent heart been investigated yet. The aim of this work was to evaluate the
ischemia–reperfusion injury and hyperlipidemia (Toufektsian et al., effects of different domestic cooking conditions namely boiling
2008), anti-inflammatory effects (He & Giusti, 2010) and prevent and steaming, on kernels on or off the cob on anthocyanins, phen-
diabetes and obesity (Tsuda, Horio, Uchida, Aoki, & Osawa, 2003). olics and antioxidant activity of purple waxy corn. Moreover, we
Waxy corn (Zea mays L. var. ceratina) is increasingly consumed determined these antioxidant compounds in residuals after
in China, Korea, Vietnam, Taiwan, Laos, Myanmar and Thailand, cooking such as cob and water, to understand the potential for
developing value-added co-products and utilizing of waste. This
⇑ Corresponding author at: Department of Plant Science and Agricultural information may have a significant impact on consumers’ selection
Resources, Khon Kaen University, Khon Kaen 40002, Thailand. Tel./fax: +66 432 of cooking methods and allow them to better preserve the
02696. nutritional value of their food.
E-mail address: [email protected] (K. Lertrat).

http://dx.doi.org/10.1016/j.foodchem.2014.05.069
0308-8146/Ó 2014 Elsevier Ltd. All rights reserved.
 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517 511

2. Materials and methods Wrolstad (2001) and Jing, Noriega, Schwartz, and Giusti (2007)
with slight modifications. Approximately 2 g of each sample were
2.1. Chemicals and reagents added to a flask containing 25 mL of 70% aqueous acetone acidified
by the addition of HCl to 0.01% and mixed well. The flasks were
Authentic anthocyanin standards: cyanidin-3-O-glucoside; shaken on a platform shaker (LabScientific Inc, USA) at 200 rpm
pelargonidin-3-O-glucoside; peonidin-3-O-glucoside, Folin–Ciocal- and room temperature for 2 h. Each sample was filtered through
teu’s phenol reagent, 2,4,6-tri(2-pyridyl)-S-triazine (TPTZ), ferulic, Whatman # 1 filter paper under vacuum using a Büchner funnel,
protocatechuic, p-coumaric, vanillic, caffeic, p-hydroxybenzoic, and the slurry was washed with 10 mL of acidified 70% acetone.
syringic, gallic, chlorogenic acids and Trolox (6-hydroxy-2,5,7, The filtrate was transferred to a separatory funnel, and 15 mL of
8-tetramethylchroman-2-carboxylic acid) were obtained from chloroform were added. The mixture was gently mixed by turning
Sigma–Aldrich (USA). 2,2’-azino-bis (3-ethylbenzothiazoline-6- the funnel upside down a few times. The samples were stored
sulphonic acid) diammonium salt (ABTS) was from Fluka overnight at 4 °C or until a clear partition between the two phases
(Switzerland). HPLC-grade methanol, acetonitrile and reagents was obtained. The solution was transferred to a centrifuge tube
were purchased from Labscan (Poland). All of the chemicals and and centrifuged at 11,538g and 4 °C for 10 min. The upper aque-
reagents used in the experiments were of analytical grade. ous layer containing the acetone/water mixture was collected, and
the chloroform/acetone layer was carefully discarded. The residual
2.2. Plant material and sample preparation acetone and chloroform were removed from the anthocyanin
extract using a rotary evaporator at 40 °C under vacuum. The vol-
For this study, the purple waxy corn variety ‘‘Khao Niew Dum’’, ume of the extracts was increased to 25 mL in a volumetric flask by
developed by the Plant Breeding Research Center for Sustainable the addition of 0.01% HCl-acidified methanol.
Agriculture, Khon Kaen University, Thailand was used. Corn was
grown during October to December 2011 and recommended prac- 2.4. Determination of monomeric anthocyanin content
tices for commercial production of corn were followed. Ears were
picked by hand at the milk stage (20 days after pollination; DAP). Monomeric anthocyanin content was measured by the pH dif-
For the analyses, only physiologically undamaged ears with the ferential method, as described by Giusti and Wrolstad (2001). A
mass 200–220 g were used. For raw corn, a length of 3 cm from UV–vis spectrophotometer (GENESYS 10S, Thermo Scientific,
the terminal tip end was removed from 10 waxy corn ears to USA) was used to measure the absorbance at 510 and 700 nm.
reduce the kernel maturity variation. Then, kernels were manually Anthocyanin levels were expressed as lg of cyanidin-3-glucoside
cut from cob and dipped into liquid nitrogen to stop enzymatic equivalents per g of dry weight (lg CGE/g DW), using the reported
activity, corn kernels and cobs were freeze-dried and finely ground molar extinction coefficient of 26,900 M 1 cm 1 and a molecular
with a sample mill, sieved through a 60-mesh screen, thoroughly weight of 449.2 g/mol.
mixed and stored at 20 °C until analysis.
Boiling and steaming were used because these are common 2.5. Quantification of specific anthocyanins
methods of cooking fresh corn. Cooking conditions were optimized
by preliminary experiments carried out for each treatment. For all Reversed-phase HPLC analysis of anthocyanins was performed
cooking treatments, the minimum cooking time to reach tender- using a Shimadzu system (Shimadzu, Japan) equipped with a bin-
ness for adequate palatability and taste was used. For cooking, ary pump (LC-20AC pump) and a diode array detector (SPD-M20A).
the following methods were used: Chromatographic separations were performed on an Xselect CHS
C-18 column (4.6  250 mm, i.d. 5 lm) (Waters, USA). The compo-
(a) 10 corn ears (2 kg) were boiled in 4L tap water in a covered sition of solvents and the gradient elution conditions used were
stainless-steel pot and cooked for 19 min in boiling water. those described by Kim et al. (2007), with slight modifications.
After cooking, boiled corn kernels were cut from the cob The mobile phases used were 0.1% hydrochloric acid in methanol
with a knife. (15:85 v/v) (phase A) and 8% formic acid (phase B), at a flow rate
(b) A single layer 10 corn ears (2 kg) were steamed in a cov- of 1 mL/min. Gradient elution was performed as follows:
ered stainless-steel steamer (45 cm diameter) for 26 min. 0–0.5 min, 0–80% phase B; 0.5–9.5 min, 80–10% phase B; 9.5–
The water in steamer was maintained boiling throughout 10 min, 10–15% phase B; 10–15 min, 15–5% phase B; 15–20 min,
the process to generate steam. After cooking, steamed corn 5–80% phase B; and a re-equilibration period of 1 min with 80%
kernels were separated from the cob by using a knife. phase B used between individual runs. Operating conditions were
(c) Fresh cut corn kernels from 20 corn ears (2 kg) were boiled as follows: column temperature 30 °C, injection volume 20 lL,
in 2L tap water in a covered stainless-steel pot and cooked and a detection wavelength of 250–600 nm (a representative
for 7 min to boiling water. wavelength of 520 nm). Solutions were injected after being filtered
(d) Fresh cut corn kernels from 20 corn ears (2 kg) were through a 0.20 lm nylon membrane filter. Anthocyanins in sam-
steamed in a covered stainless-steel steamer (45 cm diame- ples were identified by comparing their relative retention times
ter) for 12 min. The water in steamer was maintained boiling and UV spectra with those of standards and were detected using
throughout the process to generate steam. an external standard method. The results for the anthocyanins
were expressed as lg per g of dry weight (lg/g DW).
Each method was carried out in triplicate. After all cooking
experiments, samples were cooled rapidly on ice. Then, specimens 2.6. Extraction of phenolic compounds and antioxidant activity
were frozen and freeze–dried similarly to raw corn. Cooking water determination
was thoroughly mixed with a homogenizer prior to antioxidant
analyses. Free phenolic compounds in waxy corn kernels and cobs were
extracted according to the method described by Adom and Liu
2.3. Extraction of anthocyanin determination (2002), with slight modifications. Approximately 2 g of each sam-
ple were added to a flask containing 25 mL of 80% chilled ethanol.
The anthocyanins in ground waxy corn kernels were extracted The flask was shaken on a platform shaker (LabScientific Inc, USA)
according to the method described by Rodriguez-Saona and at 200 rpm at room temperature for 10 min. After centrifugation at
512 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517

11,538g and 4 °C for 10 min, the supernatant was removed; correlation test was conducted to determine the correlation
extraction was repeated two more times. Supernatants were between variables.
pooled and then evaporated at 40 °C under vacuum. The residue
from vacuum evaporation was re-dissolved in 25 mL of 80%
ethanol. Extracts were stored at 20 °C until analysis. 3. Results and discussion

2.7. Determination of phenolic contents 3.1. Effects of cooking conditions on monomeric anthocyanin and
phenolic contents
The phenolic contents were determined using the Folin–
Ciocalteu (F–C) method as described by Hu and Xu (2011). Briefly, Concentration of anthocyanin may vary among foods produced
the appropriate dilutions of the extracts were oxidized with F–C by a given plant species due to different external and internal fac-
reagent for 90 min, and the reaction was neutralized with sodium tors, such as genetic and agronomic factors, intensity and type of
carbonate. The absorbance of the resulting blue color was mea- light, temperature, processing and storage (de Pascual-Teresa &
sured at 765 nm, and the phenolic content was expressed as mg Sanchez-Ballesta, 2008). Raw corncob exhibited a monomeric
of gallic acid equivalents per g of dry weight (mg GAE/g DW). anthocyanin content (2534.1 lg CGE/g DW) higher than that of
raw corn kernels (754.0 lg CGE/g DW) (Table 1). However, the
2.8. Quantification of phenolic compounds anthocyanin content in waxy corn kernels was found to be sub-
stantially higher than those reported for pigmented corn from Boli-
Reversed-phase HPLC analysis of anthocyanins was performed via (19–717 lg CGE/g DW) (Montilla et al., 2011), Mexico (721 lg
using a Shimadzu system (Shimadzu, Japan) equipped with a bin- CGE/g DW), and the United States (307 lg CGE/g DW) (del
ary pump (LC-20AC pump) and a diode array detector (SPD-M20A). Pozo-Insfran, Brences, Sena, Saldivar, & Talcott, 2006). The cooking
Chromatographic separations were performed on an Xselect CHS process had a significant (p 6 0.05) impact on the retention of
C-18 column (4.6  250 mm, i.d. 5 lm) (Waters, USA). Butsat, monomeric anthocyanin content. Cooking cut-kernels by boiling
Weerapreeyakul, and Siriamornpun (2009) described the composi- resulted in the greatest decreases (60.7%), followed by boiled
tion of solvents and the gradient elution conditions used with whole-ears (31.7%), steamed cut-kernels (19.2%), and steamed
slight modifications. The mobile phase consisted of deionized whole-ears (3.5%). The preservation of natural pigments after ther-
water with acetic acid (pH 2.74) (phase A) and acetonitrile (phase mal processing is a major quality parameter. In a study of the effect
B) at a flow rate of 0.8 mL/min. Gradient elution was performed as of a cooking on anthocyanins in sweet potato, steaming reduced
follows: 0–5 min, 5–9% phase B; 5–15 min, 9% phase B; 15–22 min, anthocyanin content by nearly half of original amount (Kim
9–11% phase B; 22–38 min, 11–18% phase B; 38–43 min, 18–23% et al., 2012). Therefore, the various results indicated the impor-
phase B; 43–44 min, 23–90% phase B; 44–45 min, 90–80% phase tance of cooking method on nutrient retention.
B; 45–55 min, isocratic at 80% phase B; 55–60 min, 80–5% phase The lost anthocyanin content in corn could be due to degrada-
B; and a re-equilibration period of 5 min with 5% phase B used tion or decomposition of anthocyanin upon thermal treatments
between individual runs. Operating conditions were as follows: (Ioannou, Hafsa, Hamdi, Charbonnel, & Ghoul, 2012). Beginning
column temperature 38 °C, injection volume 20 lL, and UV-diode with the monomeric anthocyanin content in raw corn, it was pos-
array detection at 280 nm (hydroxybenzoic acids) and 325 nm sible calculated the percentage of loss to the cooking water and to
(hydroxycinnamic acids). Spectra were recorded from 200 to degradation in the different cooking conditions. In the case of the
600 nm. Phenolic compounds in samples were identified by com- whole-ear corns, boiling resulted in a higher percentage of antho-
paring their relative retention times and UV spectra with those cyanin degradation than steaming by around 4 times. Boiling
of standards and were detected using an external standard resulted in a higher percentage of anthocyanin being degraded
method. The results for the phenolics were expressed as lg per g than was retained in the kernels. The percentage of anthocyanin
of dry weight (lg/g DW). in cooking water was not significant (p 6 0.05). These suggest the
decrease of anthocyanin during boiling was predominantly caused
2.9. Determination of antioxidant activities by breakdown of anthocyanins rather than their release to the
cooking water. Cut-kernels showed similar result to whole-ear
The reducing ability was determined using the ferric reducing cooking, however, boiling resulted in a greater percentage loss of
antioxidant power (FRAP) assay, which was performed according anthocyanin to cooking water than steaming up to 900-folds
to the method described by Hu and Xu (2011). The FRAP values (Fig. 1). The stability of anthocyanin and other food pigments
are expressed as lmol of Fe (II) per g of dry weight (lmol Fe (II)/ decreased with increasing temperature (Xu & Chang, 2008). Jing
g DW). The linear range of the calibration curve was 10–100 lM. and Giusti (2007) observed a consistent decrease of protein at
The Trolox equivalent antioxidant capacity (TEAC) assay, which 100 °C in purple corn water extracts indicating a possible protein
measures the reduction of radical cations of ABTS by antioxidants, denaturation at high temperatures, which could result in anthocy-
was conducted as described by Lopez-Martinez, Oliart-Ros, anin complexation and precipitation leading to a decline in
Valerio-Alfaro, Lee, and Parkin (2009). Trolox was used as the ref- anthocyanin content.
erence compound. The results are expressed in lmol of Trolox Data on phenolics in the literature about cooked waxy corn are
equivalents per g of dry weight (lmol TE/g DW). The linear range very limited. Total phenolic content of corn revealed a trend
of the calibration curve was 100–1000 lM. similar to that described for anthocyanin contents. Significant dif-
ferences (p 6 0.05) in phenolic content were found among treat-
2.10. Statistical analysis ments (Table 1). In the present study, boiled cut-kernels resulted
in the greatest decreases (47.8%), followed by the boiled whole-
All data were reported as means ± standard deviation. Data ears (30.4%), steamed cut-kernels (27.5%), and steamed whole-ears
were subjected to two-way analysis of variance (ANOVA), Duncan’s (10.1%). These results are in good agreement with those reported
multiple range test (DMRT) (p 6 0.05) was used to identify signif- by Turkmen et al. (2005), who found that thermal treatments
icant differences between group means and the calculated effects decreased the total phenolics in squash, peas and leek. As com-
for the 22 factorial design were obtained by using JMP Pro software pared to the boiling treatments, steaming treatments retained
(version 10.0, SAS institute Inc., Chicago, IL, USA). The Pearson greater phenolic contents. These significant losses could be
 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517 513

Table 1
Anthocyanins, phenolic compounds and antioxidant activities in raw and cooked purple waxy corn kernel under different processing and cooking conditionsa.

Treatments/Compounds Raw kernel Whole-ear corn Cut-kernel

b b
Boiling Steaming Boilingb Steamingb
Anthocyanins
MAC (lg CGE/g) 754.0 ± 62.8 514.7 ± 6.3c 727.9 ± 32.2a 298.1 ± 36.0d 609.5 ± 11.3b
Cy-3-Glu (lg/g) 121.6 ± 6.8 61.8 ± 1.9c 82.9 ± 3.1a 40.3 + 1.3d 72.0 ± 2.5b
Pg-3-Glu (lg/g) 46.7 ± 1.8 35.7 ± 1.3b 40.7 ± 1.9a 15.2 + 1.3c 37.6 ± 0.9ab
Pn-3-Glu (lg/g) 39.0 ± 1.8 23.4 ± 0.5c 28.9 ± 0.8a 11.2 + 0.1d 24.3 ± 0.3b
Phenolic compounds
TPC (mg GAE/g) 6.9 ± 0.4 4.8 ± 0.2b 6.2 ± 0.2a 3.6 ± 0.1c 5.0 ± 0.2b
PCCA (lg/g) 9.9 ± 0.3 7.0 ± 0.2c 14.1 ± 0.3a 4.8 ± 0.2d 12.0 ± 0.1b
p-OH (lg/g) 1.8 ± 0.1 0.4 ± 0c 1.5 ± 0a 0.3 ± 0.1d 1.1 ± 0.1b
VA (lg/g) 9.8 ± 0.9 6.2 ± 0.4b 8.6 ± 0.5a 2.3 ± 0.5c 6.8 ± 0.3b
CFA (lg/g) 2.9 ± 0.1 1.5 ± 0.1b 2.3 ± 0.2a 1.1 ± 0c 1.4 ± 0.1bc
p-CA (lg/g) 11.0 ± 1.4 7.5 ± 0.3b 9.4 ± 0.2a 5.8 ± 0.6c 8.0 ± 0.4b
FA (lg/g) 23.1 ± 0.6 17.8 ± 0.8b 20.4 ± 0.7a 9.0 ± 0.9d 15.1 ± 0.6c
Antioxidant activities
FRAP (lmol Fe(II)/g) 102.3 ± 6.5 72.2 ± 1.1b 81.8 ± 0.8a 43.6 ± 1.9d 54.9 ± 3.4c
TEAC (lmol TE/g) 94.6 ± 2.5 67.4 ± 5.5b 90.6 ± 2.5a 45.6 ± 3.7c 62.7 ± 3.8b
a
Values are means ± SD. MAC, monomeric anthocyanin content; Cy-3-Glu, cyanidin-3-glucoside; Pg-3-Glu, pelargonidin-3-glucoside; Pn-3-Glu; peonidin-3-glucoside, TPC,
total phenolic content; PCCA, protocatechuic acid; p-OH, p-hydroxybenzoic acid; VA, vanillic acid; CFA, caffeic acid; p-CA, p-coumaric acid; FA, ferulic acid; FRAP, ferric
reducing antioxidant power; TEAC, Trolox equivalent antioxidant capacity.
b
Means in the same lines with different letters are significant (p 6 0.05) determined by Duncan’s multiple range test (DMRT).

Kernels
Cob

Cooking water

Degradation

(A) (B)

Fig. 1. Percentage of total monomeric anthocyanin content in cooked, cooking water and degradation of whole-ear corns (A) and cut-kernels (B) cooking as effect of different
cooking conditions. Values are expressed as the mean ± SD based on the initial contents in raw corn in dried weight.

attributed to water soluble phenolics leaching into cooking water properties (Xu & Chang, 2008). Cooking was found to give rise to an
as well as breakdown of phenolics during thermal processing. Sim- increase in the level of phenolic compounds in sweet corn
ilar to whole-ear anthocyanins, boiling showed a higher percent- (Dewanto, Wu, & Liu, 2002). Various effects can be attributed easily
age of phenolic degradation than steaming around 3 times, and to the diversity of food products used and lack of the standardiza-
the percentage of phenolics in cooking water was not significant tion of domestic cooking methods (Irina & Mohamed, 2012).
(p 6 0.05). Additionally, the percentage of phenolics retained in
kernels after boiling was less than the percentage lost due to deg- 3.2. Effects of cooking conditions on anthocyanin compositions
radation. In the case of cut-kernels, steaming resulted in greater
retention of phenolics in the kernels (73.0%) than boiling (53.0%). The concentration of cyanidin-3-glucoside decreased signifi-
These results indicated that phenolics were reduced by boiling cantly across all cooking methods. Boiled cut-kernels resulted in
by nearly half of original content. Phenolic content degradation the greatest decreases (66.9%), followed by the boiled whole-ears
showed a trend similar to that described for whole-ear cooked (49.2%), steamed cut-kernels (40.8%), and steamed whole-ears
corn. However, boiling caused phenolics to leach into cooking (31.8%) (Table 1). Pelargonidin-3-glucoside and peonidin-3-gluco-
water nearly 7 folds more than steaming (Fig. 2). These results sup- side were also decreased in the same manner; however, total
port the hypothesis that separation of kernels from the cob caused losses were lower than those of cyanidin-3-glucoside (except
loss of phenolics. Thermal treatment does not always result as the peonidin-3-glucoside in boiled cut-kernels). In contrast, peonidin
destruction of bioactive compounds. In some cases, cooking can 3-glucoside was lost to the greatest degree among all anthocyanins
induce the formation of novel compounds and improve antioxidant in black soybean during thermal processing (Xu & Chang, 2008).
514 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517

Kernels

Cob

Cooking water

Degradation

(A) (B)

Fig. 2. Percentage of total phenolic content in cooked, cooking water and degradation of whole-ear corns (A) and cut-kernels (B) cooking as effect of different cooking
conditions. Values are expressed as the mean ± SD based on the initial contents in raw corn in dried weight.

This study observed that pelargonidin-3-glucoside had the greatest and peonidin-3-glucoside degradation than those of boiling. How-
thermal stability of all anthocyanins, while, Sadilova, Stintzing, and ever, cyanidin-3-glucoside degradation was not significant different
Carle (2006) suggested that additional hydroxylation apparently between steaming and boiling (p 6 0.05), but total of anthocyanin
did not affect pigment stability when pelargonidin-3-glucoside degradation together with cooking water was 2 times less than that
was compared to cyanidin-3-glucoside. of boiling. Either anthocyanin was not detected by HPLC in steaming
It is well known that anthocyanins are readily degraded when water from whole-ears or cut-kernels, mostly the leaching effect on
exposed to heat, resulting in dramatic impact on color and their these compounds might not occur. Additionally, after cooking
health-promoting properties. Degradation starts with opening of whole-ears retained anthocyanin content better than cut-kernels.
the central ring followed by hydrolysis of the molecule, establishing This is possibly because when kernels were removed from the
colorless products. In the case of whole-ear corns, steaming retained cob, the pericarp was ruptured, which may reduce the barrier to
individual anthocyanin concentration better than boiling. As migration into the cooking water as well as increase the surface area
regards content and percentage of anthocyanin degradation simi- exposed to the cooking medium. Processing such as peeling,
larly to retain of their content in kernels; however, most of anthocy- trimming, chopping, slicing, crushing, pressing, and sieving was
anin content was preserved in the cob (Table 2). Additionally, expected to affect content, activity, and availability of antioxidant
pelargonidin-3-glucoside does not show degradation, in fact the composition (Ioannou et al., 2012). The resulting pigmented boiling
concentration of pelargonidin-3-glucoside in the cob slightly water is a potentially valuable co-product and can be simmered and
increased after cooking with either cooking method. This could be mixed with pineapple, sugar, juices and spices for cooked beverages
because the cob absorbs this compound from the cooking water. such as ‘‘Chicha Morada’’ (Aoki, Kuze, & Yashiaki, 2002). Moreover,
Bunea et al. (2008) suggested that the increase in concentrations pigmented boiling water may be used to cook rice or soak glutinous
of certain bioactive compounds after thermal treatment may be rice before cooking to make purple rice, and therefore, may be a suit-
explained either by their better release from the food matrix as a able replacement for others, more expensive source of purple color
result of the breakdown of supramolecular structures containing such as black rice or Asian pigeon wing (Clitorea ternatea), a
functional groups or their thermal stability. In the case of the cut- traditional source of purple/blue colorant in Asia.
kernels, similar to whole ears, it was found that steam cooking
resulted in retention of more anthocyanins than boiling. Addition-
3.3. Effects of cooking conditions on phenolic compounds
ally, boiling resulted in a greater percentage of anthocyanin degra-
dation than the percentage retained in the kernels (Table 3). Steam
This work focused on free phenolic acids because it has
cooking showed a lower percentage of pelargonidin-3-glucoside
been shown that the bioavailability of soluble phenolic acids is

Table 2
Distribution in percentage of anthocyanins and phenolic compounds in cooked, cooking water and degradation of whole-ear cooking in two cooking conditionsa.

Compounds Boiling Steaming

Kernel Cob Water Degradation Kernel Cob Water Degradation
Cy-3-Glu 12.0 ± 0.5e 61.6 ± 0.9b 5.0 ± 0.4f 21.4 ± 1.2c 16.2 ± 1.0d 79.2 ± 2.9a – 4.6 ± 2.1f
Pg-3-Glu 20.8 ± 0.3d 77.7 ± 2.2b 5.3 ± 0.4e – 23.8 ± 1.9c 86.5 ± 2.6a –
Pn-3-Glu 10.5 ± 0.7f 54.0 ± 1.0b 2.9 ± 0.8g 32.6 ± 2.2g 12.9 ± 0.3e 71.3 ± 0.7a – 15.8 ± 0.8d
PCCA 28.5 ± 1.8d 31.2 ± 5.7cd 3.2 ± 0.4e 37.1 ± 3.9bc 54.1 ± 1.5a 42.6 ± 4.4b 0.7 ± 0.1e 3.4 ± 2.7e
p-OH 9.7 ± 1.3e 28.5 ± 1.0c 24.7 ± 2.3d 37.1 ± 2.4a 34.0 ± 0.7b 37.4 ± 1.2a 2.3 ± 0.3f 26.3 ± 0.8cd
VA 22.3 ± 1.6d 38.9 ± 1.6b 5.0 ± 1.8ef 33.7 ± 1.4c 31.2 ± 2.1c 60.5 ± 3.1a 0.7 ± 0.1f 7.6 ± 4.7e
CFA 18.6 ± 0.6d 41.5 ± 3.7b 9.4 ± 0.7e 30.5 ± 2.8c 28.1 ± 4.2c 51.0 ± 6.5a 1.3 ± 0.1f 19.7 ± 4.3d
p-CA 23.0 ± 0.6e 39.5 ± 1.3b 1.4 ± 0.2g 36.2 ± 0.6c 28.7 ± 1.3d 53.2 ± 1.9a 0.5 ± 0.1g 17.6 ± 2.9f
FA 29.6 ± 0.3d 47.0 ± 0.7b 11.2 ± 1.1e 12.2 ± 1.6e 33.9 ± 1.4c 59.4 ± 2.4a 1.2 ± 0.1g 5.4 ± 2.1f
a
Values are means ± SD. Means in the same lines with different letters are significant (p 6 0.05) determined by Duncan’s multiple range test (DMRT). The abbreviations in
the column of compounds correspond to the same compounds represented in Table 1. Values were calculated based on the initial contents in raw corn in dried weight.
 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517 515

Table 3
Distribution in percentage of anthocyanins and phenolic compounds in cooked, cooking water, and degradation of cut-kernel cooking in two different cooking conditionsa.

Compounds Boiling Steaming

Kernel Water Degradation Kernel Water Degradation
Cy-3-Glu 33.2 ± 1.9bc 26.4 ± 2.6c 40.4 ± 3.8b 59.5 ± 6.0a – 40.5 ± 6.0b
Pg-3-Glu 32.8 ± 5.7bc 28.6 ± 4.3c 38.7 ± 4.1b 80.6 ± 3.9a – 19.4 ± 3.2d
Pn-3-Glu 28.6 ± 1.8d 27.3 ± 5.5d 44.1 ± 6.9b 63.8 ± 0.5a – 36.2 ± 0.5c
PCCA 48.3 ± 1.2b 10.5 ± 1.1d 41.2 ± 0.2c 60.0 ± 3.6a – –
p-OH 15.3 ± 4.0c 62.0 ± 2.5a 22.7 ± 1.9c 64.3 ± 5.8a – 34.6 ± 5.8b
VA 24.7 ± 4.4c 30.1 ± 5.9c 45.2 ± 9.6b 71.2 ± 0.9a 1.1 ± 0.3d 27.7 ± 1.2c
CFA 38.2 ± 3.0b 35.5 ± 5.5b 26.3 ± 3.6c 47.8 ± 0.4a – 52.0 ± 0.4a
p-CA 52.7 ± 5.7b 6.3 ± 0.2d 41.0 ± 5.5b 73.5 ± 9.3a – 26.4 ± 9.3c
FA 38.8 ± 4.2b 16.6 ± 2.1d 44.5 ± 6.2a 65.5 ± 5.3a 4.6 ± 1.0e 29.9 ± 4.4c
a
values are means ± SD. Means in the same lines with different letters are significant (p 6 0.05) determined by Duncan’s multiple range test (DMRT). The abbreviations in
the column of compounds correspond to the same compounds represented in Table 1. Values were calculated based on the initial contents in raw corn in dried weight.

considerably higher than that of the insoluble phenolic acids (Zhao, percentage of these substances undergoing degradation was
Egashira, & Sanada, 2004). To the best of our knowledge, phenolic higher than the percentage in cooking water in either whole-ear
acid profiles in processed waxy corn have not been systematically corns or cut-kernels, except p-hydroxybenzoic and caffeic acid
investigated. Three benzoic type phenolic acids (protocatechuic in boiled cut-kernels. Moreover, protocatechuic, p-hydroxybenzo-
acid, p-hydroxybenzoic acid and vanillic acid) and three cinnamic ic, caffeic and p-coumaric were not found in the water used to
type phenolic acids (caffeic acid, p-coumaric acid and ferulic acid) steamed cut-kernels (Tables 2 and 3). These results allow us to
were detected in both raw and cooked waxy corn. Gallic acid, conclude that degradation of phenolic acids was greater than
syringic acid and chlorogenic acid were not detected in either ker- losses due to leaching into cooking water. On the other hand,
nels or cob (data not shown). The predominant phenolic acids in the different results could be attributed to several factors includ-
both raw and cooked purple waxy corn were ferulic, followed by ing the type of material used, cultivars, cooking medium and ves-
p-coumaric and protocatechuic acids. In comparison to original sels, cooking time, pressure, and energy (Ioannou, Hafsa, Hamdi,
raw corn, both thermal treatments caused significant (p 6 0.05) Charbonel, & Ghoul, 2012). Some authors have indicated that
decreases in all measured phenolic compounds (except proto- pressure-cooking enhanced the antioxidant compositions and pal-
catechuic acid cooked by steaming) (Table 1). The thermal treat- atability of vegetables (Xu & Chang, 2009). However, higher
ment resulted in a 16.7–83.3% loss of p-hydroxybenzoic acid, power also could result in greater degradation (Hiemori et al.,
12.2–76.5% of vanillic acid, 20.7–62.1% caffeic acid, 14.5–47.2% of 2009). Further studies on pressure-cooking of antioxidant
p-coumaric acid, and 11.7–61.0% of ferulic acid compared to un- compositions in corn of this type need to be conducted.
cooked corn. The steamed whole-ear retained the greatest propor-
tion of phenolic acids, followed by steamed cut-kernels, boiled
whole-ears and boiled cut-kernels, respectively. However, steamed 3.4. Effects of cooking conditions on antioxidant activity
cut-kernels and boiled whole-ears were not different in vanillic,
caffeic and p-coumaric acid but were lower in ferulic acid. Boiling Thermal treatments are generally regarded as being destructive
had a more detrimental effect on phenolic acids than steaming. The to antioxidants. Antioxidant activities of raw and cooked corn,
boiling treatment causes of disruption cellular components with including FRAP and TEAC assays are presented in Table 1. Signifi-
the consequent release of these molecules into the cooking water cant differences (p 6 0.05) in FRAP values were found among most
(Migilo, Chiavaro, Visconti, Fogliano, & Pellegrini, 2008). Moreover, treatments. Boiled cut-kernels had the greatest decrease (57.4%),
the steaming treatment could cause matrix softening and followed by boiled whole-ears (46.3%), steamed cut-kernels
increased extractability of antioxidant components from the raw (29.4%), and steamed whole-ears (20.0%). The TEAC values were
materials (Huang, Chang, & Shao, 2006). The protocatechuic acid similar to FRAP, however, the boiled whole-ear was not differed
slightly increased during steam cooking of cut-kernels, indicating with steamed cut-kernels. These results allow us to conclude
thermal treatment could cause changes in phenolic substances in steam cooking preserved greater antioxidant content as compared
the corn by liberating the bound phenolic compounds into free to the boiling in all treatments by all assays. Other studies have
form (Xu & Chang, 2009). Hiemori, Koh, and Mitchell (2009) found suggested that boiling is generally regarded as being destructive
that this compound was generated with concomitant cyanidin-3- to antioxidant components (Krishnaswamy & Raghuramulu,
glucoside degradation during thermal processing. 1998). On the contrary, the antioxidant activities were increased
With its solubility in water, free phenolic acid can undergo for several vegetables such as carrots, spinach, mushroom, aspara-
large losses during corn processing due to considerable leaching gus, broccoli and cabbage after thermal treatment (Halvorsen et al.,
to the water used in cooking. This study found a high content 2006). The loss in antioxidants from cooked corn can be attributed
of phenolics in the water, especially in the boiling treatment, to synergistic combinations or interactions of several types of
which produced higher levels of phenolic compounds than steam- chemical reactions, diffusion of water soluble compounds, and
ing in both whole-ear corns and cut-kernels (Tables 2 and 3). This the formation or breakdown of them. Therefore, we investigated
result could be attributed to the losses of phenolic compositions the antioxidant activity of cooking water. Boiling water had high
in cooking water. However, the total of phenolic compounds in antioxidant values, especially in water from boiled cut-kernels, fol-
cooked samples and cooking water was similar to the amount lowed by whole-ears; whereas, steaming water had low values of
of phenolics in raw samples. Differences in phenolic content both antioxidant activity assays (Tables 2 and 3). However, the
could be due to breakdown of phenolics, since the sum of phen- sum of antioxidant activity of the cooked corn and cooking water
olics in cooked samples and cooking water consistently differed is different from antioxidant activity of raw materials. These
from their content in raw samples. Degradation of phenolics in results may suggest losses in their antioxidants are due to
corn may be a more serious problem than leaching. The breakdown of antioxidant compounds.
516 B. Harakotr et al. / Food Chemistry 164 (2014) 510–517

Table 4 Sustainable Agriculture of the Faculty of Agriculture of Khon Kaen

Correlations between antioxidant activities, three specific anthocyanins and phenolic University in Thailand also funded part of this work.
compounds found in waxy corna.

Correlation coefficient (r) FRAP TEAC Appendix A. Supplementary data

Monomeric anthocyanin content 0.77** 0.90**
Cyanidin-3-glucoside 0.77** 0.89** Supplementary data associated with this article can be found, in
Pelargonidin-3-glucoside 0.80** 0.82**
the online version, at http://dx.doi.org/10.1016/j.foodchem.2014.
Peonidin-3-glucoside 0.83** 0.89**
Total phenolic content 0.83** 0.95** 05.069.
Protocatechuic acid 0.60* 0.79**
p-Hydroxybenzoic acid 0.58* 0.77** References
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