11

Quantification of Phenolic Content, Tocopherols and

Phytosterols in Different Morphological Fractions
Obtained as Byproducts of the Corn Wet Milling
R. HERNÁNDEZ-SOTO, G. SANDOVAL-FABIAN, A. CARDADOR-MARTÍNEZ AND
M. ESTARRÓN-ESPINOZA.

ABSTRACT
Byproducts from the corn wet milling starch process may be the source
of diverse bioactive compounds such as phenolic compounds, tocopherols
and phytosterols, all of which are retained in the by-products in
considerable quantities. The extraction of such constituents from by-
products was made with ethanol or chloroform. Phenolic contents
ranging from 0.27 to 14.98 mg/g were found in all fractions. The
quantification of tocopherols was carried out by means of a new
colorimetric test, from which total tocopherol contents of 0.08 to 9.19
mg/g were found. All extracts showed remarkable antioxidant
activities. Finally, the phytosterols from the by-products were extracted
and quantified, in which the lipids fraction (LF) contains the higher
quantity of phytosterols, where β-sitosterol was the main compound.
The results suggest that the gluten and lipids fractions could be used
to obtain useful phytochemicals.

11.1. INTRODUCTION
In Mexico, the starch industry and its derivatives processed
approximately 2.2 million tons of yellow corn during the period of 1995-
2005. Corn is processed by wet milling to obtain starch, but only around
60% of the grain is used, generating 136 thousand tons/year of by-
products of low commercial value, which can only be locally

xxxxxxx

* Corresponding author: E-mail: gsandoval@ciatej.net.mx

188 Nutraceuticals and Functional Foods

commercialized and basically destined for animal feed due to their high
perishable character CANAMI (2007).

Although corn waste can hardly be classified among the most

hazardous wastes, its treatment is very important in view of the great
volume of waste materials involved (Arvanitoyannis and Tserkezou,
2008). The accumulation of these products can become an environmental
contamination problem. On the other hand, the constant growth of the
starch industry makes it necessary to diversify the uses of the generated
by-products in order to achieve a sustainable development. In the wet
milling process of corn, the main products are starch, sugars and corn
oil, therefore a large portion of fatty acids, proteins and fibers are
discarded. However, these residues may be an attractive source of diverse
bioactive compounds, such as phenolic compounds, phytosterols,
tocopherols and tocotrienols (Kurilich and Juvik, 1999; Niwa, Doi, Kato
and Osawa, 2001; Perez-Jimenez and Saura-Calixto, 2005; Zhao,
Egashira and Sanada, 2005).

Phenols, phenolic acids, cinnamic acids, cumarines, isocumarines,

chromonols, lignanes and neolignanes, flavonoids (flavonols, flavones,
isoflavones, among others) and tannins are all phenolic compounds,
stressing on the ferulic acid and diferulate contents in grains (Adom
and Liu, 2002; Martínez, Periago and Ros, 2000). Phenolic acids are
mainly found in the bran of grains. Ferulic and caffeic acid (cinnamic
acids) work to prevent the formation of carcinogens by means of
precursors and block reactions of carcinogens with cell macromolecules
(Martinez-Tome et al., 2004).

The tocopherols and tocotrienols are fat-soluble compound groups,

denominated generally as vitamin E. They are well recognized as
antioxidants in vegetable oils, and their presence increases the stability
of lipids against auto-oxidation (Xu, Hua and Godber, 2001). The
tocopherols display a high degree of relation with the polyunsaturated
fatty acids (PUFA) content in vegetal oils, comprising the major fatty
acids: linoleic, oleic and palmitic acid (Goffman and Bohme, 2001). They
also contain selenium in its reduced state and inhibit the formation of
nitrosamines at low pH thus preventing the formation of carcinogens
by means of precursors (Slavin, 2000).

Antioxidants found in cereals are water soluble, fat-soluble and

approximately one-half are insoluble. A large part of insoluble
antioxidants are bound as cinnamic acid esters to arabinoxylan side
chains of hemicelluloses. Phytic acid, present in grains, also acts as an
antioxidant by chelating metals such as iron, which may catalyze the
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 189

formation of free radicals (Slavin, 2000). Cereals have specifically been

the object of diverse studies in determining antioxidant activity using
whole grain from wheat, barley, oats and rye (Adom and Liu, 2002;
Perez-Jimenez and Saura-Calixto, 2005; Xu et al., 2001; Zielinski and
Kozlowska, 2000), corn and corn steep liquor (Niwa et al., 2001; Zhao et
al., 2005). The phenolic and tocopherol content in corn by-products
confers upon them an elevated level of antioxidant activity (Niwa et al.,
2001).

The phytosterols and their reduced forms, phytostanols, are sterols

of plant origin, widely distributed in nature and whose chemical
structure is similar to that of cholesterol. For years, hypocholesterolemic
effects have been attributed to these compounds. They are also
considered as a factor in the risk reduction of cardiovascular damage
(Moreau, Whitaker, and Hicks, 2002; Valenzuela and Ronco, 2004).
Despite the fact that the extraction of these bioactive compounds is
achieved through diverse sources, obtaining these products by means
of industrial by-products has not been completely analyzed. In the case
of corn, there are reports for obtaining these products from the corn
fiber and corn germ (Moreau and Hicks, 2005; Moreau, Singh, Nuñez,
and Hicks, 2000; Yadav, Moreau and Hicks, 2007). The presence of these
compounds has been reported as well in by-products from the ethanol
industry (Winkler, Rennick, Eller and Vaughn, 2007). The antioxidant
properties of the corn steep liquor leftover material of starch-
manufacturing processes also have been reported (Niwa et al., 2001).

Consequently, the objective of the present study was revalue the

by-products generated from processing yellow dent corn to obtain starch
by the wet milling process, by extraction and quantification of phenolic
compounds, tocopherols and phytosterols as well as its antioxidant
properties. All these compounds have phytochemicals propierties that
are higly valued in the food and pharmaceutical industries.

11.2. MATERIALS AND METHODS

11.2.1. Biological Material
The different by-products were obtained from yellow dent corn and
processed by wet milling. The byproducts were identified as follows:
corn wet fiber (CWF) and corn dry fiber (CDF); both are found in the
outer cap or fraction of grain with a high fiber content; a high humidity
content in the first case (CWF), and subject to thermal processes in the
second case (CDF). Germ is the fraction of the grain where the highest
oil content was originally found. The fraction used in this study was
190 Nutraceuticals and Functional Foods

generated once the germ was processed to obtain crude corn oil. Gluten,
the dry residue of corn after the starch and germ has been extracted.
Finally, the oily residues generated from the extraction and refining
process of crude corn oil were denominated as lipid fraction (LF).
Samples were collected according to standard sampling procedures and
stored in sealed polyethylene bags at 4oC until their analysis.

11.2.2. Reagents and Standards

³
All reagents and standards used were rated ≥ 95% purity, Folin-
Ciocalteu reagent, Gallic acid, α-tocopherol, Oleic acid, Dihydro-
cholesterol, Stigmastanol, Campesterol, Stigmasterol and β-sitosterol.
All were bought from Sigma-Aldrich Mexico. All solvents used for
extractions and other substances used were reagent grade or HPLC
grade.

11.2.3. Sample Preparation

The moisture content of corn residues was determinate according to
the NOM-F-83-1986, the quantification of total lipid and insaponificable
matter was carried out by recording the dry weight of chloroform extracts
and ethereal extracts obtained after of the saponification.

11.2.4. Extraction of Bioactive Compound

The extracts were obtained according to the drawing in Fig. 11.1. Germ,
Gluten, CDF and CWF were crushed using a food processor (Moulinex)
and later were milled to a 20 mesh particle size with a rotary-tapping
shaker (RO-TAP, RX-29, W.S. Tayler, OH, USA), until 25 g of the samples
were obtained. The LF byproducts were processed directly. All by-
products were extracted with 250 mL of chloroform or with 250 mL of
ethanol, at a temperature of 30oC for one h. The solid phase was
separated using a Wathman filter paper No 20, which was discarded.
The ethanolic and chloroform extracts were kept in amber glass bottles
and stored at 4oC until their analysis.

11.2.5. Total Phenolic Content

The total phenolic content of each extract was determined by the Folin-
Ciocalteu method (Singleton, Orthofer, Lamuela-Raventós, and Lester,
1999). Briefly, 0.06 mL of the extract was mixed with 4.74 mL of water
in a test tub and oxidized with 0.3 mL of 0.5 N Folin-Ciocalteu reagent,
then the reaction was neutralized with 0.9 mL sodium carbonate
solution (20%). The absorbance values were obtained by the resulting
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 191

Fig. 11.1: Extraction method

blue color measured at 760 nm with a Cintra 6 UV spectrophotometer

(GBC Brand, V3579) after incubation for 2 h at 25°C. Quantification
was done on the basis of a standard curve of gallic acid. Results were
expressed as micrograms (mg) of gallic acid equivalent per gram of
dry matter (DM).

11.2.6. Total Tocopherol Content

The determination of total tocopherol content was done in colorimetric
form, adapting the Chang method (Chang, Lee and Lee, 2005). The
principle of that method is the interaction between cupric agents and
free fatty acids forming colored cupric soaps with the cage-like complex,
Cu2(C18H34O2)4. Due to the fact that the tocopherols were not colored
directly by cupric acetate pyridine, and knowing the correlation between
the tocopherols and the fatty acid content, the tocopherols were mixed
with oleic acid. This was used for indirect coloration of tocopherols
(Chang et al., 2005; Goffman and Bohme, 2001). In test tubes, 10 mL of
the ethanolic extracts and 20 mL of hexane were mixed and vortex mixed.
After standing for one h, 5 mL of the upper layer was taken and 1.0 mL
of copper pyridine acetate (5% w/v, pH 6.01) was added. The mixture
was shaken in a vortex for 90 seconds and the samples were placed in
the dark for 5 minutes. Finally, the absorbance was measured with a
Cintra 6 Spectrophotometer (GBC Brand; V 3579), at 715 nm.
Quantification was done on the basis of a standard curve of α-tocopherol
mixed with oleic acid (4:1 w/w). Results were expressed as mg of α-
tocopherol equivalents (TE) per gram of DM.
192 Nutraceuticals and Functional Foods

11.2.7. Total Antioxidant Activity

Total Antioxidant activity was measured by a modified version of the
original Brand-Williams, et al. (1995) method. In a ninety-six-well plate,
0.02 mL of each extract was added, previously standardized to 150 μM
equivalents of gallic acid based on the total phenolic concentration or
0.02 mL of BHT, catequine and gallic acid, all at 150 μM. All samples and
standards were added with 0.2 mL of 2.2-diphenyl-1-picryhydrazyl
(DPPH) 150 μM in 80% methanol. The samples and standards were
prepared in triplicate. After 90 min. in the dark, the absorbance was
measured at 520 nm in a plate reader (BioRad, Model 680 XR, microplate
reader). The total antioxidant activity was calculated as μmoles of gallic
acid by gram of DM, Antioxidant activity also was calculated as % of
discoloration = [(A of control – A of sample)/ (A of control)] × 100. The control was
the DPPH solution in methanol.

11.2.8. Phytosterol Content

The chloroform extracts previously obtained were subjected to vacuum
evaporation in a rotavapor (Büchi Labortechnik AG CH 9230) at 35oC
until dry. Once the lipidic fraction was obtained, the sample was
saponified according to the Giacometti method (2001), with some
modifications. The dry sample was added to 50 mL methanolic KOH 2
N, and boiled at 55–60 oC for one h. After the reaction, once that solution
was at room temperature, 50 mL of water was added and the
unsaponifiable matter was extracted three times with 40 mL of diethyl
ether. Then the combined ether extracts were washed three times with
50 mL of water. Finally the washed etherous phase was filtered through
filter paper with anhydrous sodium sulphate. The unsaponifiable matter
was obtained after evaporating the ethyl ether. Then 8 mg of the
insaponifiable matter was mixed with 350 μl of dihydrocholesterol (2
mg/mL) external standard solution and finally dissolved in 1 mL
chloroform for GC analysis. Prior to GC analysis, 100 μl was transferred
to an Eppendorf tube containing 33 μl of the silylating mixture
(Hexamethyldisilazane, Trichloromethylsilane and Pyridine at a ratio
of 2:1:10 v/v/v). After 15 min, the mixture was centrifuged and from the
supernatant, 1 μl was taken for GC analysis; all samples were analyzed
in duplicate. A GC chromatograph (Hewlett Packard 6890) equipped
with a flame ionization detector, an HP 6890 auto sampler, and an HP-
5 column (30 m long, 0.32 mm in diameter and 0.25 μm film thickness),
was used to carry out the chromatography analysis, with the following
temperature program: the temperature increase to 220°C at 270°C a
rate of 8°C/min, then hold isothermal the rest of time analysis. The
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 193

injector and detector temperatures were set at 300oC. Flow rate of the
carrier gas, (helium) was 2 mL/min and the injection split ratio was
20:1, the injection volume was 1 μl. The overall system was controlled
by HP-MS ChemStation G17001DA (D.02.00.SP1 MSD, 2005) Software.
The identification of phytosterols was done by comparing the standard
retention time. Quantification was done on the basis of a standard curve
of phytosterols.

11.2.9. Experimental Design

All values were reported on a dry matter (DM) basis. All by-products
were extracted by triplicate, the determination of phenolics compounds,
tocopherols, phytosterols and antioxidant activity were performed twice
(3x2). The results were presented as mean ± standard deviation values.

11.3. RESULTS AND DISCUSSION

11.3.1. Total Phenolic Content
The total phenolic content measured in the ethanolic extracts of the
different by-products of corn is presented in Table 11.1. The LF fraction
showed the highest phenolic content, followed by gluten and CWF. CWF
has two times the concentration of CDF, suggesting that temperature
treatment is responsible for the decrease in phenolics. CWF and CDF
had comparable phenolic contents with similar products, although lower
than those reported by Zhao et al. (2005) for corn bran (a fraction of the
corn dry-milling industry), while the LF and Gluten fractions have a
higher content in the findings content than in the findings for coarse
fiber, and whole cereals.
Table 11.1: Total phenolic contents in the ethanolic extracts in corn byproducts
Byproduct Total phenols
(mg GAE/g)
CWF 4.70 ± 0.33
CDF 2.39 ± 0.06
Germ 0.27 ± 0.03
Gluten 5.82 ± 1.48
LF 14.98 ± 1.33

Germ fraction possessed the lowest content of phenolic compounds,

but it is important to indicate that the Germ fraction analyzed was
previously processed for corn oil extraction and dry heated, which causes
a decrease in the quantity of phenolic compounds from 20-50%
(Martinez-Tome et al., 2004).
194 Nutraceuticals and Functional Foods

Due to the nature of the materials used in the determination, and

that generally diverse solvents and analytical method were used in
the extraction of these compounds, as well as taking into account that
generally, the quantity of compounds extracted decreases with the
polarity of the solvents used (Zielinski and Kozlowska, 2000), it is not
possible to do a direct comparison with the data found in the literature.
However, it is possible to compare the results obtained with the data
generated by certain authors for similar materials, such as Adom et
al. (2002) who reported the total phenolic content of yellow corn of
2.64 mg GAE (gallic acid equivalents)/g of DM, using ethanolic extracts
(80%). On the other hand, Zhao et al. (2005) reported a content of
main phenolic acid in corn bran in 43.2 mg/g of DM. During the wet
milling of corn, two types of corn fiber are produced: coarse fiber, which
is primarily from pericarp, and fine fiber, which is from the inner
cellular endosperm portion of the corn kernel, Yadav et al. (2007)
analyzed these fibers and reported a total content of phenolics in the
coarse fiber of 4.50 mg/g of dm and 1.08 mg/g of DM in the fine fiber
using a mixture of chloroform and methanol (2:1; v/v) as solvent. Lopez-
Martinez et al. (2009) measured the total phenolic content of eighteen
Mexican maize phenotypes with a range variation from 2.15 to 34.01
mg GAE/g DM of whole grain flour.

11.3.2. Total Tocopherols Content

Table 11.2 shows the content of tocopherols determinated from different
by-products. Again, it is the LF fraction which had the highest content
of tocopherols, followed by Gluten, whereas Germ fraction had the
poorest content of tocopherols. It had just 0.08 mg/g. The content of
tocopherols, like the content of phenolic was affected by the thermal
treatment, which partially destroyed the tocopherols remaining in the
germ fraction. A similar effect occurred with the corn fiber, given the
fact that CWF contains approximately three times more tocopherols
than the CDF and is also subjected to heat drying.

Table 11.2: Total tocopherol content from the ethanolic extracts in corn byproducts
Byproduct Total tocopherols
(mg TE/g)
CWF 0.49 ± 0.08
CDF 0.18 ± 0.04
Germ 0.08 ± 0.02
Gluten 3.14 ± 0.09
LF 9.19 ± 1.25
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 195

The quantities determined for CWF and CDF fractions are lower than
in the findings reported by Moreau et al. (2005), who measured the total
tocopherol content of yellow corn, as well as corn bran, and corn germ oil,
in both wet milling and dry milling, using an ethanolic extract (50°C),
reporting 1.60 mg/g of DM of total tocopherol content in corn kernel, 0.54
mg/g by corn bran, 1.19 mg/g in corn germ (fraction of wet-milling) and
0.36 mg/g in corn germ (fraction of dry-milling), It should be noted that
the corn germ analyzed by Moreau was quantified before being extracted
industrially to obtain oil from corn germ, which explains the high content
of tocopherols reported in these fractions in comparison with the low
content of tocopherols quantified in Germ fraction in the present study.
However, the quantities determined for Gluten and LF fractions are higher
than the values reported in the same study.

On the other hand, in corn by-products from the ethanol industry,

Winkler et al. (2007) reported the tocopherol content obtained under
several extraction conditions: 1.82 mg/g extracted with hexane, 0.73
mg/g extracted with ethanol and 1.71 mg/g with CO2 in supercritical
extraction conditions.

In our results, it can be observed that the Gluten and LF fractions

not only have a higher tocopherol content than the findings for by-
products from the ethanol industry, but also that of Goffman et al. (2001)
for diverse corn varieties, 0.91 to 1.57 mg/g using petroleum ether as
the extraction solvent, as well as the total tocopherol content findings
of Panfili et al. (2003) in different cereal species, using ethanol in alkaline
digestion (70°C for 45 min) 0.0721 mg/g for oats, 0.0743 mg/g for soft
wheat, 0.0747 mg/g for barley and 0.0669 mg/g for corn. Moore et al.
(2005), found a concentration range from 0.0034 mg/g to 0.0101 mg/g of
DM in soft wheat.

The great variability of total tocopherol content in cereals not only

depends on the genotype and the environmental conditions in which
the grains develop (Perez-Jimenez and Saura-Calixto, 2005), but also
on the method of extraction (Moreau and Hicks, 2005). Our results
indicated that a significant quantity is lost not only during the oil
refining process but during the diverse steps of corn industrialization
for obtaining products such as starch, oil and ethanol. However,
considerable quantities of bioactive compounds that can be exploited to
produce nutraceuticals still remain in the discarded byproducts.

Finally, it is important to emphasize the fact that tocopherol

quantification using the colorimetric process is indirect, given that the
196 Nutraceuticals and Functional Foods

tocopherol and oleic acid contents are proportionally related in proportions

ranging from 19.5–30.5% of the total fatty acids in corn. The oleic acid
present in the sample reacts with the cupric acetate pyridine complex.
However, diverse studies cite a high positive correlation found between
tocopherols and the ratio tocopherols/PUFA (Chang et al., 2005; Goffman
and Bohme, 2001). Furthermore, according to Chang’s results, the
correlation coefficients “r” determined for several samples of tocopherols
mixed with oleic acid, by HPLC analysis and by colorimetric method was
very similar. It is possible then, to use the colorimetric method with an
elevated confiability index (Chang et al., 2005).

11.3.3. Antioxidant Activity

The antioxidant capacity measured by DPPH discoloration reaction in
the ethanolic extracts ranged from 0.10 to 28.21 μmoles of gallic acid by
gram of dry matter. These values are concentrated in Table 11.3. The
lowest antioxidant activity is presented by the Germ by-product, which
coincides with the results obtained for the phenolic compounds and
tocopherols, thus directly affecting the antioxidant activity (Xu et al.,
2001). The highest value for antioxidant activity is presented by LF
fractions, which also presents the highest contents for phenolic
compounds as well as tocopherols.

CWF and gluten showed similar values of antioxidant capacity, six

times lower than LF but two and-a-half times higher than CDF. Although
Gluten had at least six times more tocopherols than CWF, their
antioxidant capacity is similar, suggesting that the total antioxidant
capacity of a product not only originates from the sum of antioxidant
capabilities of each of its compounds, but also depends on the
microenvironment in which the component is found, as these compounds
can interact with others producing synergistic or inhibitory effects
(Zielinski and Kozlowska, 2000).

Table 11.3: Antioxidant activity from the ethanolic extracts in corn byproducts
Byproduct μMol gallic acid/g % Discoloration
CWF 5.01 ± 0.13 41.6
CDF 2.02 ± 0.16 28.6
Germ 0.10 ± 0.04 17.3
Gluten 5.68 ± 0.36 43.0
LF 28.21 ± 3.07 92.5

Levels of antioxidant activity for CWF and Germ coincided with the
findings of Martínez-Tomé et al. (2004) and Yadav et al. (2007), who
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 197

indicated that antioxidants from corn kernel are found in greater

proportion in bran. On the other hand, Beta et al. (2005), measured the
antioxidant activity in different fractions of pearled wheat, using
ethanolic extracts (80%), and reported that antioxidant activity was
concentrated in the external fraction of the grain with a 20% of DPPH
discoloration compared to only 2.5% of DPPH discoloration in the flour.
Accordingly, the indices of antioxidant activity obtained for the samples
depend on the characteristics of the pearling process.

Cereals are relatively high in antioxidant activity. Diverse data in

the literature (Adom and Liu, 2002; Perez-Jimenez and Saura-Calixto,
2005), indicated that ferulic acid, a phenolic compound mainly present
in the by-products studied, prevents the formation of carcinogens by
means of precursors, and despite being found joined to the lateral chains
of arabinoxylane of hemicellulose found in the colon, they may be
released and absorbed, increasing the defense against carcinogenic cells
(Slavin, 2000). In addition, the literature shows that the antioxidant
activity of the corn kernel is the highest, followed by wheat, oats and
rice (Zielinski and Kozlowska, 2000). Based on the results obtained in
this investigation with respect to the antioxidant properties of corn
byproducts, it is possible to consider that LF and Gluten fractions can
be considered as alternative sources to phytochemicals.

11.3.4. Total Phytosterols Content

The total phytosterols content determinated from corn byproducts are
provided in Table 11.4. CWF and CDF had similar levels of phytosterols
(0.37 and 0.30 mg/g), which is comparable to the value reported by Jiang
and Wang (2005), who measured the total content of phytosterols in
different fractions of cereal by-products, using a mixture of chloroform
and methanol (2:1) as solvent, (0.3 mg/g for corn fine fiber, 4.5 mg/g for
rice bran, 1.2 mg/g for wheat bran and 0.7 mg/g for hull oats). In contrast,
our results Moreau et al. (2000), determined a total phytosterols content
of 0.19 mg/g in corn fiber using hexane as solvent. Due to the different
extraction method as well as the different nature from the analyzed
samples, it is not possible to do a direct comparison with the data found
in the literature. However, it is possible to compare the results obtained
with the similar material. Also, the data determined in the present study
for Germ fractions differ with respect to the findings of Moreau et al.
(2000) for germ fraction from wet milling by-products, who determined
a total phytosterols content of 0.168 mg/g. In the same way, the total
phytosterol content in the Gluten fraction in the present study was
higher than the content in the starch fraction reported by Singh et al.
198 Nutraceuticals and Functional Foods

(2003), who reports a total phytosterols content of 0.006 mg/g, and the
total phytosterols content of 0.398 mg/g for wheat flour, quantified by
Buri et al. (2004). This is probably because the aleurone layer in corn,
consisting of a single layer of living cells with thick cell walls, and having
the highest content of phytosterols among the corn by-products from
wet milling (Moreau et al., 2000), was integrated within the gluten
fraction analyzed due to a deficient pearling process of grain (Beta et
al., 2005).

Table 11.4: Phytosterol content from the chloroform extracts in corn byproducts
Byproduct Total phytosterol
(mg/100 g)
CWF 0.375 ± 0.12
CDF 0.301 ± 0.04
Gluten 2.801 ± 0.12
Germ 0.647 ± 0.26
LF 42.076 ± 7.36

Finally, the LF fraction showed the highest phytosterols content

(40.01 mg/g). This value is greater than that reported by Piironen et al.
(2000), for crude corn oil (8.09 to 15.57 mg/g) and refined corn oil (7.15
to 9.52 mg/g), suggesting that during the process of industrialization of
corn, significant amounts of phytosterols are lost. Phytosterols content
differs considerably from the findings reported by Winkler et al. (2007)
in by-products of the ethanol industry, perhaps because of the differences
in both the variety of corn used as well as the fermentation process in
which the analyzed by-products were obtained.

Likewise, the total quantity of phytosterols determined in the

extracts of by-products from the ethanol industry varies with respect to
the total quantity obtained for the main compounds from by-products
of the starch industry analyzed in this work. Indeed, CWF, CDF and
Germ fractions possess contents lower than 2.05 ± 0.09 mg/g of by-
product, determined by Winkler et al. (2007) using hexane as solvent,
while the Gluten and LF fractions surpassed the stated contents. The
phytosterol content in the different corn by-products is generous if
compared with diverse foods, especially the LF, which possesses greater
phytosterol content than main sources in these compounds such as rice
bran oil (11.9 mg/g), sesame (7.14 mg/g) and sunflower seed (4.44 mg/g)
(Ling and Jones, 1995; Piironen et al., 2000). In addition, the phytosterols
in corn are found in conjugated form with ferulic acid. The phytosterol’s
ferulates have been recognized by the hipocholesterolemic effects
(Moreau et al., 2002; Nyström, Makinen, Lampi and Piironen, 2005).
Quantification of Phenolic Content, Tocopherols and Phytosterols ... 199

The variability in the content of phytosterols reported for different

analyzed corn byproducts is wide, similar to the findings of other
compounds, which may be explained by various factors such as
differences in the methods of analysis, as well as environmental and
genetic factors of the raw material analyzed.

11.4. CONCLUSION
Corn is cultivated throughout the world and is used as raw material for
obtaining diverse industrial products, generating a large quantity of
by-products of low economic value. These by-products are susceptible
to, or available for industrial exploitation in obtaining phytochemicals
of high added value such as antioxidants, tocopherols and phytosterols,
which are found in the diverse analyzed fractions, especially in the LF
and Gluten fractions. The high content of bioactive compounds in these
fractions opens the possibility of exploitation of these byproducts for
the treatment of general health problems at a global level, through their
incorporation in the production of functional foods.

11.5. ACKNOWLEDGEMENTS
The authors of this investigation wish to thank SAGARPA (Mexico) for
financing this project and Corn Products International (CPI),
Guadalajara for corn byproducts.

11.6. REFERENCES
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Agricultural and Food Chemistry, 50(21): 6182-6187.
Arvanitoyannis, I.S., and Tserkezou, P. 2008. Corn and rice waste: a comparative
and critical presentation of methods and current and potential uses of treated
waste. International Journal of Food Science and Technology, 43(6): 958-988.
Beta, T., Nam, S., Dexter, J.E., and Sapirstein, H.D. 2005. Phenolic content and
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