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JOURNAL OF

FOOD COMPOSITION
AND ANALYSIS

Journal of Food Composition and Analysis 16 (2003) 147153


www.elsevier.com/locate/jfca

Original Article

Comparison of the cholesterol content of Brazilian chicken and


quail eggs
N. Bragagnolo*, D. B. Rodriguez-Amaya
Departamento de Ci#encia de Alimentos, Faculdade de Engenharia de Alimentos, Universidade Estadual de Campinas, C.P.
6121. 13083-970, Campinas, SP, Brazil
Received 14 January 2002; received in revised form 15 May 2002; accepted 6 June 2002

Abstract
There was no signicant difference in the cholesterol levels of large, white- and dark-shelled eggs.
Although slightly lower, the cholesterol concentration of small, white-shelled eggs was also not signicantly
different in terms of mg/g yolk. The cholesterol content of the quail egg in mg/g yolk was very similar to
that of the chicken egg. No signicant difference was seen also when the cholesterol levels of raw and
cooked chicken eggs were compared. The overall average of the cholesterol content of the chicken egg was
12.0 mg/g yolk and that of the quail egg was 12.1 mg/g yolk.
r 2003 Elsevier Science Ltd. All rights reserved.
Keywords: Chicken egg; Quail egg; Cholesterol; HPLC analysis

1. Introduction
Appreciated for their nutritive value and functional properties, eggs are an important item in
the human diet. In developing countries, it is often the only animal protein source that is
accessible to the general population. It is a major source of dietary cholesterol and consumer
concern about the association of cholesterol with coronary heart disease has lowered
consumption.
There are some conicting views and results in relation to the analytical methods for
cholesterol. Colorimetric determination of the cholesterol concentrations of eggs and egg products
had been questioned because interfering compounds could lead to signicant overestimation
(Beyer & Jensen, 1989a). The extent of overestimation, however, would depend on the
colorimetric method used (Bragagnolo & Rodriguez-Amaya, 1993) and some results
*Corresponding author. Tel.:+55-19-3788-2160; fax: +55-19-3289-2832..
E-mail address: neura@fea.unicamp.br (N. Bragagnolo).
0889-1575/03/$ - see front matter r 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0889-1575(02)00129-1

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N. Bragagnolo, D. B. Rodriguez-Amaya / Journal of Food Composition and Analysis 16 (2003) 147153

(Rangachar, Setty, & Hedge, 1970; Sainz, Gonzales, Roca, & Alemany, 1983) were comparable to
those obtained by gas chromatography (GC) or high-performance liquid chromatography
(HPLC) as pointed out by Maurice, Lightsey, Hsen, Gaylord, and Reddy (1994). The
chromatographic techniques are favored because of their ability to separate and quantify
cholesterol specically, although Maurice et al. (1994) noted that reported levels obtained by GC
were higher than those determined by HPLC. Jiang, Fenton, and Sim (1991) found no signicant
difference in egg cholesterol values obtained by enzymatic, GC and HPLC methods. Based mainly
on data from colorimetric assays, the cholesterol content was estimated in the United States to be
550 mg/100 g of egg (USDA, 1975). Reecting results of chromatographic methods, the revised
value is 425 mg/100 g of egg (USDA, 2000).
The extraction and saponication steps had also been assessed. Van Elswyk, Schake, & Hargis
(1991) observed that saponication of the lipid extract resulted in a signicantly lower value than
direct saponication of the sample. However, Bitman & Wood (1980), Fenton & Sim (1991) and
Maurice et al. (1994) did not detect signicant difference in egg cholesterol determined by these
two procedures.
Natural variation between samples has also been reported, the cholesterol level in eggs
varying with species, breed, hens age, egg and yolk weight, and diet (Riad, Kicka, Osman, &
! & Villanua,
!
Kamar, 1981; Naber, 1983; Al-Zubaidy & Al-Taha, 1984; Villanua
1988;
Beyer & Jensen, 1989b, 1983; Pandey, Panda, Maitra, & Mahapatra, 1989; Jiang, Cherian,
Robinson, & Sim, 1990; Chung, Ferrier, & Squires, 1991; Jiang & Sim, 1991; Maurice et al., 1994).
Hargis (1988), however, concluded that dietary, genetic and pharmacological manipulation to
reduce cholesterol in eggs have only been marginally successful. Chung et al. (1991) considered
selection of eggs for lower cholesterol content impractical under current commercial operating
conditions.
Brazil does not have its own data on the cholesterol levels of chicken and quail eggs. This work
was therefore carried out to ll this information gap, while taking advantage of renements in the
analytical method. Quail egg has become a popular item in salad buffet tables of Brazilian
restaurants.

2. Materials and methods


2.1. Samples
Thirty one-dozen packs, 10 packs for each of large white-shelled, large dark-shelled and small
white-shelled chicken eggs, were bought at different times from different grocery stores in
Campinas, S*ao Paulo State, Brazil during a period of 8 months. From each pack, three eggs were
randomly taken, the yolks were separated from the albumen and mixed.
For the quail eggs, 10 three-dozen packs were also purchased from different grocery stores at
different times. From each pack, 10 eggs were taken at random and the yolks were separated and
homogenized.
To compare the cholesterol contents of raw and boiled eggs, three one-dozen packs of chicken
eggs were bought. Each pack was divided, the yolks of six eggs were pooled and samples were

N. Bragagnolo, D. B. Rodriguez-Amaya / Journal of Food Composition and Analysis 16 (2003) 147153

149

taken for analysis. The other six eggs were boiled for 5 min, the yolks were then taken and
homogenized.

2.2. Determination of cholesterol


Two methods were initially compared. In one method, the yolk sample (0.2570.01 g) with 5 ml
of 2% KOH in absolute ethanol was heated in a 50 C water bath according to Jiang et al. (1991),
except that heating was carried out for 120 min instead of 60 min. The mixture was then cooled
under running water and 5 ml of distilled water were added. Cholesterol was extracted twice with
10 ml of hexane. An aliquot (3 ml) of the hexane extract was dried under nitrogen, redissolved in
3 ml of acetonitrile:isopropanol (70:30) and injected into the HPLC equipment. In the second
method, lipids of the yolk sample (10 g) were extracted with chloroformmethanol (2:1) according
to Folch, Lees, & Stanley (1957). An aliquot of the lipid extract (3 ml) was dried under nitrogen
and saponied with 12% ethanolic KOH (10 ml) for 15 min at 80 C (Bohac, Rhee, Cross, & Ono,
1988). After cooling and addition of 5 ml distilled water, the unsaponiable material was extracted
twice with 10 ml of hexane, 3 ml of the extract was dried under nitrogen, redissolved in 3 ml of the
mobile phase, and cholesterol was quantied by HPLC.
Additionally, a certied reference material, SRM 1845 powdered egg (NIST, USA), was
analyzed by the two methods.
The HPLC apparatus consisted of a Varian chromatograph, equipped with a ternary solvent
delivery system (Model 9010). Rheodyne injector with a 10 ml loop, Waters photodiode array
detector (Model 990) and a Hewlett-Packard recorder (Model 2225 D). The analytical column
was Spherisorb ODS-2 (5 mm), 4.6  150 mm, preceded with a 4.6  10 mm Spherisorb ODS-2
(5 mm) guard column. The mobile phase (ow rate, 1 ml/min) consisted of acetonitrile:isopropanol
(70:30 v/v). Each run took 15 min. Absorption spectra were taken at 190300 nm and the
chromatograms at 210 nm. All solvents were reagent grade for extraction and HPLC grade for
chromatography. Quantication was carried out by external standardization. The standard curves
were linear and bracketed the samples concentrations. Calibration was done on each day of
analysis. Aside from spiking, the peaks identity and also its purity were veried by means of the
spectra obtained with the photodiode array detector, taken at the maximum and at the ascending
and descending slopes of the peak (Fig. 1). The identity and purity of cholesterol accumulated
from several runs were also shown by mass spectrometry, the spectrum being identical with that of
a cholesterol standard (Sigma, USA) with the molecular ion at m/z 386. The electron impact mass
spectrum was obtained with a VG model Quatro instrument. Direct insertion probe was used at
an ionizing voltage of 70 eV and ion source temperature of 240 C.

2.3. Statistical analysis


To verify signicant differences in relation to size and shell color, and between cooked and raw
eggs, the results were analyzed by analysis of variance, using Statgraf software, version 4.0 (1989).
The Tukeys test was utilized to compare means at a 5% signicance level.

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N. Bragagnolo, D. B. Rodriguez-Amaya / Journal of Food Composition and Analysis 16 (2003) 147153

Fig. 1. Typical HPLC chromatogram of the cholesterol in eggs and the mass spectrum and the UV spectra obtained
with the photodiode array detector, taken at the maximum and at the ascending and descending slopes of the peak.
Chromatographic conditions: Spherisorb ODS-2 column (150  4.6 mm, 5 mm); ow rate, 1 ml/min; mobile phase,
acetonitrile:isopropanol (70:30 v/v). Absorption spectra were taken at 190300 nm and the chromatogram processed at
210 nm.

3. Results and discussion


3.1. Evaluation of the method
The cholesterol contents of three samples of fresh eggs prepared by direct saponication or
saponication of the lipid extract are shown in Table 1. For the three samples analyzed, the
cholesterol levels obtained by direct saponication were higher than those obtained by
saponication of the lipid extract. This nding is in agreement with Van Elswyk et al. (1991),
but contrary to Bitman and Wood (1980), Fenton and Sim (1991) and Maurice et al. (1994), who
found the same values for both methods.

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151

Table 1
Comparison of the cholesterol concentration of egg yolk (mg/100 g) obtained by direct saponication and
saponication of lipid extract
Sample

Direct saponication

Saponication of lipid extract

1
2
3

12.872.3a
14.272.5a
13.171.2a

11.471.5b
13.372.3b
11.971.4b

Note: Means and standard deviations of three samples analyzed in triplicate. Means in the same row bearing different
letters differ signicantly (Po0.05).

Table 2
Cholesterol levels of chicken and quail eggs
Samples

Chicken egg
Large egg, white shell
Large egg, dark shell
Small egg, white shell
Overall mean
Quail eggs

Weight (g)a

Cholesterolb

Eggc

Yolk

mg/g yolk

mg/g egg

mg/egg

6472
6775
5274
6178
9.870.5

1971
1972
1472
1773
3.270.3

12.371.1a
12.370.4a
11.371.2a
12.071.2
12.170.7a

3.770.3ab
3.570.1b
3.070.4c
3.470.4
3.970.2a

233723a
234749a
158724b
209740
3972c

Note: Values in the same column bearing the same letter are not signicantly different (Po0.05).
a
Each value is the mean and standard deviation of 30 chicken eggs and 100 quail eggs.
b
Each value is the mean and standard deviation of 10 composite samples of three chicken eggs or 10 quail eggs. All
analyses were done in duplicate.
c
Edible portion.

Using direct saponication, the cholesterol content of the certied reference material (SRM
1845, powdered egg) was found to be 19.0670.08 mg/g, compared to the certied value of
19.070.2 mg/g. With saponication of the lipid extract, a value of 18.170.7 mg/g was obtained.
The recovery percentages for direct saponication were 98.170.5 and 98.970.1%, respectively,
when 0.1 and 0.05 mg of cholesterol were added.
Direct saponication was therefore adopted in the present work. It has the added advantage of
being rapid and easy to perform, apart from not employing toxic reagents. The advantage of lipid
extraction followed by saponication is that the same lipid extract for the quantication of
cholesterol can be used for the determination of total lipids and the fatty acid composition.
3.2. Cholesterol content
The cholesterol contents of chicken and quail eggs produced in Brazil are shown in Table 2.
There was no signicant difference in the cholesterol level of large, white- or dark-shelled eggs in
terms of mg/g yolk, mg/g egg and mg/egg. The cholesterol concentration of the small,

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N. Bragagnolo, D. B. Rodriguez-Amaya / Journal of Food Composition and Analysis 16 (2003) 147153

Table 3
Cholesterol levels of raw and boiled chicken eggs
Weight (g)a

Samples

Cholesterolb

Egg

Yolk

mg/g yolk

mg/g egg

mg/egg

Raw egg
Large egg,
Large egg,
Large egg,
Large egg,

white shell
white shell
dark shell
dark shell

5674
5772
5573
6571

1972
1971
1871
1972

10a
11a
13a
10a

3.4a
3.5a
4.3a
2.9a

192a
201a
233a
185a

Boiled egg
Large egg,
Large egg,
Large egg,
Large egg,

white shell
white shell
dark shell
dark shell

5873
5872
5771
6672

2072
1872
1771
1971

10a
11a
14a
10a

3.5a
3.4a
4.1a
2.9a

204a
198a
233a
189a

Note: Values in the same column bearing the same letter are not signicantly different (Po0.05).
a
Each value is the mean and standard deviation of 6 chicken eggs.
b
Each value is the mean of duplicate analysis.

white-shelled egg was also not signicantly different in terms of the mg/g yolk. On a mg/g egg and
mg/egg, the small egg had lower values, reecting the difference in weight.
Also using HPLC for quantication, Jiang et al. (1991) obtained 195 mg/egg or 11.7 mg/g yolk,
Beyer and Jensen (1989a) 199 mg/egg or 11.0 mg/g yolk and Beyer and Jensen (1989b) 195 mg/egg
and 11.7 mg/g yolk, the eggs weighing about 60 g in all cases. These values are between those
obtained in the present study for large and small eggs. Jiang et al. (1991) used direct saponication
and Beyer and Jensen (1989a,b) saponication of extracted lipids. USDAs current values
(USDA, 2000) are 247 mg/egg for 58 g egg and 276 mg/egg for 65 g egg, thus higher than our and
the other authors values.
The cholesterol concentration of the quail (Japanese Corturnix coturnix) egg was very similar to
that of the chicken egg in terms of mg/g yolk, contrary to the popular belief in Brazil that quail
eggs are lower in cholesterol content. The data obtained in the present study, however, are much
lower than the USDA (2000) values of 8.4 mg/g egg and 76 mg/egg. Maurice et al. (1994) reported
45 mg/egg, closer to the 39 mg/egg obtained in the present study.
Comparing raw and boiled chicken eggs (Table 3), no signicant difference was seen in all units
(i.e., mg/g yolk, mg/g eggs, mg/egg) calculated.

Acknowledgements
The authors wish to thank the Funda@*ao de Amparo a" Pesquisa do Estado de S*ao Paulo
(FAPESP) for the nancial support and Mr Mark Prescott (University of Liverpool) for
recording the MS spectra.

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153

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