The impact of selected nutritional factors on cis-9, trans

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Probl Hig Epidemiol 2011, 92(4): 840-843
The impact of selected nutritional factors on cis-9, trans-11
octadecadienoate content in milk of breastfeeding women.
Part I.
Wpływ wybranych czynników żywieniowych na zawartość kwasu cis-9, trans-11
oktadekadienowego w mleku kobiet karmiących. Część I.
Agnieszka Białek, Andrzej Tokarz, Mateusz Romanowicz
Katedra i Zakład Bromatologii, Warszawski Uniwersytet Medyczny
Wprowadzenie. Mleko matki stanowi najlepszy pokarm dla niemowląt
i małych dzieci. Kwas cis-9, trans-11 oktadekadienowy (kwas żwaczowy)
jest głównym izomerem spośród sprzężonych dienów kwasu linolowego,
obecnych w mleku różnych gatunków. Ma on duże znaczenie dla zdrowia
niemowląt jako czynnik wzrostu, przyczyniając się m.in. do efektywniejszego
wykorzystania pokarmu. Liczne czynniki decydują o jego ilości w tłuszczu
mleka.
Cel pracy. Oznaczenie zawartości tłuszczu i kwasu żwaczowego w mleku
kobiecym oraz określenie wpływu różnorodnych czynników żywieniowych
na te parametry.
Materiał i metody. Przebadano 67 próbek mleka kobiet karmiących,
których dietę poddano analizie pod kątem ich zwyczajów żywieniowych.
Natomiast analizę kwasów tłuszczowych wykonano techniką GC
z zastosowaniem kolumny kapilarnej i detekcją płomieniowo-jonizacyjną.
Wzorzec CLA FAME reference standard (Nu-Chek-Prep) posłużył do
identyfikacji kwasu żwaczowego. Ocenę statystyczną wyników wykonano
przy użyciu programu Statistica 8.0.
Wyniki. Średnie zawartości tłuszczu w mleku wynosiły 2,42±0,11%, zaś
kwasu żwaczowego – 1,38±0,09 mg/g tł. Stwierdzono dodatnią korelację
pomiędzy ilością tłuszczu i zawartością CLA w mleku oraz pomiędzy
zawartością CLA w mleku i CLA w tłuszczu, oraz ujemną korelację pomiędzy
zawartością tłuszczu a zawartością CLA w tłuszczu mleka. Ponadto inne
czynniki, takie jak np. spożycie mleka i napojów mlecznych wpływały na
zawartość tłuszczu lub zawartość kwasu żwaczowego.
Wniosek. Uzyskane wyniki wskazują, że czynniki dietetyczne mogą
oddziaływać na zawartość tłuszczu i kwasu żwaczowego w mleku
kobiecym.
Introduction. Maternal milk provides optimal nutrition to the breastfed
infants. Cis-9, trans-11 octadecadienoate (rumenic acid), main isomer of
conjugated linoleic acids in milk of different species can greatly influence
infant health as a potential growth factor, mainly by improving feed
efficiency. Numerous factors can affect its concentration in milk fat.
Aim. To examine the content of fat and rumenic acid in human milk as
well as to determine the influence of selected nutritional factors on that
content.
Material & methods. We investigated 67 milk samples of breastfeeding
women. They were anonymously surveyed, i.a. about their nutritional
habits. The fatty acid analysis was made with GC with capillary column
and flame-ionization detection. CLA FAME reference standard (Nu-ChekPrep) was used to identify rumenic acid. The statistic evaluation was made
with Statistica 8.0.
Results. The mean fat content in milk was 2.42±0.11% and the mean
rumenic acid content was established to be 1.38±0.09 mg/g of fat. We
observed a positive correlation between the fat and CLA content in milk
and between the CLA content in fat and in milk, as well as a negative
correlation between the fat and CLA content in fat. Moreover other factors,
such as milk and dairy beverage consumption, influence the fat or rumenic
acid content.
Conclusion. Our findings confirm that dietary factors can affect the fat
and rumenic acid content in human milk.
Key words: conjugated linoleic acid, CLA, maternal milk, breastfeeding,
maternal nutrition
Słowa kluczowe: sprzężone dieny kwasu linolowego, CLA, mleko kobiece,
karmienie piersią, żywienie matki
© Probl Hig Epidemiol 2011, 92(4): 840-843
www.phie.pl
Nadesłano: 10.06.2011
Zakwalifikowano do druku: 02.08.2011
Introduction
Maternal milk provides optimal nutrition for
infants. Human milk is a complex mixture of all
nutrients essential for normal growth and development
Adres do korespondencji / Address for correspondence
mgr Agnieszka Białek
Katedra i Zakład Bromatologii
ul. Banacha 1, 02-097 Warszawa
tel./fax (+ 48 22) 5720785, e-mail: [email protected]
of the infant. It contains about 3 to 5% of lipids [1]
which represent the main source of energy. Most of
them are TAG (>95%) and are located in globules
coated with areolas of milk and cellular proteins,
Białek A et al. The impact of selected nutritional factors on cis-9, trans-11 octadecadienoate content in milk ... Part I.
phospholipids, sterols, enzymes and other bipolar
substances. Many factors can affect the composition
of human milk, e.g.: stage of lactation and maternal
diet [1, 2]. Fatty acids of human milk fat originate
from adipose tissue (60%), diet (30%) and de novo
synthesis (10-12%) [3].
CLA (conjugated linoleic acid) refers to a group
of positional and geometric isomers of linoleic acid
with a pair of conjugated carbon-carbon double
bonds. They have been identified primarily in
ruminant milks, meats and dairy products. CLA have
been reported to produce a variety of physiological
effects, e.g.: antiatherogenic effect [4, 5], causing
body fat loss, increasing lean body mass [6, 7] and
anticancerogenic activity [8, 9]. The two major CLA
isomers with known physiologic activities are cis-9,
trans-11 octadecadienoate (rumenic acid, RA) and
trans-10, cis-12 octadecadienoate. Rumenic acid has
great meaning for infant health as a potential growth
factor, mainly by enhancing weight and improving
feed efficiency [10].
Maternal milk is the first choice for newborns
and infants, therefore its quality is very important for
infant health. The aim of our study was to determine
the influence of different dietary factors on the
content of fat and cis-9, trans-11 octadecadienoate
in human milk.
Material and methods
This study involved 67 breastfeeding women,
aged 20-42 years who gave birth to one child or two
children, mainly in The Gynecological-Obstetric
Department of Central Hospital of the Ministry of
Interior and Administration. All mothers agreed to
provide samples of breast milk. Both colostrum and
mature milk samples were examined. Mothers were
anonymously asked about their nutritional and dietary
habits during and after pregnancy.
Samples of breast milk (50-150 ml) were collected
by manual expression from both breasts. They were
immediately frozen and stored at –20ºC in sterile
polypropylene containers. The milk samples were
thawed only once and lipids extraction was prepared
according to a modified procedure by Chin et al. [11]
from 1.0 ml of breast milk using twice 5.0 ml of ethyl
ether: petroleum ether (1:1, v/v) after adding 2.0
ml of ethanol and 0.1 ml of ammonium hydroxide
(25%). Organic extracts were dried under nitrogen
and the extracted milk fat was weighed. Fatty acid
methyl esters were obtained by transesterification
according to Christie et al. [12] and separated and
quantified by gas chromatography using Shimadzu
GC-17A chromatograph equipped with flameionization detector. The injector and detector were
heated to 250ºC. The separation was performed on
841
BPX 70 capillary column (60 m × 0.25 mm i.d., film
thickness: 0.20 μm, SGE) with helium as the carrier
gas. The initial oven temperature was 170ºC for 3
min., thereafter increased by 5 ºC/min. to 200ºC and
maintained for 50 min. and increased by 10ºC/min.
to 220ºC and maintained for 20 min. The identities of
cis-9, trans-11 octadecadienoic acid were established
by comparing retention times to CLA FAME reference
standard (Nu-Chek-Prep). Other fatty acids were not
quantified in this study.
The results were evaluated with Statistica 8
(StatSoft, Poland). The data were tested for normality
with Shapiro-Wilk test. Because of the shortage of
normal distribution the data were examined using
proper non-parametric test, e.g.: Mann-Whitney U
rank test and Kruskal-Wallis test. P – value ≤0.05 was
considered significant.
Results
All examined human milk samples contained
measurable amounts of rumenic acid ranging from 0.36
to 3.99 mg/g of fat (mean±SD; 1.38±0.09 mg/g of
fat). On the milk weight basis, these values ranged from
0.92 to 9.53 mg/100 g of milk (3.17±0.20 mg/100 g
of milk). On the milk volume basis, these values ranged
from 0.84 to 9.71 mg/100 ml of milk (3.24±0.21
mg/100 ml of milk). The fat content in maternal milk
ranged from 1.11 to 5.78% (2.42±0.11%) (Table I).
There was a positive correlation between the fat and
cis-9, trans-11 CLA content in milk (r=0.3589,
p=0.0300) (Fig. 1A), between the rumenic acid
content in fat and in milk (r=0.7173, p < 0.0010)
and a negative correlation between the fat and
rumenic acid content in fat (r=-0.03116, p=0.0100)
(Fig. 1B). These data indicate that the rumenic acid
concentration in human milk depends on both the fat
content in breast milk and the fat quality.
The fat content in milk and the CLA content
in milk fat and in milk depending on the evaluated
factors are listed in Table II.
A high intake of milk and dairy beverages had
a great impact on both the fat and cis-9, trans-11 CLA
concentration in human milk and resulted in a significant
decrease in the fat content (p=0.0016) and a significant
increase in the rumenic acid content in fat (p=0.0043).
Table I. Content of fat and cis-9, trans-11 octadecadienoate in human milk
Tabela I. Zawartość tłuszczu i kwasu cis-9, trans-11 oktadekadienowego
w mleku kobiecym
Range
/Zakres
Mean±SD
/średnia±SD
%
1.11-2.78
2.42±0.11
mg/g of fat
0.36-3.99
1.38±0.09
cis-9, trans-11 CLA
mg/100 ml of milk
0.84-9.71
3.24±0.21
cis-9, trans-11 CLA
mg/100 g of milk
0.92-9.53
3.17±0.20
Variable /Zmienna
Unit /Jednostka
FAT / Tłuszcz
cis-9, trans-11 CLA
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Probl Hig Epidemiol 2011, 92(4): 840-843
Table II. Content of fat and cis-9, trans-11 CLA in human milk depending on various dietary factors
Tabela II. Zawartość tłuszczu i kwasu cis-9, trans-11 oktadekadienowego w mleku kobiecym w zależności od różnych czynników dietetycznych
FAT /Tłuszcz
Factor /Czynnik
Frequency
/Częstotliwość
Milk and dairy beverages Yes /tak
/Mleko i napoje mleczne No /nie
Cheese /Ser
Seldom /rzadko
Often /często
Beef /Wołowina
no/seldom /nie/rzadko
Often /często
Pork /Wieprzowina
Seldom /rzadko
2 a week /2 x w tyg.
3 a week /3 x w tyg.
5 a week /5 x w tyg.
Daily /dziennie
Veal /Cielęcina
Yes /Tak
No /Nie
Poultry /Drób
Seldom /rzadko
2 a week /2 x w tyg.
3 a week /3 x w tyg.
Daily /dziennie
Fish /Ryby
Yes /Tak
No /Nie
Cold meats /wędliny
No /Nie
Seldom /rzadko
2 a week /2 x w tyg.
3 a week /3 x w tyg.
Daily /dziennie
cis-9, trans-11 CLA
%
mean±SD
/średnia±SD
2.21±0.81
2.89±0.98
2.48±1.13
2.41±0.87
2.60±0.98
2.27±1.00
2.62±0.71
2.59±1.18
2.66±0.94
1.98±0.00
2.21±0.77
2.30±0.83
2.57±0.97
2.52±1.00
2.57±1.18
2.55±0.75
2.28±0.82
2.49±1.14
2.39±0.82
3.10±0,00
1.53±0,00
2.41±0,46
2.14±0,72
2.54±1,00
mg/g of fat
p
0.0016*
0.0180*
0.0926
0.1207
0.2588
0.7371
0.2763
0.2119
mean±SD
/średnia±SD
1.51±0.72
1.10±0.63
1.21±0.57
1.42±0.74
1.52±0.78
1.30±0.73
1.46±0.82
1.29±0.59
1.33±0.58
1.06±0.00
1.35±0.90
1.39±0.73
1.43±0.79
1.22±0.58
1.63±0.86
1.39±0.79
1.15±0.47
1.14±0.51
1.48±0.78
0.83±0.00
1.38±0.00
1.04±0.21
1.45±0.68
1.40±0.77
mg/100 ml
p
0.0043*
0.0498*
0.1581
0.8158
0.8260
0.3126
0.5966
0.7022
mean±SD
/średnia±SD
3.23±1.55
3.28±2.05
2.97±1.65
3.31±1.73
3.87±2.09
2.66±1.07
3.95±2.43
3.23±1.74
3.33±1.05
2.12±0.00
2.79±1.60
3.26±1.99
3.52±1.79
2.96±1.28
3.96±1.87
3.47±2.22
2.62±1.21
2.74±1.49
3.40±1.75
2.66±0.00
2.14±0.00
2.92±1.04
2.93±1.27
3.43±1.92
mg/100 g
p
0.7304
0.6781
0.0425*
0.4264
0.5622
0.1428
0.9760
0.8870
mean±SD
/średnia±SD
3.15±1.47
3.21±2.00
2.91±1.56
3.24±1.67
3.82±2.01
2.57±1.02
3.88±2.39
3.15±1.66
3.30±1.03
2.06±0.00
2.75±1.54
3.15±1.91
3.45±1.73
2.89±1.26
3.85±1.72
3.44±2.15
2.56±1.18
2.70±1.42
3.33±1.71
2.56±0.00
2.19±0.00
2.76±1.07
2.84±1.15
3.38±1.85
p
0.6901
0.6781
0.0164*
0.4430
0.4596
0.1225
0.0273*
0.8630
p – probability level/significance level, mean – arithmetic mean, SD – standard deviation, * – significant differences between groups
Frequent consumption of cheese also influenced both
these variables causing significant differences between
groups. The fat content was significantly lower and
the rumenic acid content was significantly higher in
milk of women who often consumed cheese compared
to the group of mothers who seldom ate cheese. Our
results are partly similar to those of Park et al. [13] who
observed a higher concentration of rumenic acid in milk
of women with a higher dietary rumenic acid intake.
Also Rizenthaler et al. [14] showed that consumption
of rumenic acid-enriched cheese slightly increased the
milk rumenic acid concentration. Both these data do
not confirm our results of the influence of dairy product
consumption on the fat amount in milk. The results
of Rizenthaler et al. show that a higher intake of dairy
products rich in cis-9, trans-11 CLA does not influence
the milk fat concentration [14]. Data of Park et al.
indicate that consumption of dairy products causes an
increase in the total milk fat content [13]. Moutsioulis et
al., who investigated the short-term kinetics of maternal
CLA consumption from food on CLA appearance in
breast milk of lactating women, gave the volunteers
CLA-enriched cookies made of CLA-enriched butter
and observed a significant increase in the mean breast
milk CLA concentration (0.16±0.08 vs 0.09±0.04
mg/ml) [15]. Although the mean content of rumenic
acid in milk was noticeably greater than in our study, the
observed tendency confirms our remarks of the influence
of maternal diet rich in dairy food on the rumenic acid
concentration in breast milk. Dairy products are also
a valuable source not only of calcium but also of iodide,
which is essential to fetal development [16].
Surprisingly our data also show that cis-9, trans-11
octadecadienoate consumption was lower in the group
of frequent beef consumption. The increased intake
of pork, veal and poultry did not cause any significant
changes in the fat and CLA concentration. Ruminant
meat like beef or veal are good dietary sources of
conjugated linoleic acids and their intake should
have caused higher CLA amounts in milk [17], but
in our study differences in beef and veal consumption
between groups were not significant, and this could
be the main reason of our surprising findings. Also
a frequent fish consumption influenced the rumenic
acid concentration in maternal milk, causing
a significant decrease of its amount (p=0.0273).
There were also no significant differences in the RA
or fat content in human milk among the groups of
different frequency of cold meats consumption.
Białek A et al. The impact of selected nutritional factors on cis-9, trans-11 octadecadienoate content in milk ... Part I.
Conclusions
1. The diet of breastfeeding mothers and their
nutritional habits have a great influence on the
content of both fat and rumenic acid in their
milk.
843
2. Milk and dairy products are a valuable source of
many important nutrients in the diet of pregnant
and breastfeeding women, e.g. rumenic acid,
which is significant for infant health.
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