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JOURNAL OF SCIENCE, Hue University, N
0
61, 2010


MILK FORTIFICATION WITH IRON ETHYLENE DIAMINE
TETRAACETATE (NAFEEDTA) SUPPOSED TO BE A PREVENTIVE
MEASURE AGAINST IRON DEFICIENCY FOR PRIMARY SCHOOL
CHILDREN IN THAILAND
Panomai N, Sanchaisuriya

P, Lowirakorn
Department of Nutrition, Faculty of Public Health, University of Khonkaen,Thailand
Schelp

FP

Institute of Tropical Medicine, Charite, University Medicine Berlin, Germany
SUMMARY
One cost-effective strategy for controlling iron deficiency anemia is the fortification of
food. Iron salt fortificants such as NaFeEDTA have the potential to be used widely as an iron
fortificant . Milk could be a potentially

useful food vehicle for iron fortification programs
because of the daily free school milk program in Thailand. Objectives: To investigate the
feasibility of iron fortified milk (NaFeEDTA) with a 5 mg iron/sachet/day given to a group of
primary school children. Design: One hundred and sixty school aged children (86 males and 74
females) were enrolled and divided into 2 groups: Group 1 received fortified milk (FM) and

group 2) received non fortified milk (NF). A sachet of milk was served daily for 5 days per week
as 200 ml milk containing 5 mg Fe as NaFeEDTA (FM) or not fortified milk (NF) for 3 months.
Concentrations of hemoglobin(Hb), serum ferritin (SF), mean corpuscular volume (MCV), and
mean corpuscular hemoglobin (MCH), were measured at baseline and after 3 months to
determine the developments. Results: There were statistical differences in the changes of
hemoglobin concentration (Hb), mean cell volumes (MCV), and mean corpuscular hemoglobin
(MCH), between the fortified milk and non fortified milk group at a significance level of p<0.05.
Conclusions: Fortification of milk in connection with a free school milk program is feasible and
might be used as a preventive measure against iron depletion in areas with a high rate of iron
deficiency.
Key Words: Iron, food fortification, iron deficiency anemia, school aged children

1. Introduction
Among micronutrient deficiencies, iron deficiency anemia (IDA) is an important
public health issue. According to the World Health Organization (WHO), worldwide,
about two billion people are anemic, with the main cause being a deficiency in dietary
iron. The progress in controlling IDA over the last few decades, however, is insufficient.


326
This could be due to the fact that in the past, attention had been drawn mainly to iron
deficiency anemia estimated on the grounds of the prevalence of hemoglobin
concentrations below thresholds, and not to the iron status of population groups as such.
In one recent publication, the WHO stated that ‘not all anemic people are iron deficient
and iron deficiency may occur without anemia’. A meaningful public health policy
geared towards prevention of IDA therefore might include investigation of the
importance of other causes of anemia in a given population. As far as iron deficiency is
concerned, implementing measures to improve the iron status of population groups and,
as early as possible in the life time of people at risk, i.e. for school children could
decrease IDA. Thailand is one of the countries facing the problem of IDA. The Ministry

of Public Health (MOPH) of the Thai government indicated that the prevalence of IDA
increased from 20.5 % in the year 1995 to 26.7 % in 2003. From a National Food and
Nutrition Survey conducted in Thailand around the year 2000 it is known that the
prevalence of anemia in school-age children (6-14 years of age) amounted to 13.1 % by
using hematocrit threshold values as criteria and 26.7 % when using hemoglobin values
as criteria. One way to prevent iron deficiency in school children might be iron
fortification. Since 1992 the Royal Thai government has maintained a free milk
program (200ml/child/day) to preschool children in the communities and public primary
schools
.
This program was used as a vehicle for a fortification trial by the investigators.
It is known that iron salts such as ferrous sulphate and fumarate are reasonably well
absorbed and are suitable, when storage is not expected to be long term. Compared with
the commonly used iron salt fortificants, sodium iron ethylene diamine tetraacetate
(NaFeEDTA) has the potential to be used widely as an iron fortificant in future.
NaFeEDTA absorption is 2 to 3 times higher than ferrous sulphate. Its structure affords
some protection against phytate inhibition of iron absorption. It is about one third as
well absorbed as ferrous sulphate, but has the advantages of greater stability and of
increasing the absorption of native non-haem iron and zinc in food
.
Based on these
fortunate preconditions, a fortification trial program using NaFeEDTA fortified milk
was conducted. The researchers aimed to undertake an assessment of the feasibility of
such a program, as well as to try to establish the effectiveness of the program. In
addition a short qualitative dietary survey was undertaken to estimate the frequency of
food items known to be either promoting or inhibiting iron absorption.
2. Methodology
A district approximately 660 km northeast of Bangkok was selected as a study
area. Four schools at the Si Chomphu district, KhonKaen Province; namely the Si
Chomphu Kindergarten, Ban Mai Sok Som Kob, Ban Nong Ta Kai and Choom

Chon Ban Wang Perm School participated in the project, which was conducted from
November 2007 to March 2008. The Si Chomphu district was chosen for the study
because of pronounced interest in health - and the school authorities of the district


327
facilitated cooperation with study team. The selection of schools was based on a simple
random sampling technique from schools with not less than 120 and not more than
1,500 school children. From the selected schools, out of 1.158 children in the age of 9 to
11 years 160 school children (86 boys and 74 girls) were randomly selected and asked
to join the project, and 80 children for each group were allocated either to the group
receiving fortified milk (FM) or getting non-fortified milk only (NF). Children
participating in the study had no history and symptoms of a serious disease, in particular,
they were not suffering from thalassemia or hematolytical disease, Hb concentration of
>8 g/dl and serum ferritin of 12 g/l. Permission to conduct the intervention was
received from local education authorities and school principals. Informed written
consent was obtained from the parent or guardian of each child who accompanied the
children on the first day of the survey. De-worming by 15 mg albendazole was provided
for all of the students for the elimination of intestinal parasites before starting the
intervention. Based on the study protocol, 5 mg of iron in the form of NaFeEDTA milk
(200 ml per sachet) was administered daily to each subject of the fortified milk group
(FM) at 8.30 AM for a period of 3 months. The intervention group as well as the control
group were given milk of the same brand and quantity. The quantity of NaFeEDTA were
given to the students following the Joint Expert Committee on Food Additives (JECFA)
recommendations
.
Both the fortified and the non fortified milk were provided by a milk
company located within the Khon Kaen province. The concentration of iron added to
the fortified milk was 5 mg of iron per 200 ml of milk as NaFeEDTA. Food grade-
NaFeEDTA was manufactured and provided by AkzoNobel Chemicals Pte. Ltd.,

Singapore. The milk was distributed to each child 5 days/week by the teacher, or a staff
member of the project. In order to ensure accurate intake, the milk was consumed by the
pupils under the complete supervision of the investigator and teachers. At the same time,
detailed information about the milk consumption of each child was recorded on a
consumption sheet to check the acceptance of the milk by the students. A short
qualitative dietary assessment about food items promoting and inhibiting iron
absorption was undertaken at the beginning of the study.
A 3 ml venous blood sample was taken at the beginning and at the end of the
study period for the measurement

of hemoglobin concentration (Hb), serum ferritin (SF),
Mean Cell Volume (MCV), and Mean Corpuscular Hemoglobin (MCH). Each sample
was inserted into EDTA-coated tubes. The tubes were kept cool and transported to the
laboratory at the Research Center, Faculty of Associated. Medical Science, and Faculty
of Medicine, Khon Kaen University. All of bloods samples were analyzed to determine
iron storage (serum ferritin) which was determined by COBAS INTEGRA
®
800
analyzer, and red blood cell indices (Hb, serum ferritin(SF), MCV, and MCH) by the
Sysmex
®
SF-3000 automated hematology analyzer, Kobe, Japan.
Statistical analysis was done by using the STATA program. Percent mean,


328
standard deviation, and median to describe general information were calculated.
Students’t-test and Wilcoxon sign rank test were applied to examine the difference
changes of hemoglobin (Hb), serum ferritin (SF), mean cell volume (MCV), and mean
corpuscular hemoglobin (MCH) over the study period for 3 months.

3. Results
The age of the students was 9.9 + 0.3 and 9.8+0.4 years old for males and
females, respectively. The mean of weight for males was 27.7+5.5 kilograms and for
females 29.7+8.8 kilograms. The height of males was 129.6 +7.8.5 cm and of females
133.4+9.2 cm. The results derived from boys and girls had been quite similar and no
interesting aspect had been discovered by evaluating sexes separately so that in the
following the results are given for both sexes combined except for table 2.
Part of the results of a qualitative dietary survey is given in Table 1. Frequent
intake of protein supporting iron absorption is mainly restricted to poultry and plants
while intake of food known to somehow inhibit iron absorption is mainly due to the
digestion of egg yolk and soybean products.
Table 1. Number and percent of food items being enhancers or inhibitors of iron absorption
consumed 1 to 2 times /week by 160 school children
Food items being
enhancer of iron
absorption*
N (%)
Food items being
inhibitor of iron
absorption**
N (%)
Beef, meat 23 (14.4) Seasame seeds, black 5 (3.1)
Chicken, drumstick 37 (23.1) Seasame seeds, white 13 (8.1)
Chicken, thigh 27 (16.9) Tea 17 (10.6)
Chicken, wing 27 (16.9) Coffee 11 (6.9)
Chicken, liver 25 (15.6) Egg (hen, yolk) 28 (17.5)
Leadtree, leaves 22 (13.7) Egg (duck, yolk) 19 (11.9)
Onion 30 (18.8) Soybean paste, black 16 (10.0)
Ya-nang leaves*** 20 (12.5) Soybean paste, white 7 (4.4)
Cassia, leaves 30 (18.8)

Tomato, ripe 24 (15.0)
Fennel, leaves 24 (15.0)
Sesbania, leaves 15 (9.4)
Tamarind, young 26 (16.2)


329
Based on a qualitative food frequency questionnaire of 160 children. Multiple
answers were possible
*Enhancer: Food items rich in heme, ascorbic acid, citric acid, or vitamin A
** Inhibitors: Food items rich in phutate, polyphenols (tannin), soy protein, or
egg yolk
***Antiaris toxicaria
Before starting the intervention children had been asked how they tolerated
drinking milk in the past. The results are shown in Table 2. Most of the males did not
complain about side effects after drinking the milk; 28 children (65.1 %) admitted that
drinking the milk did not stimulate their appetite, 11 children (25.6 %) complained
about flatulence, 5 children vomited (11.6 %) and 4 (9.3 %) boys complained about
having diarrhea. Milk did not stimulate appetite for 22 girls, (59.5 %), flatulence and
vomiting and diarrhea was rare for the group of girls as well.
Table 2. General health, side effects evaluation for boys and girls.
Response Factor
Rating Scale
Male(n=43) Female(n=37)
YES NO YES NO
n (%) n (%) n (%) n (%)
Appetite 15(34.9) 28(65.1) 15(40.5) 22(59.5)
Flatulence 11(25.6) 32(74.4) 3(8.1) 34(91.9)
Vomiting 5(11.6) 38(88.4) 3(8.1) 34(91.9)
Diarrhea 4(9.3) 39(90.7) 1(2.7) 36(97.3)

Table 3 presents the hemoglobin (Hb), mean cell volume (MCV), mean
corpuscular hemoglobin (MCH), and serum ferritin (SF) values before and after 3
months of intervention and the differences before and after the study as means and 95%
CI. The p-value indicates the statistical significance between the means of the
differences between the group of children receiving fortified milk and the controls.
Mean values of Hb for the control group decreased and for the intervention group
increased. The difference before and after intervention for both groups is statistically
significant. The variation of ferritin was not altered by the intervention. Furthermore it
also shows the changes of red blood cell indices after 3 months of the study. The results
indicate that the intervention statistically altered the difference between before and after
the provision of fortified milk of MCV and MCH for the FM group above the NF group.


330
Table 3. Hemoglobin(Hb), mean cell volume(MCV), mean corpuscular hemoglobin, (MCH),
and serum ferritin(SF) changes in school aged children after 3 months of intervention according
to groups.
Parameters

FM NF
P-value
Before

After
Mean
diff
Before

After
Mean

diff
(95%CI)

Hb
12.2
±1.1
12.2±1.0

-0.0
(-0.1
to -
0.2)
12.6 ±
0.8
12.3±0.9

-0.3
(-0.4
to -0.2)
0.0004*
MCV
74.00
(54.00,

85.00)
76.40
(56.50,
88.40)
2.60
(2.37

to
2.90)
76
(54.00,
87.00)
78.80
(55.40,
89.20)
2.05
(1.78
to 2.40)
0.0002*
MCH
25.00
(18.00,

29.00)
25.25
(18.50,
29.90)
0.50
(0.40
to
0.70)
26.00
(18.00,
31.00)
25.90
(17.90,
31.30)

0.15
(0.00
to 0.30)
0.0001*
Ferritin
53.2
(20.3,
159.9)
51.3
(14.1,
159.9)
0.7
(-4.2
to
7.8)
55.3
(16.5,
172.6)
58.6
(12.4,
186.5)
7.9
(-5.8
to 21.9)
0.08
Data are presented as the mean value + SD. The data were analyzed by the
student t test
*The data were analyzed by the Mann-Whitney U test and presented as the
median value.
*Significance difference at P<0.05.

4. Discussion
The study site is known to be an area with a high prevalence of iron deficiency.
Iron deficiency anemia among school children had been assessed on district level at the
Khon Kaen province. A School Health Report by the Si Chomphu Hospital, Si


331
Chomphu district, Khon Kaen province found that the prevalence of iron deficiency
anemia on the primary school level was as high as 21.3 %. The situation is not reflected
by the school children co-operating in this study, because of ethical reasons anemic
children could not be included into the control group. So only quite healthy children had
been investigated, thus it was not expected that the results will achieve great
improvements in the hematological status of the children in the intervention group.
Further limitations of financial resources and lack of adequate laboratory facilities
prohibited the author from using other indicators of iron status such as transferrin
saturation, serum zinc protoporphyrin or the transferrin receptor
.
Although ferritin
values also reacts to infections,

which might have happened here. It is supposed that at
least the majority of children taking part in this study were prone to iron depletion
which is indicated by the fact that a small but significant improvement of the
hematological status of the children of the intervention group could be observed. This
was indicated by an improvement in the red cell volume (MCV) and red cell
hemoglobin concentration (MCH). Food items supporting but also inhibiting iron
absorption are balanced in the dietary intake as indicated by the results given in table 1.
Other micronutrients besides iron such as vitamin A might contribute to the risk of the
children having insufficient hematological status. Thus supplementing the school milk
program by NaFeEDTA might be well justified.

5. Conclusion
Obviously the milk is well tolerated by the children as the results given in table
2 indicate. Lactose intolerance is known to be a problem in Thailand but obviously is
not a factor important enough to prevent continuation the milk school program since
only a few children report occasional intestinal disturbance when drinking milk. The
amount of iron in food fortification of other studies usually has been higher than that
used in this study. The use of NaFeEDTA-fortified milk in this study was only 5 mg of
iron per person per day which is a quite low dose of NaFeEDTA. Supplementation
however might be sufficient to maintain an optimal iron store in children with iron
depletion. Additional studies however are required to substantiate this. It is supposed
that this also holds true for primary school children, at least when exposed to relatively
low doses of iron supplementation, but help children who are in a pure state of iron
depletion.
In conclusion, the result of this study strongly suggests continuing the
supplementation of the school milk program with NaFeEDTA. However further
investigations of the iron status of school children are recommended.
Acknowledgements
This paper is one part of the results of a Ph.D thesis (Public Health, Graduate
School, Khon Kaen University, Thailand). To finish the project would not have been


332
possible without the invaluable support and assistance of the directors and teachers of 4
government schools; Choomchon Ban Wang Perm, Nong Ta Kai Pittaya, Ban Mai Sok
Som Kob, and Si Chomphu Kindergarten school, Khon Kaen province, Thailand. The
study was supported by AkzoNobel Chemical Company, who supplied a research grant
and NaFeEDTA fortificants.

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