Robyn
L. McKenzie,
Christine
D.
M.H.
Thomson,
Sc.;
Ph.D.,
The
those
reported
determined New
by
Zealand
and
types
diets
had
blood children.
low eaten.
was
Zealand
1413-1418,
intake
to decrease
with
found adults,
of the
with
blood
between peroxidase reflecting
Journal
ofClinical
Sci.,
F. Robinson, of
other
countries.
children
which,
geographically,
Ph.D.,
phenylketonuria
and
on these
blood
Se levels
activities their
of the
lower
blood
and children
Zealand low
maple
the length
the
New
reflects syrup
with diets.
Se
the
urine
A strong
Se concentrations.
low
than
lower
primarily
Se content
of
in quantities
disease
on synthetic
on normal
diets,
correlation
(r
peroxidase lower
were was
differences
children
glutathione were
children
blood
age, and with
compared
of time
M.H.Sc.
This in turn,
with
Se concentrations
activities activities
Am.
and 0.62,
=
107
for
in
observed
J. Clin.
31:
Nutr.
1978.
Many arable areas of New Zealand, particularly in the South Island, have low soil selenium (Se) concentrations (1). As a result pastures and crops are low in Se, and Seresponsive diseases occur in sheep and cattle. New Zealand adults have a lower Se status compared with North American adults as assessed by low dietary intakes (1, 2), blood concentrations (3) and 24-hr urinary outputs of Se(4). Se is an integral part of the enzyme glutathione peroxidase (GSHPx) (5, 6), and GSHPx activity in blood has been correlated with dietary Se of chicks (7) and rats (8), and blood Se concentrations in sheep, cows, and pigs (9). A high correlation was found between Se concentrations and GSHPx activities in the blood of 264 New Zealand residents (10). Lombeck (1 1) reported low serum Se levels and erythrocyte GSHPx activities for healthy German subjects during infancy and early childhood; Se levels and enzyme activities of infants and children with phenylketonuria (PKU) and maple syrup urine disease (MSUD) were lower still, due to very low Se intakes from synthetic protein diets (12, 13). As yet, a human Se deficiency has not been demonstrated. However, it is possible that if The American
Dip.
concentrations in
Se also varied and
Glutathione
(Se) living
Children
B. Sc.,
Marion
selenium
children
dietary
Blood
Se intakes
Se was seen
P < 0.001) New
the
and
blood for
soils.
of food low
M. Rea,
M.H.Sc.;
ABSTRACT than
Heather
Nutrition
3 1 : AUGUST
such a state exists, then it might be found in New Zealand residents. In particular, infants and young children, and children on synthetic protein diets might be “at risk”. This paper reports Se concentrations and GSHPx activities in several groups of infants and children resident in New Zealand. Methods Subjects Random blood samples were collected from infants and children living in five areas of New Zealand: Auckland and Wellington, urban areas of the North Island where the soil Se is greater than 0.5 g g; Christchurch and Dunedin, urban areas of the South Island where the soil Se is less than 0.5 tg g’ and Tapanui, a rural area near Dunedin where Se responsive diseases occur in livestock and the soil Se is less than 0.5 zg g’. Blood samples were also collected from healthy adults living in Auckland, Dunedin, and Tapanui. The subjects were divided into the following groups (number of subjects in parenthesis): 1) healthy infants under 1 year of age and children living in Auckland (one infant, 14 children), I From the Otago, Dunedin, 2 Supported
Department of Nutrition, New Zealand. by the Medical Research
University Council
Zealand and the Medical Research Distribution mittee. 3 Address reprint requests to: Heather M. partment of Nutrition, University of Otago, Dunedin, New Zealand.
1978,
pp.
1413-14
18. Printed
in U.S.A.
of
of New ComRca, Box
Dc56,
1413
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
Selenium concentration and glutathione peroxidase activity in blood of New Zealand infants and children13
1414
MCKENZIE
ET
AL.
ground to homogeneity were stored in a freezer. Analytical
where
necessary.
Liquid
foods
methods
Se in blood and synthetic foods was determined by a modification (15) of the diaminonaphthalene fluoriinetric method of Watkinson (16). Reproducibility of the method and recoveries of added Se were good (3). GSHPx activities were assayed in whole blood of 107 of the subjects (including six PKU children) by a modification (10) of the coupled enzyme assay of Paglia and Valentine (17) using t-butyl hydroperoxide as substrate.
Diet
Results
Dietary information relevant to Se intakes was collected for most subjects. Diets for children with MSUD and PKU were classified into: 1) diets containing synthetic foods (S diets); 2) restricted diets which excluded meat, fish, eggs, and poultry (R diets); 3) unrestricted or normal diets (U diets). Samples of the synthetic foods consumed by the. MSUD infant and by the infants and some of the children with PKU were obtained from Auckland hospital for analysis. Daily Se intakes of all subjects on S diets were estimated from 24-hr dietary recall using values for the Se content of synthetic foods as well as values for foods analysed in our department (14; G. M. Friend, unpublished data). Collection
and
storage
Table 1 gives mean blood Se concentrations for all subjects in each group as well as for those in which enzyme activities were estimated. Values have been included for healthy adults from Auckland, Dunedin, and Tapanui (18). Comparisons of blood Se values were made for the complete groups. There was little difference between the means for the complete group and for the smaller group for which enzyme activities had been measured. There was no difference in blood Se concentrations or in GSHPx activities between male and female subjects. Plasma Se was always less than erythrocyte Se, and these concentrations were reflected in that of whole blood. For 36 children from Tapanui, plasma
of samples
Samples of whole blood were collected into heparinized vacutainers (Becton-Dickinson and Co., Rutherford, N.J.), separated into plasma and erythrocytes where possible and stored below 4 C. The solid foods were dried to constant weight under an infrared lamp and
TABLE 1 Se concentrations of children and New Zealand”
and adults
GSHPx activities living in
Area
in whole
Age
blood
No.
Se concentration
yr
Healthy children and Auckland children
ig/m1
7 ± 3 7 ± 3 36 ± 4
15
Tapanui
children Adults
11 ± 2 35 ± 13
Hospitalized Wellington
children 7
± ± ±
±
± 0.0 17 ± 0.012 ± 0.012
18 59 12”
0.059 0.062 0.068
±
50 49
0.048 0.060
± 0.010 ± 0.012
15
0.063 0.059
±
0.043 0.042
± 0.010 ± 0.012
7.3
±
2.8
0.049 0.042
±
0.010 0.006
10.1
±
3.3
122
9 33 25
3 12 3
3
b
7 ± 2 Christchurch
Dunedin
Values
are
means
un#{252}s/gHb
0.064 0.060 0.083
13b
children Adults
a
GSHPx . activities
adults
Adults Dunedin
.
± SD
b
7 ± 4 3 ± 3
26
5 ± 3 3 ± 2
39
Subjects
10b
5b
for whi ch enzyme
activi
ties
assayed.
± ±
0.01 1 0.012 0.009
0.021 ± 0.012
±
10.6 ± 3.2 12.9 ± 3.1 10.6
±
2.2
10.9
±
2.5
9.3 ± 2.5
10.3 ± 3.3
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
Dunedin (no infants, 19 children) and Tapanui (no infants, 50 children); 2) hospitalized infants and children in Wellington (one infant, 14 children), Christchurch (three infants, 23 children) and Dunedin (four infants, 35 children); 3) infants and children with PKU hiving in Auckland (three infants, 11 children) and Christchurch (one infant, three children) and one Auckland infant with MSUD. Second blood samples were collected from four children with PKU 5 to 8 months after the first bloods were taken; 4) healthy adult blood donors living in Auckland (122 subjects), Dunedin (59 subjects), and Tapanui (49 subjects.)
BLOOD
SELENIUM
CONCENTRATIONS TABLE Blood and
CHILDREN
2 Se concentrations
1415
of infants
children ,,
Country (reference)
Age group
Venezuela (20) Guatemala (21) Thailand (22) USA (23) West Germany (12)
children” 3-8 yrb 1-4 yrb Infantsb 1 mo-l yre 1-5 yrc 5_l2yrc S mo 1-5 yr 5-10 yr 1 mo-l yrb I_5 yr”
New
Zealand
9 7 16 9 10 7 1 6 24 8 24 29
5-lOyr” a Living in seleniferous Serum Se concentrations.
area.
b
0.8 13 0.23 ± 0.05 0.120 ± 0.036 0.097 0.051 0.081 0.097 0.036 0.057 ± 0.015 0.054 ± 0.009 0.035 ± 0.006 0.048 ± 0.012 0.057±0.017
Hospitalized.
18
16
.
14
12
D
.
19
t
o
I
0
102
004 Se concentrobon.
I
I
006
DIR
I
Ill
pg/mt
FIG. 1. Se concentrations and GSHPX activities in whole blood of New Zealand children (ages 1 month to 16 years); healthy and hospitalized children (#{149}), children with PKU (0).
differences among mean GSHPx activities of the different groups with the exception of the Christchurch hospitalized group that was lower than all the healthy groups of children (Auckland, P < 0.02; Dunedin, P < 0.002; Tapanui, P < 0.05). The mean GSHPx activity for the Auckland children was lower than that of 122 Auckland adults (P < 0.02). However, there were no differences in GSHPx activities between the healthy or hospitalized Dunedin children and a group of 12 Dunedin young adults, whose mean age was 25 ± 3 years (19).
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
Se, and erythrocyte Se was 0.036 ± 0.007 and 0.061 ± 0.010 tg Se/mi, respectively, and whole blood Se was 0.047 ± 0.007 zg Se/ml, which was almost identical with the mean for all the Tapanui children. The mean hematocrit for these children was 0.39 ± 0.02. Such minor differences in hematocrit could not account for differences in whole blood Se concentrations. For the healthy groups, the mean blood Se concentration of the Tapanui children was lower than those of the Auckland and Dunedin groups (P < 0.01). For the hospitalized children the mean blood Se concentration of the Wellington group was higher than those of the Christchurch and Dunedin groups (P < 0.01) while that of the Christchurch group was lower than that of the Dunedin group (P < 0.02). Dunedin was the only area in which blood Se concentrations were determined in both healthy and hospitalized infants and children; the healthy group had a higher mean blood Se concentration than the group in hospital (P < 0.001). Overall, the mean Se concentration of all the healthy subjects (0.053 ± 0.0 13 jzg Se/ml) was similar to that of hospitalized subjects (0.050 ± 0.014 sg Se/ml). However, the mean of the total healthy group was heavily weighted by the lower values for the Tapanui children. Blood Se of the one healthy infant from Auckland (0.034 jsg/ml) and of the eight hospitalized infants from Wellington, Christchurch, and Dunedin all fell within the range of 0.021 to 0.042 .tg/ml. The mean for infants’ blood, 0.035 ± 0.006 .tg/ml was less than means for children in age groups of 1 to 5 and 5 to 10 years old (Table 2). There was a tendency for blood Se levels to increase with age in the healthy Dunedin group but not in any other group. The mean blood Se concentration of Dunedin children was a little lower than that recorded for 59 Dunedin adults (0.062 ± 0.012 sg Se/ml); the mean for the Auckland infants and children was lower than that for 122 Auckland adults (0.083 ± 0.012 Se/mi, P < 0.001) and the mean for the Tapanui children was lower than that for 49 Tapanui adults (0.060 ± 0.012, P < 0.001) (18). Figure 1 shows a strong correlation between the blood Se levels and GSHPx activities for 107 infants and children (r = 0.62; P < 0.001). However, there were no significant
OF
1416
MCKENZIE
Infants
and
children
with
MSUD
and
PKU
TABLE 3 Se content of synthetic
foods
Branch chain free amino acid mixture (jg/g) Lofenalac powder (jzg/g) Aminogram food supplement (jsg/g) Aminogram mineral mixture (jg/g) Low phenylalanine bread (jtg/g) Prosparol (jzg/ml) Ketovite tablets (j.tg/tablet)
0.059 0.030 0.010 0.005 0.002 0.002 0.002
O8
.
#{149}
S
o7 O16 S
E
005 004
0#{149}03 002
cb 0#{149}01
0
2
I
I
I
I
I
I
4
6
8
10
12
14
age.yr FIG. 2. Se concentrations in whole blood of infants and children with MSUD and PKU; PKU synthetic diet (0), PKU nonsynthetic diet (#{149}), MSUD synthetic diet (L5).
AL.
which was almost the length of time that they had been on these diets. Blood Se concentration ofthree ofthe PKU children, from whom a second sample was taken, had decreased by 0.006 to 0.012 j.tg Se/mi: the fourth child showed no significant change in blood Se, which remained very low at 0.016 ig Se/mi. GSHPx activities were assayed for six PKU children living in Auckland, five on S diets. The mean GSHPx activity for these children was 8.9 ± 3.3 units per gram of hemoglobin, lower than that for 13 healthy Auckiand children. Discussion It has been well established that the blood Se concentrations of New Zealand adults are lower than those of adults living in other countries (3) and that New Zealand produced foods contain iess Se than most overseas foods (1, 14). The present study indicates that the blood Se of New Zealand children was also low compared with that of children in many other countries (Table 2). Estimations were performed by fluorimetry (with diaminonaphthalene) in all studies, except for that of the West German chiidren (12) in which serum Se only was estimated by neutron activation analysis. Their values for serum Se were much higher than those for plasma Se of Tapanui children. The major dietary sources of Se for man other than fish are cereals and animal products and the Se content of these foods is determined largely by the soil Se concentrations in the areas where they are produced (1); Se content of liver and kidney in low Se areas can also vary with the time animals are slaughtered after drenching with Se. Dietary Se intakes are, therefore, likely to vary geographically and this was reflected in differences in blood Se concentrations of children living in different areas of the country. Geographical variations had also been observed in blood Se within the New Zealand adult population (3). Serum Se concentration determined in some healthy West German infants and children increased from a median value of 0.051 I.Lg Se/mi, for nine infants ages 1 month to 1 year, to 0.097 jig Se/mi for seven children ages 5 to 15 years (12). Whole blood Se of New Zealand infants was also lower than the
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
Se contents of the synthetic foods are presented in Table 3. Low phenylalanine bread had a lower Se content than normal white bread (0.015 jsg Se/g dry weight) but no other comparisons could be drawn between the synthetic foods and normal infant foods. The mean daily intake for the 12 subjects on S diets was 5 ± 2 zg Se day’, with little difference between the older children and the infants. Figure 2 shows the blood Se concentrations of infants and children with PKU and MSUD. The mean blood Se concentration of those on S diets (0.038 ± 0.013 tg Se/ml) was lower than the blood Se of six subjects on R and U diets (0.074 ± 0.009 g Se/mi, P < 0.001). The blood Se concentrations of subjects on S diets appeared to decrease with age
ET
BLOOD
SELENIUM
CONCENTRATIONS
CHILDREN
1417
being on an intake of less than 5 .tg Se day’ for 13 years. This study then indicated that the low blood Se of New Zealand children is determined primarily by low dietary Se intake, and this was reflected in lowered GSHPx activities. Lombeck et al. (13) reported lower GSHPx activities for 19 PKU children. In the present study the mean GSHPx activity for six PKU children was lower than that for heaithy children but this was not significant for the small number of subjects studied. The strong correlation between blood Se levels and GSHPx activities for the infants and children confirms the previously established correlation for 264 New Zealand subjects ages between 2 and 92 years (10), and shows the same kind of regression as for values less than 0. 1 .tg Se/mi whoie blood. Se concentrations of 0.016 tg Se/mi whole blood and 0.009 ig Se per milliliter of plasma found for subject Q.R. were the lowest in this study and lower than most values recorded in the literature. A clinical examination of subject Q.R. indicated that he was in good health. Low concentrations for serum Se, 0.002 to 0.008 .tg Se/mi, were aiso recorded for infants and children on special PKU and MSUD diets in West Germany ( 12). These subjects were reported to be “in good clinical state and thriving well”. Although vitamin E intake of these children on synthetic diets and those in the present study was not determined, it is unlikely that it was inadequate and may protect against the effects of low Se intake. However, Gross (26) has observed that hemolytic anemia can still occur in premature infants on milk formulas even in the presence of adequate serum tocopherol levels. He found that feeding infants a low Se milk formula resulted in decreasing blood Se concentrations and GSHPx activities. But it was the additional oxidant stress of high polyunsaturated fatty acid content or added iron which resulted in reduced hemoglobin, increased reticulocyte response and hydrogen peroxide fragility values and also peroxidation-induced membrane damage as indicated by decreased erythrocyte phosphatidyl ethanolamine. This requires further investigation in some of our New Zealand infants. U and
The authors are Miss Veronica
very grateful Doesburg
to Miss for their
Gaylene technical
Friend assist-
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
children but a tendency to increase with age above 1 year was seen oniy in the Dunedin healthy children. Furthermore, mean blood Se concentrations of groups of children in different parts of the country were lower than values recorded for adults in corresponding areas (Table 1). These variations with age possibly reflect differences in the quantities and types of foods eaten. Griffiths (2) found the range of daily Se intakes of some New Zealand adults living in Dunedin was 6 - 70 jig Se daf’ with intakes greater than 35 tg Se day’ occurring only on days when Se-rich foods, such as fish, liver, and kidney were eaten. Using our values for the Se content of foods (14; G. M. Friend, unpublished data) and 24-hr dietary recali data collected for healthy New Zeaiand infants ages 2 months to 5 years (24; F. Davidson, personai communication) an estimate of a daily intake of 5 to 8 .tg Se day’ was obtained. It appears that the dietary Se intake of New Zealand residents might increase from about 5 jg Se day’ during infancy up to 20 to 30 jsg day’ during adolescence and adult life. Elderly subjects (over 70 years) had lower blood Se concentrations than younger adults (19) and this might also reflect the reduced food intake of this age group. The lower mean blood Se concentration of the hospitalized Dunedin group, as compared with the healthy group, might have resulted from a lower food intake during ill health. However, tissue wasting during illness at a time when growth is important could also have influenced blood Se levels. Infants and children with MSUD and PKU are fed restricted diets containing synthetic proteins for at least the first 3 years of life and often for longer periods. Since Se is mainly associated with the protein fraction of food (25), it was not surprising that the Se intake was very low for subjects on synthetic diets and remained low for the older children. The blood Se concentrations of the older chiidren were much lower than those on restricted or unrestricted diets (Fig. 2). This trend was further illustrated by the decreased Se concentration of the second blood samples from three PKU children (Fig. 2). A fourth child, subject Q.R. 13 years old showed little change. This might indicate that the blood Se had stabilized around 0.016 tg Se/mi after
OF
1418
MCKENZIE
13.
14. 15.
References 16. 1. 2.
3.
4.
5.
6.
7.
J. H. The selenium status of New Zealanders. N. Z. Med. J. 80: 202, 1974. GRIFFITH5, N. M. Dietary intake and urinary excretion ofselenium in some New Zealand women. Proc. Univ. Otago Med. Sch. 51: 8, 1973. GRIFFITHS, N. M., AND C. D. THoMsoN. Selenium in whole blood of New Zealand residents. N. Z. Med. J. 80: 199, 1974. THOMSON, C. D. Urinary excretion of selenium in some New Zealand women. Proc. Univ. Otago Med. Sch. 50: 31, 1972. ROTRUCK, J. T., A. L. POPE, H. E. GANTHER, D. G. HAFEMAN AND W. G. HOEKSTRA. Selenium: biochemical role as a component of glutathione peroxidase. Science 179: 588, 1973. FLOHE, L., W. A. GUNZLER AND H. H. SCHOCK. Glutathione peroxidase: a selenoenzyme. Febs. Lett. 32: 132, 1973. NooucHl, T., A. H. CANTOR AND M. L. Scorr. Mode of action of selenium and vitamin E in prevention of exudative diathesis in chicks. J. Nutr. 103: WATKINSON,
1502, 8.
17.
18.
19.
,
).
21.
1973.
D. G., R. A. SUNDE AND W. G. HOEKEffect of dietary selenium on erythrocyte and liver glutathione peroxidase in the rat. J. Nutr. 104: 580, 1974. 9. THOMPSON, R. H., 0. H. MCMURRAY AND W. J. BLANCHFLOWER. The levels of selenium and glutathione peroxidase activity in blood of sheep, cows and pigs. Res. Vet. Sci. 20: 229, 1976. 10. THOMSON, C. D., H. M. REA, V. M. DOESBURG AND M. F. RoBINSoN. Selenium concentrations and glutathione peroxidase activities in whole blood of New Zealand residents. Brit. J. Nutr. 37: 457, 1977. 1 1. LOMBECK, I., K. KASPEREK, H. D. HARBISCH, L. E. FEINENDEGEN AND H. J. BREMER. The selenium state of healthy children. Europ. J. Pediat. 125: 81, 1977. 12. LOMBECK, I., K. KASPEREK, L. E. FEINENDEGEN AND H. J. BREMER. Serum-selenium concentrations in patients with maple-syrup-urine disease and HAFEMAN,
STRA.
22.
23.
24.
25.
26.
AL. phenylketonuria under dietotherapy. Chin. Chim. Acta 64: 57, 1975. LOMBECK, I., H. D. HARBISCH, K. KASPEREK, L. E. FEINENDEGEN AND H. J. BREMER. Decreased serum selenium concentrations and reduced glutathione peroxidase activity in dietetically treated patients with metabolic diseases. Chem. Scripta 8A: 109, 1975. RoBINSoN, M. F. The moonstone. J. Human Nutr. 30: 79, 1976. THOMSON, C. D. The metabolism and nutritional importance of selenium in the human population of New Zealand. University of Otago: Ph.D. Thesis, 1973, p. 27. WATKINSON, J. H. Fluorimetric determination of selenium in biological material with 2,3-diaminonaphthalene. Anal. Chem. 38: 92, 1966. PAGLIA, D. E., AND W. N. VALENTINE. Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J. Lab. Clin. Med. 70: 158, 1967. RoBINSoN, M. F., C. D. THOMSON, R. D. H. STEWART. H. M. REA AND R. L. MCKENZIE. Selenium in human nutrition in New Zealand residents. In: Proceedings of Third International Symposium on Trace Element Metabolism in Man and Animals, edited by M. Kirchgessner. TEMA 3, 1978, in press. THOMSON, C. D., H. M. REA, M. F. RoBINSoN AND 0. W. CHAPMAN. Low blood selenium concentrations and glutathione peroxidase activities in elderly people. Proc. Univ. Otago Med. Sch. 55: 18, 1977. JAFFE, W. G., D. M. RUPHAEL, M. C. MONDRAGON AND M. A. CUEVAS. Estudio clinico y bioquimico n ninos escloares de una zona selenifera. Arch. Latinoamer. Nutr. 22: 595, 1972. BURK, R. F., W. N. PEARSON, R. P. WooD AND F. VITERI. Blood selenium levels and in vitro red blood cell uptake of75Se in kwashiorkor. Am. J. Chin. Nutr. 20: 723, 1967. LEVINE, R. J., AND R. E. OLSON. Blood selenium in Thai children with protein-calorie malnutrition. Proc. Soc. Exptl. Biol. Med. 134: 1030, 1970. RHEAD, W. J., E. E. CARY, W. H. ALLAWAY, S. L. SALTZSTEIN AND G. N. SCHnAUZER. The vitamin E and selenium status of infants and the sudden infant death syndrome. Bioinorg. Chem. 1: 289, 1972. HARDING, W., A. VAN Ru AND F. DAVIDSON. Diets ofNew Zealand pre-schcol children. J. N. Z. Dietet. Assoc. 29: 17, 1975. BURK, R. F. Selenium in man. In: Trace Elements in Human Health and Disease, edited by AS. Prasad. New York: Academic Press 1976, vol. II, p. 105. GROSS, S. Hemolytic anemia in premature infants: Relationships to vitamin E, selenium, glutathione peroxidase and erythrocyte lipids. Seminars Hematol. 13: 187, 1976.
Downloaded from https://academic.oup.com/ajcn/article-abstract/31/8/1413/4655909 by University of Rhode Island user on 16 October 2018
ance, to the following people for collecting blood samples from the children: Professor D. Lines, Department of Paediatrics, Auckland Hospital; Dr I. C. T. Lyon, Dcpartment of Community Health, Auckland University Medical School; Professor H. J. Weston, Department of Paediatrics and Child Health, Wellington Hospital; Professor F. T. Shannon and Dr R. Ford, Department of Paediatrics, Christchurch Hospital; Professor J. M. Watt, Department of Paediatrics, Otago University Medical School; and Dr P. G. Snow, Tapanui; and Miss P. King, Dietary Department, Auckland Hospital, for collecting samples of the synthetic foods.
ET
L. McKenzie,
Christine
D.
M.H.
Thomson,
Sc.;
Ph.D.,
The
those
reported
determined New
by
Zealand
and
types
diets
had
blood children.
low eaten.
was
Zealand
1413-1418,
intake
to decrease
with
found adults,
of the
with
blood
between peroxidase reflecting
Journal
ofClinical
Sci.,
F. Robinson, of
other
countries.
children
which,
geographically,
Ph.D.,
phenylketonuria
and
on these
blood
Se levels
activities their
of the
lower
blood
and children
Zealand low
maple
the length
the
New
reflects syrup
with diets.
Se
the
urine
A strong
Se concentrations.
low
than
lower
primarily
Se content
of
in quantities
disease
on synthetic
on normal
diets,
correlation
(r
peroxidase lower
were was
differences
children
glutathione were
children
blood
age, and with
compared
of time
M.H.Sc.
This in turn,
with
Se concentrations
activities activities
Am.
and 0.62,
=
107
for
in
observed
J. Clin.
31:
Nutr.
1978.
Many arable areas of New Zealand, particularly in the South Island, have low soil selenium (Se) concentrations (1). As a result pastures and crops are low in Se, and Seresponsive diseases occur in sheep and cattle. New Zealand adults have a lower Se status compared with North American adults as assessed by low dietary intakes (1, 2), blood concentrations (3) and 24-hr urinary outputs of Se(4). Se is an integral part of the enzyme glutathione peroxidase (GSHPx) (5, 6), and GSHPx activity in blood has been correlated with dietary Se of chicks (7) and rats (8), and blood Se concentrations in sheep, cows, and pigs (9). A high correlation was found between Se concentrations and GSHPx activities in the blood of 264 New Zealand residents (10). Lombeck (1 1) reported low serum Se levels and erythrocyte GSHPx activities for healthy German subjects during infancy and early childhood; Se levels and enzyme activities of infants and children with phenylketonuria (PKU) and maple syrup urine disease (MSUD) were lower still, due to very low Se intakes from synthetic protein diets (12, 13). As yet, a human Se deficiency has not been demonstrated. However, it is possible that if The American
Dip.
concentrations in
Se also varied and
Glutathione
(Se) living
Children
B. Sc.,
Marion
selenium
children
dietary
Blood
Se intakes
Se was seen
P < 0.001) New
the
and
blood for
soils.
of food low
M. Rea,
M.H.Sc.;
ABSTRACT than
Heather
Nutrition
3 1 : AUGUST
such a state exists, then it might be found in New Zealand residents. In particular, infants and young children, and children on synthetic protein diets might be “at risk”. This paper reports Se concentrations and GSHPx activities in several groups of infants and children resident in New Zealand. Methods Subjects Random blood samples were collected from infants and children living in five areas of New Zealand: Auckland and Wellington, urban areas of the North Island where the soil Se is greater than 0.5 g g; Christchurch and Dunedin, urban areas of the South Island where the soil Se is less than 0.5 tg g’ and Tapanui, a rural area near Dunedin where Se responsive diseases occur in livestock and the soil Se is less than 0.5 zg g’. Blood samples were also collected from healthy adults living in Auckland, Dunedin, and Tapanui. The subjects were divided into the following groups (number of subjects in parenthesis): 1) healthy infants under 1 year of age and children living in Auckland (one infant, 14 children), I From the Otago, Dunedin, 2 Supported
Department of Nutrition, New Zealand. by the Medical Research
University Council
Zealand and the Medical Research Distribution mittee. 3 Address reprint requests to: Heather M. partment of Nutrition, University of Otago, Dunedin, New Zealand.
1978,
pp.
1413-14
18. Printed
in U.S.A.
of
of New ComRca, Box
Dc56,
1413
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Selenium concentration and glutathione peroxidase activity in blood of New Zealand infants and children13
1414
MCKENZIE
ET
AL.
ground to homogeneity were stored in a freezer. Analytical
where
necessary.
Liquid
foods
methods
Se in blood and synthetic foods was determined by a modification (15) of the diaminonaphthalene fluoriinetric method of Watkinson (16). Reproducibility of the method and recoveries of added Se were good (3). GSHPx activities were assayed in whole blood of 107 of the subjects (including six PKU children) by a modification (10) of the coupled enzyme assay of Paglia and Valentine (17) using t-butyl hydroperoxide as substrate.
Diet
Results
Dietary information relevant to Se intakes was collected for most subjects. Diets for children with MSUD and PKU were classified into: 1) diets containing synthetic foods (S diets); 2) restricted diets which excluded meat, fish, eggs, and poultry (R diets); 3) unrestricted or normal diets (U diets). Samples of the synthetic foods consumed by the. MSUD infant and by the infants and some of the children with PKU were obtained from Auckland hospital for analysis. Daily Se intakes of all subjects on S diets were estimated from 24-hr dietary recall using values for the Se content of synthetic foods as well as values for foods analysed in our department (14; G. M. Friend, unpublished data). Collection
and
storage
Table 1 gives mean blood Se concentrations for all subjects in each group as well as for those in which enzyme activities were estimated. Values have been included for healthy adults from Auckland, Dunedin, and Tapanui (18). Comparisons of blood Se values were made for the complete groups. There was little difference between the means for the complete group and for the smaller group for which enzyme activities had been measured. There was no difference in blood Se concentrations or in GSHPx activities between male and female subjects. Plasma Se was always less than erythrocyte Se, and these concentrations were reflected in that of whole blood. For 36 children from Tapanui, plasma
of samples
Samples of whole blood were collected into heparinized vacutainers (Becton-Dickinson and Co., Rutherford, N.J.), separated into plasma and erythrocytes where possible and stored below 4 C. The solid foods were dried to constant weight under an infrared lamp and
TABLE 1 Se concentrations of children and New Zealand”
and adults
GSHPx activities living in
Area
in whole
Age
blood
No.
Se concentration
yr
Healthy children and Auckland children
ig/m1
7 ± 3 7 ± 3 36 ± 4
15
Tapanui
children Adults
11 ± 2 35 ± 13
Hospitalized Wellington
children 7
± ± ±
±
± 0.0 17 ± 0.012 ± 0.012
18 59 12”
0.059 0.062 0.068
±
50 49
0.048 0.060
± 0.010 ± 0.012
15
0.063 0.059
±
0.043 0.042
± 0.010 ± 0.012
7.3
±
2.8
0.049 0.042
±
0.010 0.006
10.1
±
3.3
122
9 33 25
3 12 3
3
b
7 ± 2 Christchurch
Dunedin
Values
are
means
un#{252}s/gHb
0.064 0.060 0.083
13b
children Adults
a
GSHPx . activities
adults
Adults Dunedin
.
± SD
b
7 ± 4 3 ± 3
26
5 ± 3 3 ± 2
39
Subjects
10b
5b
for whi ch enzyme
activi
ties
assayed.
± ±
0.01 1 0.012 0.009
0.021 ± 0.012
±
10.6 ± 3.2 12.9 ± 3.1 10.6
±
2.2
10.9
±
2.5
9.3 ± 2.5
10.3 ± 3.3
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Dunedin (no infants, 19 children) and Tapanui (no infants, 50 children); 2) hospitalized infants and children in Wellington (one infant, 14 children), Christchurch (three infants, 23 children) and Dunedin (four infants, 35 children); 3) infants and children with PKU hiving in Auckland (three infants, 11 children) and Christchurch (one infant, three children) and one Auckland infant with MSUD. Second blood samples were collected from four children with PKU 5 to 8 months after the first bloods were taken; 4) healthy adult blood donors living in Auckland (122 subjects), Dunedin (59 subjects), and Tapanui (49 subjects.)
BLOOD
SELENIUM
CONCENTRATIONS TABLE Blood and
CHILDREN
2 Se concentrations
1415
of infants
children ,,
Country (reference)
Age group
Venezuela (20) Guatemala (21) Thailand (22) USA (23) West Germany (12)
children” 3-8 yrb 1-4 yrb Infantsb 1 mo-l yre 1-5 yrc 5_l2yrc S mo 1-5 yr 5-10 yr 1 mo-l yrb I_5 yr”
New
Zealand
9 7 16 9 10 7 1 6 24 8 24 29
5-lOyr” a Living in seleniferous Serum Se concentrations.
area.
b
0.8 13 0.23 ± 0.05 0.120 ± 0.036 0.097 0.051 0.081 0.097 0.036 0.057 ± 0.015 0.054 ± 0.009 0.035 ± 0.006 0.048 ± 0.012 0.057±0.017
Hospitalized.
18
16
.
14
12
D
.
19
t
o
I
0
102
004 Se concentrobon.
I
I
006
DIR
I
Ill
pg/mt
FIG. 1. Se concentrations and GSHPX activities in whole blood of New Zealand children (ages 1 month to 16 years); healthy and hospitalized children (#{149}), children with PKU (0).
differences among mean GSHPx activities of the different groups with the exception of the Christchurch hospitalized group that was lower than all the healthy groups of children (Auckland, P < 0.02; Dunedin, P < 0.002; Tapanui, P < 0.05). The mean GSHPx activity for the Auckland children was lower than that of 122 Auckland adults (P < 0.02). However, there were no differences in GSHPx activities between the healthy or hospitalized Dunedin children and a group of 12 Dunedin young adults, whose mean age was 25 ± 3 years (19).
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Se, and erythrocyte Se was 0.036 ± 0.007 and 0.061 ± 0.010 tg Se/mi, respectively, and whole blood Se was 0.047 ± 0.007 zg Se/ml, which was almost identical with the mean for all the Tapanui children. The mean hematocrit for these children was 0.39 ± 0.02. Such minor differences in hematocrit could not account for differences in whole blood Se concentrations. For the healthy groups, the mean blood Se concentration of the Tapanui children was lower than those of the Auckland and Dunedin groups (P < 0.01). For the hospitalized children the mean blood Se concentration of the Wellington group was higher than those of the Christchurch and Dunedin groups (P < 0.01) while that of the Christchurch group was lower than that of the Dunedin group (P < 0.02). Dunedin was the only area in which blood Se concentrations were determined in both healthy and hospitalized infants and children; the healthy group had a higher mean blood Se concentration than the group in hospital (P < 0.001). Overall, the mean Se concentration of all the healthy subjects (0.053 ± 0.0 13 jzg Se/ml) was similar to that of hospitalized subjects (0.050 ± 0.014 sg Se/ml). However, the mean of the total healthy group was heavily weighted by the lower values for the Tapanui children. Blood Se of the one healthy infant from Auckland (0.034 jsg/ml) and of the eight hospitalized infants from Wellington, Christchurch, and Dunedin all fell within the range of 0.021 to 0.042 .tg/ml. The mean for infants’ blood, 0.035 ± 0.006 .tg/ml was less than means for children in age groups of 1 to 5 and 5 to 10 years old (Table 2). There was a tendency for blood Se levels to increase with age in the healthy Dunedin group but not in any other group. The mean blood Se concentration of Dunedin children was a little lower than that recorded for 59 Dunedin adults (0.062 ± 0.012 sg Se/ml); the mean for the Auckland infants and children was lower than that for 122 Auckland adults (0.083 ± 0.012 Se/mi, P < 0.001) and the mean for the Tapanui children was lower than that for 49 Tapanui adults (0.060 ± 0.012, P < 0.001) (18). Figure 1 shows a strong correlation between the blood Se levels and GSHPx activities for 107 infants and children (r = 0.62; P < 0.001). However, there were no significant
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MCKENZIE
Infants
and
children
with
MSUD
and
PKU
TABLE 3 Se content of synthetic
foods
Branch chain free amino acid mixture (jg/g) Lofenalac powder (jzg/g) Aminogram food supplement (jsg/g) Aminogram mineral mixture (jg/g) Low phenylalanine bread (jtg/g) Prosparol (jzg/ml) Ketovite tablets (j.tg/tablet)
0.059 0.030 0.010 0.005 0.002 0.002 0.002
O8
.
#{149}
S
o7 O16 S
E
005 004
0#{149}03 002
cb 0#{149}01
0
2
I
I
I
I
I
I
4
6
8
10
12
14
age.yr FIG. 2. Se concentrations in whole blood of infants and children with MSUD and PKU; PKU synthetic diet (0), PKU nonsynthetic diet (#{149}), MSUD synthetic diet (L5).
AL.
which was almost the length of time that they had been on these diets. Blood Se concentration ofthree ofthe PKU children, from whom a second sample was taken, had decreased by 0.006 to 0.012 j.tg Se/mi: the fourth child showed no significant change in blood Se, which remained very low at 0.016 ig Se/mi. GSHPx activities were assayed for six PKU children living in Auckland, five on S diets. The mean GSHPx activity for these children was 8.9 ± 3.3 units per gram of hemoglobin, lower than that for 13 healthy Auckiand children. Discussion It has been well established that the blood Se concentrations of New Zealand adults are lower than those of adults living in other countries (3) and that New Zealand produced foods contain iess Se than most overseas foods (1, 14). The present study indicates that the blood Se of New Zealand children was also low compared with that of children in many other countries (Table 2). Estimations were performed by fluorimetry (with diaminonaphthalene) in all studies, except for that of the West German chiidren (12) in which serum Se only was estimated by neutron activation analysis. Their values for serum Se were much higher than those for plasma Se of Tapanui children. The major dietary sources of Se for man other than fish are cereals and animal products and the Se content of these foods is determined largely by the soil Se concentrations in the areas where they are produced (1); Se content of liver and kidney in low Se areas can also vary with the time animals are slaughtered after drenching with Se. Dietary Se intakes are, therefore, likely to vary geographically and this was reflected in differences in blood Se concentrations of children living in different areas of the country. Geographical variations had also been observed in blood Se within the New Zealand adult population (3). Serum Se concentration determined in some healthy West German infants and children increased from a median value of 0.051 I.Lg Se/mi, for nine infants ages 1 month to 1 year, to 0.097 jig Se/mi for seven children ages 5 to 15 years (12). Whole blood Se of New Zealand infants was also lower than the
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Se contents of the synthetic foods are presented in Table 3. Low phenylalanine bread had a lower Se content than normal white bread (0.015 jsg Se/g dry weight) but no other comparisons could be drawn between the synthetic foods and normal infant foods. The mean daily intake for the 12 subjects on S diets was 5 ± 2 zg Se day’, with little difference between the older children and the infants. Figure 2 shows the blood Se concentrations of infants and children with PKU and MSUD. The mean blood Se concentration of those on S diets (0.038 ± 0.013 tg Se/ml) was lower than the blood Se of six subjects on R and U diets (0.074 ± 0.009 g Se/mi, P < 0.001). The blood Se concentrations of subjects on S diets appeared to decrease with age
ET
BLOOD
SELENIUM
CONCENTRATIONS
CHILDREN
1417
being on an intake of less than 5 .tg Se day’ for 13 years. This study then indicated that the low blood Se of New Zealand children is determined primarily by low dietary Se intake, and this was reflected in lowered GSHPx activities. Lombeck et al. (13) reported lower GSHPx activities for 19 PKU children. In the present study the mean GSHPx activity for six PKU children was lower than that for heaithy children but this was not significant for the small number of subjects studied. The strong correlation between blood Se levels and GSHPx activities for the infants and children confirms the previously established correlation for 264 New Zealand subjects ages between 2 and 92 years (10), and shows the same kind of regression as for values less than 0. 1 .tg Se/mi whoie blood. Se concentrations of 0.016 tg Se/mi whole blood and 0.009 ig Se per milliliter of plasma found for subject Q.R. were the lowest in this study and lower than most values recorded in the literature. A clinical examination of subject Q.R. indicated that he was in good health. Low concentrations for serum Se, 0.002 to 0.008 .tg Se/mi, were aiso recorded for infants and children on special PKU and MSUD diets in West Germany ( 12). These subjects were reported to be “in good clinical state and thriving well”. Although vitamin E intake of these children on synthetic diets and those in the present study was not determined, it is unlikely that it was inadequate and may protect against the effects of low Se intake. However, Gross (26) has observed that hemolytic anemia can still occur in premature infants on milk formulas even in the presence of adequate serum tocopherol levels. He found that feeding infants a low Se milk formula resulted in decreasing blood Se concentrations and GSHPx activities. But it was the additional oxidant stress of high polyunsaturated fatty acid content or added iron which resulted in reduced hemoglobin, increased reticulocyte response and hydrogen peroxide fragility values and also peroxidation-induced membrane damage as indicated by decreased erythrocyte phosphatidyl ethanolamine. This requires further investigation in some of our New Zealand infants. U and
The authors are Miss Veronica
very grateful Doesburg
to Miss for their
Gaylene technical
Friend assist-
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children but a tendency to increase with age above 1 year was seen oniy in the Dunedin healthy children. Furthermore, mean blood Se concentrations of groups of children in different parts of the country were lower than values recorded for adults in corresponding areas (Table 1). These variations with age possibly reflect differences in the quantities and types of foods eaten. Griffiths (2) found the range of daily Se intakes of some New Zealand adults living in Dunedin was 6 - 70 jig Se daf’ with intakes greater than 35 tg Se day’ occurring only on days when Se-rich foods, such as fish, liver, and kidney were eaten. Using our values for the Se content of foods (14; G. M. Friend, unpublished data) and 24-hr dietary recali data collected for healthy New Zeaiand infants ages 2 months to 5 years (24; F. Davidson, personai communication) an estimate of a daily intake of 5 to 8 .tg Se day’ was obtained. It appears that the dietary Se intake of New Zealand residents might increase from about 5 jg Se day’ during infancy up to 20 to 30 jsg day’ during adolescence and adult life. Elderly subjects (over 70 years) had lower blood Se concentrations than younger adults (19) and this might also reflect the reduced food intake of this age group. The lower mean blood Se concentration of the hospitalized Dunedin group, as compared with the healthy group, might have resulted from a lower food intake during ill health. However, tissue wasting during illness at a time when growth is important could also have influenced blood Se levels. Infants and children with MSUD and PKU are fed restricted diets containing synthetic proteins for at least the first 3 years of life and often for longer periods. Since Se is mainly associated with the protein fraction of food (25), it was not surprising that the Se intake was very low for subjects on synthetic diets and remained low for the older children. The blood Se concentrations of the older chiidren were much lower than those on restricted or unrestricted diets (Fig. 2). This trend was further illustrated by the decreased Se concentration of the second blood samples from three PKU children (Fig. 2). A fourth child, subject Q.R. 13 years old showed little change. This might indicate that the blood Se had stabilized around 0.016 tg Se/mi after
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1418
MCKENZIE
13.
14. 15.
References 16. 1. 2.
3.
4.
5.
6.
7.
J. H. The selenium status of New Zealanders. N. Z. Med. J. 80: 202, 1974. GRIFFITH5, N. M. Dietary intake and urinary excretion ofselenium in some New Zealand women. Proc. Univ. Otago Med. Sch. 51: 8, 1973. GRIFFITHS, N. M., AND C. D. THoMsoN. Selenium in whole blood of New Zealand residents. N. Z. Med. J. 80: 199, 1974. THOMSON, C. D. Urinary excretion of selenium in some New Zealand women. Proc. Univ. Otago Med. Sch. 50: 31, 1972. ROTRUCK, J. T., A. L. POPE, H. E. GANTHER, D. G. HAFEMAN AND W. G. HOEKSTRA. Selenium: biochemical role as a component of glutathione peroxidase. Science 179: 588, 1973. FLOHE, L., W. A. GUNZLER AND H. H. SCHOCK. Glutathione peroxidase: a selenoenzyme. Febs. Lett. 32: 132, 1973. NooucHl, T., A. H. CANTOR AND M. L. Scorr. Mode of action of selenium and vitamin E in prevention of exudative diathesis in chicks. J. Nutr. 103: WATKINSON,
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D. G., R. A. SUNDE AND W. G. HOEKEffect of dietary selenium on erythrocyte and liver glutathione peroxidase in the rat. J. Nutr. 104: 580, 1974. 9. THOMPSON, R. H., 0. H. MCMURRAY AND W. J. BLANCHFLOWER. The levels of selenium and glutathione peroxidase activity in blood of sheep, cows and pigs. Res. Vet. Sci. 20: 229, 1976. 10. THOMSON, C. D., H. M. REA, V. M. DOESBURG AND M. F. RoBINSoN. Selenium concentrations and glutathione peroxidase activities in whole blood of New Zealand residents. Brit. J. Nutr. 37: 457, 1977. 1 1. LOMBECK, I., K. KASPEREK, H. D. HARBISCH, L. E. FEINENDEGEN AND H. J. BREMER. The selenium state of healthy children. Europ. J. Pediat. 125: 81, 1977. 12. LOMBECK, I., K. KASPEREK, L. E. FEINENDEGEN AND H. J. BREMER. Serum-selenium concentrations in patients with maple-syrup-urine disease and HAFEMAN,
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AL. phenylketonuria under dietotherapy. Chin. Chim. Acta 64: 57, 1975. LOMBECK, I., H. D. HARBISCH, K. KASPEREK, L. E. FEINENDEGEN AND H. J. BREMER. Decreased serum selenium concentrations and reduced glutathione peroxidase activity in dietetically treated patients with metabolic diseases. Chem. Scripta 8A: 109, 1975. RoBINSoN, M. F. The moonstone. J. Human Nutr. 30: 79, 1976. THOMSON, C. D. The metabolism and nutritional importance of selenium in the human population of New Zealand. University of Otago: Ph.D. Thesis, 1973, p. 27. WATKINSON, J. H. Fluorimetric determination of selenium in biological material with 2,3-diaminonaphthalene. Anal. Chem. 38: 92, 1966. PAGLIA, D. E., AND W. N. VALENTINE. Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J. Lab. Clin. Med. 70: 158, 1967. RoBINSoN, M. F., C. D. THOMSON, R. D. H. STEWART. H. M. REA AND R. L. MCKENZIE. Selenium in human nutrition in New Zealand residents. In: Proceedings of Third International Symposium on Trace Element Metabolism in Man and Animals, edited by M. Kirchgessner. TEMA 3, 1978, in press. THOMSON, C. D., H. M. REA, M. F. RoBINSoN AND 0. W. CHAPMAN. Low blood selenium concentrations and glutathione peroxidase activities in elderly people. Proc. Univ. Otago Med. Sch. 55: 18, 1977. JAFFE, W. G., D. M. RUPHAEL, M. C. MONDRAGON AND M. A. CUEVAS. Estudio clinico y bioquimico n ninos escloares de una zona selenifera. Arch. Latinoamer. Nutr. 22: 595, 1972. BURK, R. F., W. N. PEARSON, R. P. WooD AND F. VITERI. Blood selenium levels and in vitro red blood cell uptake of75Se in kwashiorkor. Am. J. Chin. Nutr. 20: 723, 1967. LEVINE, R. J., AND R. E. OLSON. Blood selenium in Thai children with protein-calorie malnutrition. Proc. Soc. Exptl. Biol. Med. 134: 1030, 1970. RHEAD, W. J., E. E. CARY, W. H. ALLAWAY, S. L. SALTZSTEIN AND G. N. SCHnAUZER. The vitamin E and selenium status of infants and the sudden infant death syndrome. Bioinorg. Chem. 1: 289, 1972. HARDING, W., A. VAN Ru AND F. DAVIDSON. Diets ofNew Zealand pre-schcol children. J. N. Z. Dietet. Assoc. 29: 17, 1975. BURK, R. F. Selenium in man. In: Trace Elements in Human Health and Disease, edited by AS. Prasad. New York: Academic Press 1976, vol. II, p. 105. GROSS, S. Hemolytic anemia in premature infants: Relationships to vitamin E, selenium, glutathione peroxidase and erythrocyte lipids. Seminars Hematol. 13: 187, 1976.
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ance, to the following people for collecting blood samples from the children: Professor D. Lines, Department of Paediatrics, Auckland Hospital; Dr I. C. T. Lyon, Dcpartment of Community Health, Auckland University Medical School; Professor H. J. Weston, Department of Paediatrics and Child Health, Wellington Hospital; Professor F. T. Shannon and Dr R. Ford, Department of Paediatrics, Christchurch Hospital; Professor J. M. Watt, Department of Paediatrics, Otago University Medical School; and Dr P. G. Snow, Tapanui; and Miss P. King, Dietary Department, Auckland Hospital, for collecting samples of the synthetic foods.
ET
