ZINC NUTRITIONAL STATUS OF PRESCHOOL CHILDREN IN SELECTED 
COMMUNITIES OF SOUTHERN GHANA
BY
ETOR ERIC KOMLA TAKYI
A THESIS SUBMITTED TO THE DEPARTMENT OF NUTRITION AND 
FOOD SCIENCE IN PARTIAL FULFILMENT OF THE 
REQUIREMENTS FOR THE AWARD OF THE M.PHIL. DEGREE IN 
NUTRITION
DEPARTMENT OF NUTRITION AND FOOD SCIENCE 
UNIVERSITY OF GHANA 
LEGON
DECEMBER 1994.
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Q -146391
T X  3,61, CS  T I
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This study was undertaken by me. as presented, under the 
supervision of Dr. Ebenezer Asibey-Berko, of the Department of 
Nutrition and Food Science.
University of Ghana 
Legon.
DECLARATION
( CANDIDATE ) ' ( SUPERVISOR )
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DEDICATION
To God Almighty, for His love, protection, and enablement 
which made it possible for me to study at this rather advanced 
age.
To my family for their help and everything that they have 
sacrificed to enable me complete this study.
To all my lovely cohorts who without least hesitation 
willingly participated in the study. During my recent visits 
to them, they were very happy to see me and I feel I can hear 
them telling me 'Please, come again'. I shall always remember 
them when I pick up this thesis to read.
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ACKNOWLEDGEMENT
I thank my supervisor, Dr. Ebenezer Asibey-Berko, for all the 
help he ably gave me in the successful execution of this project.
I also wish to thank the following companies who generously 
donated some of their products to be used as incentives,thus making 
the management of the project relatively easy:
1. Nestle Ghana Ltd.
2. Cocoa processing company, Tema.
3. Cadbury Ghana Ltd.
The National Institute of standards and Technology, USA, also 
donated free of charge, standard bovine liver which was used to 
test the accuracy of my analyses. I wish to thank them most 
sincerely.
I wish to thank Mr. Eric Harrison, Mr.Emmanuel Quansah, and 
Mr. J. W. Asaku, for their invaluable assistance in the field work. 
For providing me with invaluable literature on zinc and other 
supports, I wish to thank Professor F. K. Nkrumah, Director, NMIMR.
Dr. R. Acquaah of Biochemistry Department, University of 
Ghana, generously donated p-Nitrophenol phosphate for the 
determination of alkaline phosphatase activity, as well as giving 
some useful tips on the assay. I thank him most sincerely.
All the data collected would have been of no use if not 
properly analyzed statistically. It is for this reason that I thank 
both Mr. George Mensah and Mr. Patrick Chinery of the epidemiology 
unit of NMIMR, for their assistance in the analyses of my results.
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Last but not the least, I thank all others, who in any way 
have helped me to complete this study.
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PAGE
CONTENT
TITLE PAGE 
DECLARATION
DEDICATION 111
ACKNOWLEDGEMENT 1V
TABLE OF CONTENTS vl
SUMMARY X
CHAPTER ONE 1
INTRODUCTION L
1.2. General objective 8
1.3. Specific objectives 8
CHAPTER TWO 9
LITERATURE REVIEW 9
2.1. Elements essential to man 9
2.2. Recognized trace element deficiencies in man 10
2.2.1. Iron 10
2.2.2. Copper 12
2.2.3. Chromium, manganese and iodine 15
2.2.4. Zinc 17
2.2.4.1. Distribution of zinc in human body 18
2.2.4.2. Biochemistry and the physiological roles of zinc 19
2.2.4.3. Zinc deficiency 21
2.2.4.4. Human zinc nutrition and deficiency 24
2.2.4.5. Zinc Toxicity 32
2.2.4.6 . Assessment of zinc status in man 33
TABLE OF CONTENTS
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2.2.4.7. Estimation of zinc concentration 35
2.2.5. Variables measured and justification for their
choice 36
2.2.5.1. Variables 3 6
2. 2. 5. 2. Justification 36
CHAPTER THREE 41
METHODOLOGY 41
3. Study design and plan of work 41
3.1. Study population 41
3.2. Subjects selected for the study 41
3.3. Study areas 41
3.4. Plan of work 42
3.4.1. Anthropometric measurements 42
3.4.2. Sampling of hair 43
3.4.3. Blood collection 43
3.5. Laboratory analyses 43
3.5.1. Determination of hair zinc 43
3.5.1.1. Sampling of hair 43
3.5.1.2. Washing of Hair 44
3.5.1.3 Wet-ashing of hair & analyses by AAS 44
3.5.1.4. Quantification of zinc 45
3.5.2. Determination of plasma and erythrocyte zinc 45
3.5.2.1. Preparation of plasma and erythrocytes 45
3.5.2.2. Erythrocyte zinc analysis 46
3.5.3. Determination of Plasma zinc levels 46
3.5.4. Determination of Plasma total protein 47
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3.5.5 Albumin determination 48
3.5.6. Measurement of Alkaline phosphatase activity 48
3.5.7 Statistical analysis of results 50
CHAPTER FOUR 52
RESULTS 52
4.1. Age distribution of the cohorts 52
4.2. Anthropometric classification of cohorts 52
4.3. Biochemical data based on nutritional/growth
status 56
4.4. Correlation among some indicators of zinc nutriture
and anthropometric parameters 62
CHAPTER FIVE 63
DISCUSSION OF RESULTS 63
CHAPTER SIX 71
CONCLUSIONS AND RECOMMENDATIONS 71
6.1. CONCLUSIONS 71
6.2. RECOMMENDATIONS FOR FUTURE WORK 72
7. APPENDIX 73
8 . REFERENCES 79
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LIST OF TABLES
Table 1. Anthropometric classification of the cohorts
Table 2. Mean values of the anthropometric and biochemical 
parameters of the cohorts
LIST OF FIGURES
Fig. la. Age distribution of cohorts in the various 
communities
Fig lb. Level of malnutrition ( wasting, stunting, wasting 
plus stunting) among preschoolers in the communities 
studied.
Fig. 2a. Nutritional status in the various age groups
Fig. 2b. Skinfolds and mid-upper arm circumference 
measurements of the cohorts
Fig 3a. Trends in the mean plasma protein, albumin, and A/G
ratios in the various communities
Fig. 3b. Plasma zinc and rbc zinc levels in the various
communities
Fig. 4a. Trends in the mean Albumin/Globulin ratios in the 
types of growth
Fig 4b. Albumin levels in the different nutritional states
Fig. 5a. Trends in the mean alkaline phosphatase activity 
in different types of growth
Fig 5b. Plasma zinc levels in the various nutritional states
in the communities
Fig. 6a. Trends in the mean hair zinc in the various 
nutritional states in the communities
Fig. 6b. Mean height and alkaline phosphatase activity in the 
communities
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55
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60
60
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SUMMARY
Zinc is a micronutrient indispensable for growth, development, 
reproduction, and for the activities of over 200 enzymes embracing 
all physiological activities. Zinc deficiency in the preschool age 
group therefore leads to growth faltering, wasting, stunting and 
general degeneration in physiological activities.
In rural areas of Egypt and Iran where zinc deficiency was 
first identified, dietary factors of plant origin ( especially 
phytic acid and fibre), which impair zinc absorption, were the main 
etiological factors of zinc deficiency. For the fact that recent 
studies have suggested that zinc deficiency might be as alarming as 
iron deficiency and the fact that zinc deficiency predominates in 
populations which subsist largely on vegetable diet with little 
animal protein, it was felt that this study, is both relevant and 
necessary in the Ghanaian context.
Studies were carried out in 4 communities on 200 preschool 
(nursery) children aged 3 to 5 years, to determine if zinc 
deficiency occurs within this age group; and if it does, whether it 
relates in anyway to any of the anthropometric indices of the 
cohorts.
The four communities were Ashalley Botwe, Kwabenya,( typical 
rural southern villages), Dome (periurban) and New Achimota 
( urban), all located in Greater Accra region, southern Ghana.
In the study, age and the anthropometric indicators of 
nutritional status ( weight, height, mid upper arm circumference, 
triceps, and subscapular skin-folds), and the biochemical
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indicators of zinc nutriture [ hair zinc, plasma zinc, red blood 
cell (rbc) zinc, and alkaline phosphatase activity], as well as 
indicators of protein nutriture (plasma protein, albumin, and A/G
ratio) were determined.
Results of anthropometric measurements indicated that the mean 
percentage of pre-schoolers affected by Wasting, Stunting and 
Wasting plus stunting in the four communities combined were 3.5%, 
16.5%, and 1.5% respectively, with 78.5% of normal status. When the 
results were considered for each community, 69.6% (Ashalley Botwe), 
81.2% (Kwabenya), 80.9% (Dome), and 84.1% (New Achimota),
respectively, were of normal stature. Percentage wasting was 3.6%, 
0%, 4.4% and 4.5% while stunting levels were 25%, 18.8%, 13.2%, and 
9.1% respectively. The percentage wasting plus stunting was 1.8%, 
0%, 1.5%, and 2.3%, respectively.
Statistical analysis using Duncan's and Least significant 
difference (LSD) multiple comparison tests, indicated that there 
were no significant differences (p>0.05) in the mean values for the 
indicators of zinc nutriture (plasma zinc, rbc zinc, hair zinc, and 
plasma alkaline phosphatase activity) in the different nutritional 
states (normal, wasted, stunted, wasted plus stunted).
Further comparison with reference values indicated that there 
was no zinc deficiency in any of the groups- eg. the mean plasma 
zinc values obtained for the normal, wasted, stunted, and wasted 
plus stunted groups ( for the 200 cohorts) were 1.13+0.35, 1.16 
± 0.37, 1.04 + 0.23 and 0.95 + 0.29 ppm, respectively, as compared 
to a normal range of 0.50-1.50ppm. The corresponding values for the
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hair zinc were 247.0 + 101.6, 200.9 +. 65.2, 220.0 +, 83.8 and 157.6 
+ 40 ppm, as compared to a normal level of >70 ppm. Also, the mean 
plasma, red blood cell, and hair zinc values for all the cohorts 
were normal.
Analysis of indicators of protein nutritional status (total 
plasma protein, albumin, and A/G ratio) revealed that the Plasma 
Protein values for the various anthropometric states were within 
the normal reference ranges eg. the mean Albumin/Globulin ratio 
for the normal, wasted, stunted and wasted plus stunted groups 
( for the 200 cohorts) were 1.6 +0.5, 1.2 + 0.3, 1.6 + 0.5, and
1.5 + 0.5, respectively, as compared to a normal range of 1-2.5. 
This indicates that the protein nutritional status was adequate.
It was- concluded that there was no zinc deficiency in the 
cohorts, and that zinc nutritional status did not differ in the 
various nutritional states of the cohorts.
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CHAPTER ONE
In line with the generally accepted idea that the nutritional 
status of a child is perhaps the most important determinant factor 
in the survival and/ or health status of that child, a lot of 
efforts have been made to address this problem.
Despite these efforts, malnutrition continues to worsen in the 
developing world. The lack of progress is due to the many varied 
and complex factors which affect the nutritional status of children 
but which can not be easily controlled or met, mainly due to 
adverse socio-economic factors.
There are four principal nutritional deficiency diseases known 
in the world today1. These are:
1.Protein-energy malnutrition (PEM)- caused by deficiency of 
protein (leading to kwashiorkor) or energy (leading to 
marasmus) or deficiency of both protein and energy (leading to 
marasmic-kwashiorkor).
2. Anaemia- due mainly to iron deficiency.
3. Goitre and hypothyroidism- caused by iodine deficiency.
4. Xerophthalmia- caused by vitamin A deficiency.
Whereas iodine and vitamin A deficiency diseases are found in 
definite areas with inadequate iodine in the soil, and vitamin A in 
the diet, PEM and nutritional anaemias are rather widespread.
Iron deficiency anaemia is the most prevalent nutritional 
deficiency disease in the world and it is estimated that about 29%
INTRODUCTION
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of the world population or 43% of people in Africa suffer from iron 
deficiency anaemia2. In fact, it is the only nutritional deficiency 
showing a general deterioration in many parts of the world, such as 
south Asia and sub-saharan Africa2.
Iodine deficiency, which causes endemic goitre and 
hypothyroidism leading (if untreated) to retarded growth, mental 
retardation, deafness and deaf-mutism (in infants) affects about 
400 million people world-wide with a further 400 million people at 
risk3. Vitamin A deficiency affects about half a million children 
globally each year leading to partial or total blindness with about 
two-third of this number eventually dying, within a few months of 
loosing their sight4.
It is now becoming clear that many nutritionally-important 
trace elements may become deficient in the diets of apparently 
healthy individuals, thus a lot of attention is now being focused 
on this area.
As at now, fourteen elements have been shown to be required by 
the human body in quantities less than a few micrograms per day. 
These are chromium, cobalt, copper, fluorine, iodine, iron, 
manganese, molybdenum, nickel, selenium, silicon, tin, vanadium and 
zinc5- Most nutritionists believe that the list would be extended 
in future as more and more evidence become available. However, 
deficiency states which have responded to specific replacement or 
supplementation have been described for only iron, iodine, copper, 
chromium, manganese and zinc6.
These micronutrients are important nutritionally because they
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are mostly components of many enzymes7. The metals function in the 
enzymes by (a)direct participation in catalysis, (b) combination 
with substrate to form a complex upon which the enzyme acts 
(c)formation of metalloenzyme which binds substrate (d) combination 
of metal with a reaction product and / or (e) maintenance of 
quaternary structure of the enzyme7. Thus,in the absence of these 
micronutrients, vital physiological processes are inhibited.
Even though the biological importance of zinc has been 
recognised for over a 100 years now, human deficiency states were 
not described until 19638. As at now, about 200 zinc-metalloenzymes 
have been identified9; all key metabolic pathways have zinc 
metalloenzymes5 -
When an animal is given a zinc-deficient diet, plasma zinc 
levels fall in the first day and within three days the animal 
becomes anorexic ( loss of appetite) and stops growing5. Forced 
feeding, particularly of a high-protein diet, leads to severe 
metabolic disturbances. As the deficiency continues, skin lesions 
develop, particularly around the body orifices. These progress to 
form ulcers which quickly become colonized with bacteria. At this 
stage, there is immuno-deficiency, particularly of the cell 
mediated immune system. Diarrhoea ensues and the animal enters a 
phase of gross wasting and finally death2. In the early stages, 
zinc supplementation leads to reversal of the condition within an 
hour, in the latter stages within a day or so5. This pattern has 
been observed in at least fifteen species of animals, including 
man5 -
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It has been shown that zinc is an essential micronutrient, 
implicated in many biological processes including growth, enzyme 
function, protein synthesis, sexual maturation, wound healing and 
host defence reactions10'14. In conditions of zinc deficiency or 
inadequacy therefore the above processes are retarded or abolished, 
depending on the level of deficiency.
The etiology of growth retardation is unknown but it is 
suggested to be related to nutrition or genetics or both15; however 
it is believed that,in the pre-school age, nutritional factors 
predominate over genetic factors16.
Features of nutritional zinc deficiency in animals and man 
include anorexia, hypogonadism, (under-development of sex 
organs),lethargy, behavioral changes, increased susceptibility to 
infection and impaired development of the immune system10'14. 
Diminished height has been described in infants and children with 
poor zinc nutriture15. In fact zinc deficiency has been described 
as the cause of pronounced growth retardation, dwarfism and 
impairment of sexual maturation in young people living in Egypt and 
Iran who, consume diets of very low zinc bioavailability17. A milder 
form of zinc deficiency with slight growth retardation, poor 
appetite and impaired taste acuity was described over two decades 
ago in children of middle or upper-income families in Denver, 
Colorado. These children were presumed to be of a good nutritional 
status18. Since then, more studies have identified marginal zinc 
deficiency in preschool and school children of several countries19'
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Due to the ubiquitous presence of zinc in foods, classical 
symptoms of severe zinc deficiency are rarely observed. However, 
nutritional surveys and recent clinical findings with human 
patients suggest that marginal or mild zinc deficiency may be 
widespread in developed nations22. Consequently, human zinc 
deficiency may represent health concern throughout the world, 
particularly during preschool, adolescent, pregnancy, old age, 
stress or in disease.
Evidence is also slowly accumulating that deficiencies of 
other trace elements may be as prevalent as that of iron and that 
alteration of their metabolism may be the cause of many diseases 
bothering mankind5.
In Ghana, both protein-energy malnutrition and micronutrient 
deficiency abound. A national nutrition survey in 1961 reported 
that about 40-50% of all preschool children were underweight23- A 
similar survey in 1986 reported that 58.4% of children aged 0-5 
years were underweight. A World bank report in 1988 indicated that 
8.2% of preschool children were suffering from a severe form of PEM 
in the form of marasmus or kwashiorkor. This figure of 8.2% is 
about twice that usually found in low income countries24.
Nutritional surveys in 1988 have also revealed that about 28% 
of preschool children in Ghana were stunted25. The exact cause(s) 
of stunting have not been clearly identified; however, PEM and 
micronutrient deficiencies are implicated.
In the past, research in Ghana has concentrated largely on 
finding the causes and /or management of protein-energy
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malnutrition, vitamin A,iron and lately iodine deficiencies. In 
contrast, nothing is known about the zinc status of any age group 
in Ghana. In fact Golden and Golden5 believe that we must ask 
ourselves critically if children with malnutrition are indeed zinc 
deficient since malnutrition and experimental zinc deficiency have 
clinical signs in common. Anorexia, diarrhoea, stunting growth, 
wasting, skin desquamation with ulceration, a reduction in lymphoid 
tissue and increased susceptibility to infection are common to the 
two conditions5.
This study was therefore planned to ascertain if zinc
deficiency is one of the factors associated with malnutrition in 
preschool children in some selected communities in Ghana.
Zinc deficiency was considered a likely etiological factor 
in malnutrition in preschool children due to the fact that
Ghanaians of all age groups consume cereals and vegetables as their 
staple food with only a little amount of animal protein. Zinc in 
foods is primarily associated with proteins and nucleic acids26. It
is particularly abundant in red meats, some sea foods and the
embryo portions of grains. However, zinc in plant products is 
generally less available than that supplied by animal proteins27. 
A major contributing factor is the presence of phytic acid, which 
forms an insoluble zinc-chelate complex ( zinc phytate) that is not 
absorbed from the gastrointestinal tract27. High dietary calcium has 
been shown to enhance the adverse effect of phytic acid. Fibre also 
reduces zinc bioavailability while food processing can also 
markedly alter zinc bioavailability from that present in the
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unaltered food28 -
It has been shown in rural Iran and Egypt where zinc 
deficiency was first identified that dietary factors especially 
phytic acid and fibre were the main etiological factors of zinc 
deficiency17. These plant materials impair zinc absorption. Also in 
the USA, 10 out of 338 middle and upper class children consuming 
diets consisting of little meat and a great deal of cow milk - 
which is a relatively poor source of zinc - had very low hair zinc 
levels17.
Thus the bioavailability of zinc in the largely vegetable 
Ghanaian diet would be further reduced in the body with the result 
that the RDA of zinc (10-15 mg) might not be met in majority of 
people. This is especially in preschoolers( where zinc requirement 
is increased as a result of rapid growth rates) leading to zinc 
deficiency with its attendant consequences.
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1.2. General objective.
To determine if zinc deficiency exists in preschool children 
in selected communities in Ghana; and if so, whether the 
zinc levels relate to the various nutritional states of the 
cohorts .
1.3. Specific objectives
i. To determine the extent of malnutrition in preschool 
children in the selected communities using anthropometric 
measurements.
ii. To study the correlation between zinc status and growth in 
normal and malnourished preschool children in the selected 
communities.
iii. To determine the level of hair zinc, plasma zinc 
erythrocyte zinc and alkaline phosphatase activity in the 
subjects.
iv. To determine plasma total protein, albumin, and 
Albumin/Globulin ratio as indicators of protein nutritional
status in the cohorts.
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CHAPTER TWO
LITERATURE REVIEW
2.1. Elements essential to man.
Of the ninety naturally occurring elements, twenty six are 
known to be essential to animal life6. Eleven of these are regarded 
as major elements because of their relative abundance in the body 
each is present in the body at concentration of more than 0.005% 
body weight7 - these are: carbon, hydrogen, oxygen, nitrogen,
sulphur, calcium, phosphorus, potassium, sodium, chlorine and 
magnesium.
The remaining essential elements are known as trace elements 
because they are present in animal tissues in such relatively small 
amounts ( each is less than 0.005% body weight), that the early 
researchers were unable to measure their concentrations accurately.
Trace elements which are essential in various animal species 
are iron, chromium, copper, fluorine, iodine, manganese, 
molybdenum, nickel, selenium, tin,cobalt silicon,vanadium and 
zinc5. In man, however, deficiency states which have responded to 
specific replacement or supplementation have been described for 
only iron, copper, chromium, manganese, iodine and zinc6'29' 34 (table
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2.2. Recognized trace element deficiencies in man- Table 1 16'29-34
Element Some effects of deficiency
Iron Anaemia,impaired learning ability
Iodine Endemic goitre, congenital 
hypothyroidism leading to 
retarded growth, mental retarda­
tion, deafness and deaf-mutism.
Copper Falling levels of serum copper 
and ceruloplasmin,failure of iron 
absorption, neutropenia, 
leucopenia, bone demineralization, 
failure of erythropoiesis and 
death.
Chromium Impaired glucose tolerance, poor 
growth, decreased nitrogen reten­
tion, ataxia, defective peripheral 
ne rve c onduc t i on.
Zinc Impaired appetite, growth 
retardation, delayed closure of 
epiphyses, pica, hypogeusia and 
delayed sexual maturation.
Manganese Low clotting factor
2.2.1. Iron
Iron deficiency and associated anaemia remain the most common 
and widespread nutritional disorder in the world today5, and affect 
over one billion people worldwide35. There is concern that the 
adverse effects of inadequate iron may not be limited to anaemia. 
It has been found that the function of haem proteins and the 
morphology of some subcellular structures can be disturbed and the 
future learning ability may be adversely affected by iron 
deficiency during infancy34.
Although iron is essential for the existence of all plants and
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animals, only trace amounts are present in living cells. These 
small but biologically indispensable amounts are related to the 
relative insolubility of iron and to the specialised systems 
required for its transport and incorporation into iron proteins: 
haemoglobin, myoglobin, haem enzymes( cytochromes),non-haem iron 
enzymes (flavoproteins) , ferritin, transferrin,and hemosiderin36-
The body carefully controls its stores of iron by limiting 
absorption and by reusing iron derived from breakdown of 
haemoglobin and catabolizing of nonhaem iron proteins. Although 
there is no planned iron excretion as such, there is a daily 
physiological loss of about 1 mg iron that must be replaced with 
intestinal absorption of 5-10% of the dietary 10-30 mg/ day37.
The iron needs must be satisfied by absorption from haem and 
nonhaem food iron. In infants and children, the daily requirements 
are relatively high: from birth to 6 months, 10 mg/day; from 6
months to 3 years, 15 mg/day; for boys 11 to 18 years, 18 mg/day, 
dropping to 10 mg for older men, while it is 18-23 mg/day for 
girls/women aged 10 to 50 years, dropping to 10 mg in the older 
ones7.
Iron deficiency is the most common cause of anaemia and is 
significantly more frequent in women of childbearing age37. In the 
developing countries anaemia is frequent in both sexes and in 
children of low socioeconomic background36.
The classical symptoms of anaemic-hypoxia such as weakness, 
palpitation, pallor, and dizziness may be absent in even moderate 
to severe anaemia ( less than lOOg haemoglobin/L blood), whereas
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persons with latent iron deficiency (low serum iron, depleted iron 
stores,but no anaemia), may complain of these symptoms38.
More recent investigations show a significant relationship 
between iron deficiency and physical fitness and work capacity in 
adults39 and between iron deficiency and mental performance and 
irritability in children40. In iron deficiency, monoamine oxidase 
activity is decreased leading to increased excretion of 
norepinephrine in urine. Norepinephrine is believed to influence 
behaviour in humans40,41.
2.2.2. Copper
Despite evidence dating back to Josephs since 193l42, it has 
repeatedly been stated that copper deficiency did not or could not 
exist in man43. Deficiency states of copper have now been found in 
a number of conditions such as severely malnourished infants 
rehabilitated on milk-based, low copper diets44; in untreated 
malnourished infants45, premature infants46, malnourished infants 
alimented exclusively by the intravenous route47, and in adults48.
Copper is essential for life because of its presence in 
certain metalloenzymes29,30'32. For example, as a component of 
cytochrome oxidase, copper is important in mitochondrial oxidative 
phosphorylation which produces Adenosine triphosphate6.
Copper is essential for the function of lysyl oxidase, an enzyme 
required in the cross-linking that occurs in collagen and elastin. 
Tyrosinase, a copper-dependent enzyme, is required for formation of 
melanin.
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As a component of superoxide dismutase, copper may be 
important in protecting newborn lungs from oxygen damage since this 
enzyme protects cells from damage by superoxide radicals.
Copper is also an important component of ceruloplasmin, a 
metalloenzyme synthesised in the liver in response to increased 
dietary copper, estrogens, androgens and leucocyte endogenous 
mediator6. Ceruloplasmin has many important functions6-30,32 ; these 
include oxidation of ferrous iron to ferric iron, the form in which 
iron is bound by apotransferrin for iron transport to the bone 
marrow; it oxidises amines such as epinephrine, and may serve as a 
transport protein for copper49 -
In copper deficiency, anaemia could result. This is 
attributable to a lack of copper-containing ferroxidase, including 
ceruloplasmin that are required for oxidation of ferrous iron to 
ferric iron and this prevents transport of iron to bone marrow. 
Deficiency of copper-containing amine oxidases required for cross- 
linking of elastin and collagen leads to defects in the walls of 
blood vessels; in animals, death may result from rupture of a major 
artery. Melanin synthesis is also impaired by copper deficiency50.
In a study of copper-deficient infants hospitalised in Lima, 
Peru, it was found that neutropenia and leucopenia were the prime 
early indicators of copper deficiency. Anaemia due strictly to 
copper deficiency occurred only late in the course of the 
deficiency and bone disorders still later50.
There are two well-defined genetic diseases, Menkes' syndrome 
and Wilson's disease in which copper metabolism is disrupted and
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hypocupremia occurs inspite of adequate dietary copper intake51. 
Menkes' syndrome is a sex-linked, genetic disease in human male, 
characterised by slow growth, cerebral degeneration and early 
death. The pathology is similar to that observed for copper- 
deficient animals; patients have low levels of copper in serum, 
liver, and brain and depressed levels of serum oxidase activity. It 
is believed that the basic defect is the failure to transport 
copper across membranes51.
Wilson's disease is also characterised by low plasma copper 
associated with excessive accumulation of copper in tissues such as 
liver, brain, kidney and cornea. Patients suffer from neurologic 
symptoms, liver cirrhosis and corneal degeneration. Treatment 
involves use of chelating agents such as penicillamine, to aid in 
the elimination of excess copper especially from the central 
nervous system51.
Etiological factors that contribute to copper deficiency 
include general malnutrition, diarrhoea, prolonged feeding with 
milk-based diets, intestinal malabsorption syndromes, inherited 
defects in copper metabolism, prolonged perinatal feeding and 
prematurity34,52 .
Normally, healthy term infants do not appear to be at risk of 
copper deficiency even if fed with relatively low-copper milk 
products since neonatal copper stores are usually adequate to tide 
over the young infant until he is fed other foods rich in copper34.
In contrast, premature infants do not start post-natal life 
with similar copper stores, thus low- birth weight infants are at
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risk from severe symptomatic copper deficiency by three months 
after birth46,52 -
Wilson and Lahey53 reported that a daily intake of 15/xg copper/ 
kg body weight for low birth weight infants during the first two 
months of post-natal life was sufficient to avoid any gross signs 
of copper deficiency; Cordano54 has suggested a level of 90/xg/ kg 
body weight.
2.2.3. Chromium, manganese and iodine
Chromium is essential as a component of nucleic acids and as 
a component of a factor that potentiates the action of 
insulin29,32,55 . Chromium is biologically active when it is complexed 
in a naturally-occurring, organic compound known as glucose 
tolerance factor. In this form, chromium absorption from the small 
intestine and its action as an insulin cofactor are several times 
greater than inorganic trivalent chromium compounds55. Current 
hypothesis concerning mechanism of action of glucose tolerance 
factor is that it facilitates the interaction between insulin and 
its receptors.
Chromium deficiency in man is characterised by impaired 
glucose tolerance, decreased nitrogen retention, ataxia and 
defective peripheral nerve conduction56,57 . Animals fed chromium- 
deficient diet develop glycosuria, fasting hyperglycaemia, corneal 
opacities and aortic plaques56,59 .
Plasma chromium concentration is also lower during pregnancy 
and it has been explained that the mild glucose intolerance during
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pregnancy may be due in part to decreased maternal chromium, since 
the fetus accumulates chromium at the expense of maternal chromium6-
It has been shown that in the US., tissue chromium decreases 
with age and it has also been found that one of the causes of 
glucose intolerance in the elderly is chromium deficiency60. The 
recommended daily allowance has not been established for pregnant 
women nor for newborns6.
Manganese is important for life due to the fact that manganous 
ion is known to be an activator of many enzymes7.
In 1972,the first report of manganese deficiency in man appeared61. 
Until then there was doubt as to whether manganese deficiency could 
occur in man. The symptoms were weight loss, transient dermatitis, 
occasional nausea and vomiting, changes in the colour of hair and 
beard and slow growth of hair and beard61-
Manganese deficiency has also been observed in an adult man in 
association with vitamin K deficiency62. The clotting factor of this 
patient was low until given manganese and vitamin K. In other 
species, manganese deficiency is associated with impaired growth, 
skeletal abnormalities, disturbed reproductive function, ataxia in 
the newborn and defects in lipid and carbohydrate metabolism29, 
which may relate to its role with pyruvate carboxylase7.
Iodine is essential for the synthesis of thyroid hormones that 
are important in cellular oxidation, growth and development,and 
gonadal function33. In conditions of iodine deficiency,a spectrum 
of symptoms referred to as Iodine Deficiency Disorders (IDD) 
occur. These include goitre, endemic cretinism characterised most
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commonly by mental deficiency, deaf-mutism and spastic diplegia, 
and some neurological defects, impaired mental function, increased 
still births, increased perinatal and infant mortality33 -
From the May 1993 conference on iodine deficiency at the 
Franklin Institute, Philadelphia, USA, came the following 
consensus: ' The employment of prevention of iodine deficiency is 
urgent because even a modest degree of it has a deleterious effect 
on cognition and neuromotor functions'. IDD therefore threatens the 
health, wellbeing, social and economic productivity of the 
sufferers.
In 1960, WHO estimated a population of 200 million as 
suffering from IDD, Most recent estimates greatly exceeded this 
figure in spite of extensive iodation programmes. There is a 
consensus that 800 million people are at risk of suffering from IDD 
worldwide33.
IDD is a good example of a major nutritional disorder for 
which the techniques for treatment, control, and prevention are 
easily available and affordable. All that it takes is a strong 
will, wider awareness and cooperation among those who hold the key 
to the solution of the problem.
2.2.4. Zinc
There are forty or more substances known to be essential in 
the human diet, but of these only three micronutrients- vitamin A, 
iron and iodine are thought to be commonly deficient9. There is a
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growing suspicion however that zinc might also be included in this 
category. This is based on two separate considerations. On the one 
hand, the pervasive nature of zinc-dependent enzymes in metabolic 
processes. On the other hand, zinc supplementation is beneficial in 
many disease states; for example, in malnourished children, vitamin 
A status has been improved and immune response corrected, and even 
the duration of diarrhoeal diseases seem to be reduced by zinc 
supplementation9.
The biological importance of zinc was established first in 
plants in 18 6 9 63 . In 1934, Todd et al64 reported that zinc was
necessary for animal life and suggested that this might be the case 
of humans too. This was confirmed by Prasad et al65. Today a number 
of zinc-responsive syndromes are known, many of which reflect 
underlying zinc deficiency states.
2.2.4.1. Distribution of zinc in human body.
In the late forties, McCance and Widdowson9 showed that the 
adult human body contains about two grams of zinc. Sixty percent of 
body zinc is in the muscle, 20% in bone, 5% in blood and liver and 
3% in skin and the gastrointestinal tract.
Scoular and Macy did balance studies in preschool children in 
the early forties to show that five milligram of zinc were retained 
out of an intake of 16mg a day9. Such a retention, five times 
greater than that of iron for example, seemed to speak against the 
classification of zinc as a trace element. This rethinking has been 
made possible with the introduction of atomic absorption
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spectroscopy in the investigation of the importance of zinc in 
human nutrition9 -
Zinc is present in virtually all cells, but certain tissues in 
animals have a higher abundance22. Moreover, zinc distribution among 
tissues is similar in different animal species. Concentrations 
typically are between 10 and 100 /xg/g wet weight22. The zinc 
concentrations of most soft tissues such as muscle, brain, lung,and 
the heart are relatively stable and unresponsive to amounts of 
dietary zinc over most ranges of intake, however zinc 
concentrations in other tissues such as bone, testes, hair, and 
blood tend to reflect dietary zinc intake22.
Zinc is the most abundant trace element inside most cells, 
with the exception of the erythrocytes where iron has its oxygen- 
carrying functions. Even the macro- element calcium is less 
abundant than zinc in all other cells except in bone cells9.
2.2.4.2. Biochemistry and the physiological roles of zinc
The biological importance of zinc appears to be related 
primarily to its role in many enzyme systems. Zinc is not limited 
as are calcium and iron to a few functional cells but is present in 
practically all cells of the body22.
Zinc is a functionally essential component of more than 200 
enzymes9, pervading all metabolic pathways and at least one zinc 
metalloenzyme is known in every major enzyme classification5.
The role of zinc in such enzymes can be either catalytic and/ or 
structural. Enzymes which contain zinc or require adequate zinc
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nutriture in human and/ or experimental systems for activity 
include alkaline phosphatase, alcohol dehydrogenase, carbonic 
anhydrase, lactic dehydrogenase, glutamic dehydrogenase, 
carboxypeptidase, RNA and DNA polymerase, thymidine kinase and 
delta aminolevulinic acid dehydratase66.
Some non-enzyme ligands with which zinc forms complexes 
include alpha2-macroglobulin glycoprotein in plasma which firmly 
binds about 30% of plasma zinc, albumin, which loosely binds about 
66% of plasma zinc and amino acids such as histidine and cysteine 
with which about 2% of circulating zinc is reported to be 
complexed. Zinc can also bind to transferrin, metallothionin and 
nucleoproteins66.
Zinc also helps to stabilize membrane structures and protects 
membrane integrity by the reduction of free radical formation, thus 
preventing lipid peroxidation9
Thus zinc has been shown to be important in a variety of 
physiological processes10'14. In its deficiency, many clinical 
manifestations become apparent. These include anorexia, growth 
retardation, delayed closure of epiphysis, hypogeusia, dysgeusia, 
delayed sexual maturation, alopecia, diarrhoea17. Other symptoms are 
delayed healing of wounds, mental depression and lethargy, 
increased susceptibility to infections, eye lesions such as 
retinitis and congenital malformations17. These symptoms and signs 
are to some extent dependent on age, acuteness of onset, duration 
and severity of zinc depletion and on the cause of zinc 
deficiency17. In the paediatric age however, retarded growth is an
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early, consistent and prominent feature17.
The vital importance of zinc for normal growth and development 
may be explained at least in part by its central role in nucleic 
acid metabolism. Zinc is needed in the synthesis of both DNA and 
RNA and may also have a role in the structure of nucleic acids17. 
Zinc therefore has a functional role in gene replication, 
activation and repression, is critical for transcription and 
translation, and affects nucleic acid metabolism9.
Growth of young rats on a zinc-deficient diet stops within 24 
hours, probably due to the lack of gene regulatory proteins9- These 
proteins contain a common structure "Zinc finger" which are loops 
of chains of amino acids held together at the base by a zinc atom. 
Gene regulatory proteins may contain eleven such "fingers" which 
reach down into the grooves of DNA helix and promote
transcription9.
The critical participation and importance of zinc in the 
synthesis of nucleic acids, proteins and in the structure and
functions of biomembranes account for its roles in immunity and
wound healing66.
Zinc also mediates the activity of growth hormone9. When
growth hormone attaches to its receptor sites on a cell membrane, 
zinc atom is needed to make the connection. The resulting complex 
is called "zinc-sandwich".
2.2.4.3. Zinc deficiency
Due to the critical importance and roles of zinc in metabolic
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processes, all systems of the body can be adversely affected by 
zinc deficiency. This is more so if the deficiency occurs at a time 
when cells of the particular system are rapidly dividing, growing 
or synthesising proteins66. Processes which are profoundly affected 
by experimental zinc deficiency include RNA metabolism, DNA 
metabolism, protein metabolism, and mucopolysaccharide metabolism66. 
These effects of zinc deficiency are reflected in the intact animal 
by poor utilization of dietary nitrogen and sulphur and prompt 
arrest of growth66- In fact it has been suggested that zinc 
deficiency causes a block in protein and nucleic acid synthesis5.
Reproduction is severely affected by zinc deficiency. Gonadal 
maturation is severely retarded in animals and humans. In adult 
animals, atrophy and fibrosis of gonads occur. If severe zinc 
deficiency is induced in rats in early pregnancy, there is a very- 
high incidence of teratology and abortion. If induced later, 
parturition is severely impaired and fetal growth is retarded but 
no gross teratology66.
The teratogenic effects of congenital zinc deficiency on the 
nervous system is particularly pronounced. When deficiency is 
induced during the last trimester, adverse effects on the fetal 
brain are limited to abnormal composition with a decrease in total 
DNA and an increased ratio of protein to DNA66. When such rats are 
raised to adulthood, the males have decreased tolerance to stress; 
the females show increased aggression. Thus zinc deficiency during 
critical period of brain growth and development appears to cause 
irreversible damage which adversely affects behaviour in adult
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life. It is unknown whether this effect holds true in humans66.
The effect of zinc deficiency on carbohydrate metabolism is 
less clear. However,it has been noticed that in zinc deficiency, 
there is intolerance to parenteral glucose in rats and humans, 
while the oxidation of glucose does not seem to be impaired. It has 
also been suggested that zinc in some ways participate in the 
uptake of glucose by cells66.
It has been shown that experimental zinc deficiency is 
associated with decreased synthesis of retinol-binding protein 
(RBP) and release of vitamin A from its hepatic stores. Such 
mechanisms could explain low levels of vitamin A and RBP in 
alcoholics with liver diseases67.
In experimental zinc deficiency, the activity of zinc 
metalloenzymes is decreased in certain tissues, but not in others. 
For example, a decrease in serum alkaline phosphate activity has 
been used clinically to diagnose zinc deficiency in humans68.
Another system particularly sensitive to zinc deficiency is 
the integument66. Zinc-deficient swine develop dermatitis with 
abnormally high hyaluronic acid content of the skin. Zinc-deficient 
rats also develop dermatitis and alopecia (baldness). 
Histologically, hair follicles are decreased and collagenous. 
Subcutaneous tissues appear less dense. Electron microscopic 
examination of these tissues show abnormal appearance of collagen 
and fibroblasts while chemical studies of collagen suggest impaired 
synthesis and cross-linking, impaired incorporation of methionine 
into proteins and of thymidine into DNA, decreased thymidine kinase
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activity in collagen and abnormal RNA metabolism. The gross effects 
of these abnormalities is impaired wound-healing. The clinical 
equivalents of these animal abnormalities in humans include 
impaired wound-healing, acrodermatitis enteropathica, and 
dermatitis in infants alimentated parenterally without adequate 
zinc66.
Zinc deficiency also has adverse effects on the gustatory 
system. In rats, a loss of taste acuity, as measured by preference 
for sodium chloride, occurs within three days and anorexia is 
evident in two days. Abnormal taste and smell acuity, responsive to 
zinc supplementation, have also been described in humans. In 
humans, however, abnormal taste and smell can occur as a result of 
other unrelated stresses66.
2.2.4.4. Human zinc nutrition and deficiency.
Etiological factors:
The etiological factors responsible for human zinc deficiency 
have been identified and these are presented in 
table 217:
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Table 2. Etiological factors in human deficiency17
State Cause
Inadequate dietary intake Diets low in animal protein, 
zinc lost in food processing.
Dietary factors inhibiting 
zinc absorption
Phytic acid, fibre.
Disease states inhibiting 
zinc absorption
Steatorrhoea ( cf. some body 
zinc is bound to fat 
'Acrodermatitis enteropathica
Genetic defects in zinc 
metabolism
Acrodermatitis enteropathica
Excessive loss of zinc Hyperzincuria eg.in chronic 
alcoholics, diaphoresis, 
chronic blood loss.
Increased zinc requirement Rapid growth, pregnancy and 
lactation, wound healing. !
It now appears that a marginal zinc deficiency may be quite 
common in paediatric populations especially in 0 to 6 month
babies. It also occurs in infants and children who are ingesting 
adequate quantities of other nutrients66, since the risk of zinc
A lethal inherited zinc deficiency which results in 
eczematoid skin lesions, diarrhoea, low plasma zinc levels, poor 
growth and development, malnutrition, intercurrent bacterial and 
yeast infections and eventual death if left untreated. It usually 
shows up in infancy upon changing from breast milk to cow's milk. 
It has been postulated that cow milk contains a peptide that these 
children can not digest and that chelates zinc and prevents its 
absorption69. The treatment is to provide zinc sulphate in amounts 
large enough to overcome this chelating effects and allow zinc 
absorption. Neldner and Hambidge70 found that zinc supplementation 
at a level of 22 mg of elemental zinc was enough to cause remission 
of the disease in an adult.
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deficiency is enhanced at the time of increased requirement such as 
periods of rapid growth, pregnancy and lactation.
In rural areas of Egypt and Iran where zinc deficiency was 
first identified, dietary factors, especially phytic acid and 
fibre,that impair zinc absorption, were the major etiological 
factors17. Studies of rats support this concept66.
Other pathogenic factors that contributed to zinc deficiency 
are hookworm and schistosomiasis in Egypt and geophagia in Iran and 
Turkey. High environmental temperatures may be another factor since 
sweat is rich in zinc- as much as 1 mg. per liter66.
Conditioned zinc deficiency appears to be a potential problem 
in patients with alcoholism and/ or cirrhosis, patients with 
chronic renal disease , patients with severe trauma, patients with 
chronic infections or inflammatory diseases of the bowel and 
malabsorption syndrome. The causes of the zinc deficiency are 
multiple. They include inadequate intake of zinc in diets or 
parenteral fluids as well as increased losses from the body in 
association with catabolism, protein deprivation, and PEM in 
infants66.
Other potential causes include abnormal loss of zinc due to 
proteinuria or aminoaciduria and impaired intestinal absorption66. 
It appears that the rapidity of onset of symptoms on zinc-deficient 
diet is due to the fact that there is no functional body store of 
zinc since virtually all body store is locked up in bone or 
protein5.
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Zinc and foetus
Animal studies have emphasised the nutritional importance of 
adequate maternal zinc nutrition for normal growth and development 
of the foetus. The Food and Nutrition Board of the National Academy 
of Science, USA, has recommended a daily intake of 2 0mg zinc during 
pregnancy or about 5 mg more than for non-pregnant adult17* It has 
been found that human fetal size correlated with zinc 
concentrations in amniotic fluid.
A teratogenic role for zinc deficiency in humans has also been 
suggested on account of high incidence of congenital malformations 
in the offsprings of women with acrodermatitis enteropathica. In 
fact the striking effects of experimental zinc deficiency in rats 
prompted speculations that the greater incidence of congenital 
abnormalities in certain developing countries is due to habitual 
consumption of diets low in available zinc17.
Studies on zinc nutrition in pregnancy are hampered by lack of 
established biochemical indices of zinc status. Plasma zinc levels 
are normally depressed during pregnancy due to raised levels of 
circulating estrogens17- It is also known that the causes of 
maternal mortality in developing countries include excessive 
bleeding and hypertension, both of which are associated with zinc 
deficiency66.
Zinc and the infants.
In well nourished populations,neonatal tissue concentrations 
of zinc are similar to those of adult even though the young infant
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does not have stores of zinc in contrast to those of iron and 
copper. The Food and Nutrition Board recommended a daily dietary 
intake of 3 mg zinc from 0-6 months, and 5 mg from 7-12 months17.
There is reason for concern that zinc requirements may not be 
met adequately in formula-fed infants, at least after the first two 
months or so of lactation. This is because most of the zinc in milk 
is associated with the protein fraction. Thus when the protein 
content in cow milk is reduced during the preparation of infant 
milk formulas, to approximate that of human milk,there is a 
corresponding reduction in zinc content. Hence the final zinc 
concentration of these formulas are generally less than 2mg/L 
unless supplemented with additional zinc71.
It has been shown that zinc levels of breast-fed infants were 
significantly greater than those of formula-fed infants even when 
the concentration of zinc in the formulas was about three times 
that of breast milk71. Zinc in infant formulas is added as inorganic 
salts while in breast milk the zinc is bound mostly to high 
molecular weight proteins; thus it is not only the amount of zinc 
but also the molecular localisation of zinc in milk that affects 
the extent of absorption71.
Zinc deficiency in infants may also occur in association with 
protein-energy malnutrition. Hypozincemia in the acute state may be 
due to hypoproteinemia and to superimposed infections17.
Zinc and preschool children
This is an age group that both merits and has received
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considerable priority in nutrition surveys and nutrition 
intervention programmes but it is only recently that studies have 
included biochemical indices of zinc nutritional status.
It has been observed that the levels of zinc in plasma and 
hair in preschoolers in some countries such as USA were low as 
compared to those of older children and adults72. Though these lower 
levels may be in part physiological at a time of rapid growth, 
similar low levels had not been observed in countries of Europe and 
Far East73. One factor responsible for the low zinc in American 
children may be the low zinc content of some popular infant 
formulas, prior to their recent supplementation with zinc through 
legislation.The majority of the children with low zinc status were 
also of low height percentile.
The reduced zinc intake particularly among growing children is 
accompanied by low hair zinc content, anorexia, stunting and 
lowered taste acuity20. Functional disorders appear in children with 
hair zinc content below 70 ppm20. It has also been suggested that 
plasma zinc levels below 70 fig per 100 mL indicate hypozincemia67.
Results of study on zinc nutrition of preschool children in 
Denver Head Start Programme indicated that inadequate zinc 
nutrition may be common among preschool children from low income 
families16.
Zinc in School children and adolescents.
Many studies have revealed an association between low zinc 
levels and low growth percentiles in school children considered as
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healthy. Some of these children had impaired taste acuity. Dietary 
supplementation with 0 .2 mg zinc per kg body weight per day was 
followed within a few months by normalisation of taste perception 
and a suboptimal increases in hair zinc content. Thus it appears 
that suboptimal zinc deficiency may not be uncommon in otherwise 
healthy school-age children16,72.
The first report of human zinc deficiency was related to the 
syndrome of adolescent nutritional dwarfism in Egypt and Iran. 
Since then, additional studies have identified marginal zinc 
deficiency in preschool and school children in several 
countries20,71,74,75 . The main features of this syndrome were growth 
retardation, and failure of sexual maturation.
Numerous tests have shown that these adolescents were zinc- 
deficient and zinc therapy under controlled conditions led to the 
conclusion that zinc deficiency was a major factor responsible for 
retarded growth and delayed sexual maturation. Other features of 
zinc depletion included anorexia, lethargy, dry and roughened 
skin17.
It is possible that a variety of factors are responsible for 
zinc deficiency states in different areas of the world. In Iran, 
the major factor is the large quantities of phytic acid and fibre 
present in the unleavened bread that is a major dietary staple in 
rural areas. These dietary components inhibit zinc absorption 
through formation of insoluble zinc phytate17.
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Zinc and the adult.
Once growth and development are complete, zinc requirements 
are comparatively lower and the effects of zinc deficiency, except 
during pregnancy, are likely to be less severe17- Nevertheless, a 
number of zinc-responsive syndromes have been reported in adults . 
For example, zinc supplementation has been widely used in surgical 
patients to promote wound healing. Conflicting reports on the 
efficacy of such therapy are probably due to the fact that the 
patients were not selected on the basis of their zinc status17.
Wound healing is impaired in the zinc-deficient animals and 
in humans with low plasma levels and healing is accelerated with 
zinc therapy. Similarly improvements in the rate of healing of 
venous leg ulcers have been observed with zinc therapy, but only in 
patients with hypozincuria. Zinc therapy has also been reported to 
accelerate the rate of healing of benign gastric ulcers17.
In a study of the zinc status of elderly black Americans from 
urban low-income house-holds, Wagner et al found that eleven 
percent of 135 subjects had hair zinc levels less or equal to 70 
/xg/g or serum zinc concentration less or equal to 70/ig/g and 
suggested that the zinc status of this elderly population may be 
less than ideal76 -
In summary , it is apparent that zinc is not only essential, 
but because it is involved in so many important physiological 
processes, it may even be the "first limiting factor" in these 
processes. This means that it could be the critical limiting factor 
in the diet and the consequences of zinc deficiency are likely to
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be extensive, if not catastrophic for the organism.
2.2.4.5. Zinc Toxicity
Although the consequences of zinc deficiency have been 
recognised for many years, it is recently that attention has been 
directed to the potential consequences of excessive zinc intakes.
Zinc is considered to be relatively non-toxic, particularly if 
taken orally; however, manifestations of overt toxicity symptoms 
such as nausea, vomiting, epigastric pain, lethargy and fatigue 
will occur in high intakes77. At low intakes, but at concentrations 
well in excess of RDA ( ie. 100-300 mg zinc/day vrs.an RDA of 15 
mg/day) evidence of induced copper deficiency with attendant 
symptoms of anaemia and neutropenia as well as impaired immune 
functions and elevated ratio of high density lipoprotein(HDL) to 
low density lipoprotein(LDL) have been reported77. Even low levels 
of supplementation, close to RDA, have been suggested to interfere 
with the utilization of copper and iron and to adversely increase 
HDL77.
It has also been shown that the form of zinc ingested may 
influence the ensuing symptoms. For example, an emetic dose of 
l-2g. zinc sulphate (corresponding to 225-450mg elemental zinc) 
caused nausea, vomiting, epigastric pain, abdominal cramps and 
diarrhoea or dysentery while ingestion of 12g of elemental zinc 
produced lethargy,light-headedness, slight staggering of gait and 
difficulty of writing; all symptoms disappearing with chelating 
therapy; interestingly, no gastrointestinal symptoms were
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reported77.
2.2.4.6. Assessment of zinc status in man
Several analytical methods involving the use of flame or 
flameless atomic absorption spectrometry have been used to 
determine body zinc status from biological tissues and fluids such 
as plasma, serum, whole blood, erythrocytes, urine, hair,and 
saliva78'82.
1. Plasma and serum: Plasma and serum have been most widely used 
samples, but such results are greatly affected by various states 
such as acute infection, stress, myocardial infarction, and 
physical exertion; thus a low plasma or serum zinc concentration 
per se, does not necessarily reflect the actual body zinc 
status83'85. It is therefore desirable to carry out determinations on 
a variety of biological tissues or fluids such as plasma or serum, 
erythrocytes, hair and urine to arrive at the zinc status in an 
organism84.
2. Erythrocytes and hair: The turnover of zinc in erythrocytes and 
hair is slow and as such lowered values in these materials would be 
indicative of a long-term deficient state. On the other hand the 
turnover in plasma or serum is relatively fast hence a low value 
here would reflect a current zinc deficiency , but as explained 
earlier, the level is affected by a variety of factors84.
3. Neutrophils: Neutrophils have a high zinc content and a short 
life-span, so acute zinc status might be expected to be reflected 
more promptly in these cells84.
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4. Urine zinc: A decreased excretion of zinc in urine appears to 
be a valuable indicator of zinc deficiency in man; however, 
hypozincuria may be indicated in other conditions such as liver 
cirrhosis and in sickle cell disease85.
5. Metabolic balance studies: Measurements such as metabolic 
balance study, turnover rates and 24-hour exchangeable pool for 
zinc using zinc-65 may provide additional tools for assessing zinc 
status in man17.
Zinc balance studies also provide a good basis for assessing 
the zinc status. A positive retention of zinc would be indicative 
of zinc deficiency. Using zinc-65, it was shown that in zinc 
deficiency, plasma turnover was increased, the 24-hour exchangeable 
pool was decreased and the cumulative excretion of zinc-65 in urine 
and stool was low85.
6 . Zinc metalloenzymes: Certain zinc metalloenzymes in blood may 
be used for assessment of zinc status in man. The activity of 
Alkaline phosphatase was found to be significantly decreased in 
serum or plasma of zinc-deficient rats, pigs, dairy cows, calves, 
chicks and swine05. Consequently, serum Alkaline phosphatase 
activity has been shown to increase following zinc supplementation 
to zinc-deficient dwarfs in the middle East8.
7. Carbonic anhydrase: Studies of sickle cell disease subjects 
revealed that a significant number of these patients are zinc 
deficient and show an increased activity of ribonuclease in the 
plasma86.
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8 . Plasma ribonuclease: Studies of sickle cell patients revealed 
that a significant number of these are zinc deficient and show an 
increased activity of ribonuclease in the plasma. Zinc is an 
inhibitor of ribonuclease and an increased activity of this enzyme 
has been observed in zinc-deficient tissues of experimental 
animals; thus, measurements of plasma ribonuclease may provide 
another parameter for assessing zinc status in man86.
9. Urinary sulphate excretion following an ingestion of S35- 
cysteine to zinc-deficient rats is enhanced due to the fact that 
rats utilise sulphur-containing amino acids for proteosynthesis87. 
It remains to be tested whether such an approach will be fruitful 
in man87.
2.2.4.7. Estimation of zinc concentration
Zinc has been determined colorimetrically using dithizone88,89 
and fluorimetrically as a 8-hydroxyquinoline complex stabilised by 
gum arabic at ph 8 with activation at 375nm and emission at 517nm90.
The method of choice now is atomic absorption spectrometry91,92 
which has several advantages, such as, simplicity, increased 
sensitivity, and accuracy, over the old traditional methods. A 
characteristic absorption at 213.8nm provides sensitivity to < 1.5 
limol/ L ( O.lmg/L) with excellent accuracy and precision22. Also, 
provided the instrument is fitted with a high solid burner such as 
the Boling 3 slot burner which allows direct aspiration of high 
solid-containing samples without clogging, determination can be 
carried out directly on suitably diluted samples of serum or urine.
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The increased viscosity of the test compared to the aqueous 
standards can cause some matrix problems. Parker et al. , and 
Hackley et al., overcame this problem by preparing standards in 
albumin (40g/L)91 and dextran (30g/L)92, respectively.
Preparation of other samples for analyses involves combustion 
of organic material (wet or dry ashing) ,followed by solubilization 
of the ash in acid. Cells and organelles can be hydrolysed in base 
such as 0.2 mol. NaoH/L before atomic absorption analysis22.
Fluids are usually diluted with water or dilute acids to 
appropriate concentrations, without ashing, before analysis. X-ray 
emission spectroscopy is gaining increased attention as a semi- 
quantitative method to subcellular zinc localization and 
measurement22. Atomic absorption spectrophotometry has been used in 
the present work to determine zinc concentration in hair, 
erythrocytes and plasma.
2.2.5. Variables to be measured and justification for their choice
2.2.5.1. Variables
i. hair zinc
ii. alkaline phosphatase activity
iii. total plasma protein
iv. albumin
v. plasma and erythrocyte zinc
2.2. 5.2. Justification
Right now, there is no single laboratory measurement that
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reliably reflects zinc status. However, it has been suggested that 
the determination of the zinc concentration in two or more tissues, 
such as serum, plasma, erythrocytes, hair and/ or urine should 
provide a better data-base from which zinc nutriture can be 
evaluated93, 94 .
(i). Zinc statue based on hair zinc erythrocyte and plasma zinc 
analysis:
The use of hair for the evaluation of human zinc nutriture 
remains somewhat controversial94'96. It has been suggested that 
contamination from exogenous sources94, beauty treatments95, 96 , and 
variable hair growth rates, may complicate the interpretation of 
hair zinc data. However, evidence from animal studies demonstrates 
that hair zinc concentration reflects chronic or long-term zinc 
nutriture and that it correlates with the zinc content of bone and 
diet9,97.
Flynn et al98., have emphasised that hair can be very useful 
in nutritional assessment if attention is paid to proper sampling 
techniques and appropriate statistical analysis.
It is generally agreed that homogeneity in the distance of the 
sample from the scalp is an important factor and that the hair 
closest to the scalp should be sampled for zinc analysis. This 
newly-grown hair is presumably more reflective of current zinc 
metabolic status and is less subject to contamination from 
exogenous sources98.
Since grease, dust and other foreign bodies on the surface of
37
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hair may contribute most of the zinc of hairs, the hair must be 
washed. Even though the importance of a standardised washing 
procedure is widely acknowledged, different laboratories use 
different washing procedures and there is no consensus on how the 
washing should be done". For example, the following washing 
solutions with variable volumes and washing times, have been used: 
distilled water, ionic or nonionic detergents, combinations of 
aqueous detergents and organic solvents, chelating agents and even 
mineral acids". The effects of these washing agents/ procedures 
have been stated to differ. Moreover, mammalian hair may continue 
to loose material for an indefinite number of washes. Thus the 
problem in hair analysis is not the reliability of the measurements 
but preparation of the sample ie. sampling, washing agents, length 
(time) of washing, and number of successive washings. These could 
be the source of discrepancy between results of various workers".
The use of scalp hair has the advantage of non-invasiveness, 
ease of collection, preservation and transport ( including airmail 
postage to distant specialised laboratories)".
A depressed level of circulating zinc in plasma or serum has been 
associated with human zinc deficiency16,100,101, but circulating zinc 
levels are not always a reliable index of zinc nutriture93,94,100 . One 
must rule out any haemolysis, hypoproteinemia, heritable 
hyperzincemia, or redistribution of zinc from plasma to tissues or 
from tissues to plasma, in order for this parameter to be an 
accurate indicator of zinc status93, 94 . Due to the high zinc content 
of the erythrocytes, any slight haemolysis would add zinc to serum.
38
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Beside, numerous factors, as indicated earlier are known to 
influence the concentration of zinc in plasma and for that matter 
serum94, 102"105. Despite these drawbacks, serum/plasma zinc and hair 
analysis are the indicators most often employed in zinc status 
assessments16.
Whitehouse et al84,have proposed that zinc in other tissues 
such as erythrocytes and leucocytes may more closely reflect body- 
zinc status. Since erythrocytes have an average lifespan of 120 
days, data on zinc concentrations in erythrocytes have mainly been 
useful for indicating long-term zinc status.
(ii). Plasma total protein and albumin:
Zinc in plasma is distributed between two major fractions- 
about 60% are loosely bound to albumin and the rest firmly bound to 
globulin and other cellular components.
It has been shown that as a result of modifications of cow's 
milk to produce infant formulas in which the original high protein 
level is reduced to equivalent amounts present in human breast 
milk, these infant formulas become zinc-deficient. In animal 
tissues, zinc is carried as a zinc-protein complex, so any 
deficiency of protein could affect the amount or availability of 
zinc to tissues. Indeed, work by Lonnerdal et al.104, revealed that 
in human milk, 14% of zinc is complexed with casein, 28% with serum 
albumin, 29% is associated with low molecular weight proteins such 
as metallothionein105, while the rest, that is 29%, is associated 
with fat.
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Following oral administration of zinc-65, the isotope is 
associated exclusively with albumin which suggests that albumin 
transports zinc from the intestinal cells17. Thus any deficiency of 
protein and/ or albumin could lead to zinc deficiency.
It is therefore important to measure plasma proteins and albumin 
levels in any studies on zinc nutriture. The determination of 
albumin/globulin ratios will also give an indication of the protein 
nutrition status of the subjects.
(iii)• Alkaline phosphatase activity:
Many studies have shown that serum Alkaline phosphatase 
activity responds rapidly to zinc depletion through a significant 
reduction of its activity17,106. In the studies by Roth and 
Kirchgessner106, this enzyme lost 27% and 48% of its activity within 
two and four days, respectively, after the zinc-deficient diet was 
initiated.
In limited studies, however, either there was no reduction in 
activity or there was no significant difference in the enzyme 
activity in zinc-deficient or zinc-supplemented pair-fed control 
rats107'108.
An accurate determination of Alkaline phosphatase activity may 
therefore give invaluable clue about the zinc nutritional status of 
the cohorts. It must be noted that alkaline phosphatase activity 
can be affected by many factors unrelated to zinc status.
40
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CHAPTER THREE
METHODOLOGY
3. Study design and plan of work.
3.1. Study population:
Children attending nursery/kindergarten school in four 
communities: Kwabenya village, Ashalley Botwe village, Dome and New 
Achimota, constituted the reference population. These villages were 
chosen because they are easily accessible by road. Most of the 
target group attend the nursery schools in which the studies were 
carried out. These conditions made the project manageable within 
the limit of the available resources.
3.2. Subjects selected for the study:
Children aged 3 to 5 years attending nursery/ kindergarten in 
the four communities were recruited for the study. All the children 
were used since none showed any overt signs of infection such as 
pneumonia, measles, or upper respiratory tract infection.
Informed consent of chiefs/assembly men/parents/ 
guardians/teachers were sought for children participating in the 
study.
3.3. Study areas:
The study was carried out in the nursery/ kindergarten schools 
in the four communities with different socio-economic status, 
judged by the number of sandcrete houses, aluminium/zinc-roofed 
houses and infracstructural developments such as places of
41
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convenience, and potable water. This was to determine if zinc 
status was dependent on socio-economic status. Accordingly, two of 
these communities ( Ashalley Botwe and Kwabenya) are typical 
villages; Dome is peri-urban while New Achimota is almost urban.
The socio-economic status can therefore be ranked as follows: 
New Achimota > Dome > Ashalley Botwe > Kwabenya village.
The estimated total population of the 4 communities was about 
3500 of which about 15% are within the age group of 3-5 years. 
Depending on the community, the adult inhabitants ( ie, parents/
guardians of the cohorts) are either farmers, artisans, taxi
drivers, teachers, architects, engineers, and medical officers, who 
work in Accra. There are no health centres in the first three 
communities (ie. Ashalley Botwe, Kwabenya, Dome); the inhabitants 
depend on native doctors as well as the clinics at the Ghana Atomic 
Energy Commission, Madina or Legon.
3.4. Plan of work:
The correct ages of the subjects were mostly ascertained from 
health or weighing cards ( 99%) or from mothers' interview(l%).
Children who fell within the ages of 3-5 years, were recruited for 
the study:
3.4.1. Anthropometric measurements.
Standing heights were measured, using a stadiometer. Height was 
measured twice the second measurement being after repositioning 
of the subjects.
Weights were measured using a bathroom scale. Scales were 
calibrated daily with a 10kg standard weight and zeroed before each
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measurement, when necessary. Mid-upper arm circumference (MUAC) was 
measured ( to the nearest 0.1 cm) on the left arm using tapes. The 
tape measure used was a non-stretchable polyvinyl-coated 
fibreglass. Triceps and subscapular skin fold thicknesses (in 
duplicate to the nearest 0 .1mm) on the left arm were measured with 
Harpenden skinfolds calipers. Triceps skinfolds and MUAC were both 
measured half-way down the left arm between the tip of the acranial 
process of the scapular and the olecranon process of the ulna.
3.4.2. Sampling of hair:
Hair samples (about 200-400mg) were cut from all the 
participating children for analysis for hair zinc concentration.
3.4.3. Blood collection:
Five (5) mL of blood were collected via venipuncture of all 
the participating children for the following analyses: plasma total 
protein, albumin, alkaline phosphatase activity, erythrocyte and 
plasma zinc.
3.5. Laboratory analyses:
3.5.1. Determination of hair zinc
3. 5.1.1. Sampling of hair:
Head hair was combed out and the top cut off uniformly until 
it was easy to comb using a selected gauge of fine comb. About 
200- 4 00mg of the hair were then cut close to the scalp, in the 
occipital region, onto white sheets of paper (using a pair of 
stainless steel scissors), and collected into thin plastic bags, 
labelled, sealed, and brought back to the laboratory. Samples were
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stored in a desiccator until processed for analysis.
3.5.1.2. Washing of Hair
The hair samples were individually treated as follows109: 
Samples were introduced into labelled 100 mL acid-washed plastic 
containers. Hexane (100 mL) was added and tubes transferred to 
mechanical flask shaker( Yamato shaker model SA-31), and shaken for 
2 0 minutes, using the highest speed. After pouring out the hexane 
washings, the hair was washed in succession with 100 mL of 95% 
ethanol and three changes of reagent water. The water was an ultra 
pure deionised water obtained by passing tap water through 
millepore filters. The hair samples were then removed onto filter 
papers in acid-washed plastic petri dishes , blotted with acid- 
washed filter paper and air- dried at room temperature for 30 mins. 
The samples were finally dried in an oven at 70°C for 10 mins. 
After cooling in a desiccator over silica gel , the samples were 
transferred into thin plastic bags, labelled and stored in a 
desiccator until used for assay.
3.5.1.3. Wet-ashing of hair and analysis by AAS:
Dried hair samples, weighing between 15 to 15.7 mg, were 
weighed for each sample in duplicate into acid-washed test- tubes 
and 1.5mL of a digest mixture (of 1 part of 60-70% perchloric acid 
and two parts of 3 0% hydrogen peroxide ) added and samples placed 
in an oven at 70°C for one hour, to solubilise the hair110. After 
cooling, 5 mL of reagent water were added, vortex-mixed and stored 
at 4°C until used for zinc analysis by atomic absorption
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spectrometry (Shimadzu Atomic Absorption/ Flame emission 
spectrophotometer AA-630-12: AAS) at the following settings: 
Wavelength =213.9 nm 
Slit width: 1.9 A"
Acetylene gas flow rate: 10 L min . "1 
Air flow rate: 2.4 L min . " 1 
Burner height: 4 mm.
All the necessary precautions, including an overnight soaking 
of containers in 2M hydrochloric acid, use of special deionised 
water and preferential use of plastic containers to glass 
containers, were taken.
3.5.1.4. Quantification of zinc:
A calibration curve was prepared from a commercial zinc 
standard (lOOOppm) containing 0, 20, 40, 60, 80 and 100 fig Zn/ 100 
mL in perchloric acid/hydrogen peroxide mixture. The best line was 
fitted by regression and used to calculate hair zinc concentration. 
Each determination was in duplicate on two samples of the same hair 
for each child. The coefficient of variation (CV) was determined by 
analyzing 10 replicates of the same hair sample.
3.5.2. Determination of plasma and erythrocyte zinc:
3.5.2.1. Preparation of plasma and erythrocytes84
Blood (5 mL) was withdrawn by venipuncture with a disposable
needle(Nipro;21G x 1.5 ) fitted to polypropylene syringe (10 mL) .
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The needle was removed and blood introduced into acid-washed 
polypropylene tubes, containing 60 units of zinc-free sodium 
heparin. The tube was inverted gently several times to mix and then 
placed on ice and transported to the laboratory, where it was 
centrifuged at 2250 x g for 15 minutes at 4°C.
The plasma was transferred into 2 mL polypropylene tubes using 
acid-washed Pasteur pipette, capped, and promptly frozen at 
-30-40°C. The erythrocytes were resuspended in 10 mL of 9g/L cold 
saline and stored at 4°C for subsequent washing, the next day.
3.5.2.2. Erythrocyte zinc analysis:
The packed erythrocytes were washed three times by 
resuspending the cells in 10 mL of cold 9 g/L saline and 
centrifuging (2250 x g for 15 min. at 4°C) . After each 
centrifugation, the saline was aspirated with pasteur pipette and 
10 mL of fresh cold saline added and cells resuspended by gentle 
inversion several times84. After the final washing, the tubes were 
flicked gently about four times. A volume (200 ptL) of washed cells 
was wet- ashed in 6 mL of the digest mixture for 1 hr. at 70°C. 
Thereafter, the clear solution was transferred into acid-washed 10 
mL volumetric flask, and the volume made up to the mark with the 
reagent water and used for zinc determination using Shimadzu Atomic 
Absorption Spectrometer, as in 3.5.1.3. Triplicate determinations 
were made as well as the coefficient of variation.
3.5.3. Determination of Plasma zinc levels.
Plasma was diluted with distilled water in ratio of 1:4 and 
the zinc levels determined using AAS as outlined earlier. A
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standard graph was prepared as before with the exception that zinc 
solutions were made up in 5% glycerol,to cater for matrix effect76. 
Duplicate determinations were made.
3.5.4. Determination of Plasma total proteins
This was done using Biuret reagent method of Weischselbaum111. 
Determination were done in triplicate using the following scheme:
blank standard test 
Biuret reagent(mL) 4.0 4.0m 4.0
Water 1.0
Plasma(1:9 dilution) - 1.0
Standard protein-BSA 1.0
Total volume (mL) 5.0 5.0 5.0
The content of each tube was mixed using vortex mixer and 
allowed to stand at room temperature, with occasional mixing, for 
3 0 minutes and the optical density measured at 55 0nm using a 
Shimadzu spectrophotometer against the reagent blank, which was 
used to zero the instrument. Determinations were carried out in 
triplicate.
A standard graph was drawn from measurements of solutions 
containing 2.0, 4.0, 6.0, 8.0, and 10.0 mg BSA/mL and the best line 
fitted by regressing analysis and then used to compute the 
concentration of the test plasma. Percentage recoveries and CV were 
determined.
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3.5.5. Albumin determination
This was done using Bromo Cresol Green method ( BCG)112:
Plasma (25/zL) was added to 5mL of the working BCG dye solution, 
mixed and the optical density measured within 30 seconds, at 63 0nm 
using a Shimadzu spectrophotometer against a reagent blank. 
Standard graph using BSA (1.0, 1.5, 2.0, 2.5, 3.0, 3.5, and
4.0g/dL) was prepared and best line fitted by regressing equation 
and used to compute albumin levels of the test plasma. 
Determinations were carried out in triplicate. The percentage 
recovery as well as the CV, were determined.
3.5.6. Measurement of Alkaline phosphatase activity (ALP):
This was determined using the recommended method of the German 
Society on enzymes36. The conditions of reaction were:
Buffer: Diethanolamine(DEA), 1.0 mmol L ' 1 
ph: 9.8
Reaction temperature: 30 + 1°C
Substrate concentration: p-Nitrophenol phosphate, disodium salt,
(p-NPP) , 10 mmol L' 1
Mg+2 : 0.5 mmol L ' 1 
Reaction volume: 3.02 7 mL 
Fraction volume: 0.009
Plasma was removed from storage and kept overnight at 4°C for 
10- 12 h. to reactivate alkaline phosphatase113. Three (3) mL of 
freshly-prepared substrate buffer, prewarmed to 30°C were 
introduced into a cuvette and used to zero the spectrophotometer
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set at 4 05 nm.
Plasma (27 /xL) was added and quickly mixed by placing 
cellophane seal at the mouth of the cuvette and inverting it for 
about 4 times. A timer( stopwatch) was simultaneously started. The 
outside of the cuvette was quickly wiped dry and then returned into 
the spectrophotometer. The absorbance was immediately taken and 
then at 30 seconds interval for a total of six minutes.
For the sample blank, 3 mL of the neat buffer and 27 /xL of 
plasma were mixed in a cuvette and the absorbance measured. For the 
solvent blank, the absorbance of substrate-buffer and 27 fib of 
deionised water was measured. Duplicate measurements were initially 
made( for about thirty samples), but since it was found out that 
there were no significant difference between the readings, single 
determinations were made, thereafter. This was also necessitated by 
the fact that the quantities of p-NP, p-NPP and DEA, which had to 
be borrowed, were limited. The cv was measured as before. 
Calculations:
Catalytic activity of ALP ( U L"1 ) ,
V
----------------- x ( &A/j\t )  1
e x 1 x v
where V is reaction volume (L),e is the micromolar 
absorbtivity ( L x t^mol' 1 x cm'1) , 1 is pathlength of 
cuvette (cm), v is sample volume (L),^A is the 
change in absorbance, and t is the reaction time (min).
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Since the micro molar absorptivity must be determined in each 
laboratory in order to obtain good results114, the following 
procedures were used115:
p-Nitrophenol was crystallised from boiling water, air-dried 
at room temperature for 3 0 min. and then stored in a desiccator 
over silica gel for a week to obtain dry crystals.
1 mmol/L solution of p-nitrophenol(pNP) was prepared by dissolving
139.1 mg of dried pNp in distilled water and making up the volume 
to 1L with distilled water. Twenty (20) mL of this solution were 
transferred into a 500mL volumetric flask and the volume made up 
with 10 mmol L"1 sodium hydroxide solution. A second solution was 
made using DEA buffer to make up the volume, just for comparison. 
The absorbance of the two solutions were measured at 405nm using 
either neat sodium hydroxide or neat DEA as blank.
3.5.7 Statistical analysis of results.
Lotus 123 was used to calculate the values of all the biochemical 
parameters measured using standard graphs. This was done for 
protein, albumin, plasma zinc, hair zinc and rbc zinc.
Data were entered into the computer using D-base. For the 
anthropometric classification of the cohorts, WHO/NCHS 
EPINFO/EPINUTR. programme was used to classify the cohorts into the 
various nutritional states. Thereafter, SPSS statistical package 
was used to calculate the various values of the parameters 
measured, and then used to group these into the appropriate 
nutritional states.
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Multiple range (comparison) tests , using Duncan's and LSD methods 
were used to compare means. SPSS packages were used to derive 
regression equations as well as establishing correlation 
coefficients. Harvard graphics (hg3) was used to draw the bar 
(histograms) as well as the line graphs.
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CHAPTER FOUR
RESULTS 
4.1. Age distribution of the cohorts
The age distribution of the cohorts used in the present study is 
depicted in figure la. Of the 200 cohorts, 3% (6/2 00) were below 
36mo; 20% (40/200) were 36-47mo; 44.5% (89/200) were 48-60mo; while 
the rest, 32.5% (65/200) were above 60mo., but below 72mo.
4.2. Anthropometric classification of cohorts:
The basis of classification of the cohorts was as follows: 
Normal= weight-for height and height-for Age are both > -2SD of the 
NCHS reference standards; Wasted= weight-for-height is < -2SD of 
the reference standards; Stunted= Height-for age < -2SD of 
reference standards; Wasted plus stunted= weight-for-height and 
height-age are both <-2SD of reference standards. This scheme is 
similar to the classification scheme proposed by Waterlow116. The 
results are indicated in Table 1 ( see appendix).
Of the 200 cohorts screened, 78.5% (157/200) were of a normal 
nutritional status, 3.5% (7/200) wasted, while 16.5% (33/200) were 
stunted and 1.5% (3/200) both wasted and stunted.
The breakdown by communities indicated that 69.6% (39/56),
81.2% (26/32), 80.9% (55/68) and 84.1% (37/44) of the cohorts in 
Ashaley Botwe, Kwabenya, Dome and New Achimota, respectively had 
normal nutrition. In the same sequence, 25% (14/56), 18.8% (6/32), 
13.2% (9/68) and 9.1% (4/44) were stunted, while 3.6% (2/56), 0%, 
4.4% (3/68) and 4.5% (2/44) were wasted. In Ashalley Botwe, 1.8%
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(1/56) of the children were wasted plus stunted; the corresponding 
figures for Kwabenya, Dome, and New Achimota were 0%, 1.5% (1/68) 
and 2.3% (1/44) . These are presented graphically in figure lb.
When the growth status was assessed by age (fig 2a) , it was 
seen that in children below 3 years (n=6) , none was wasted or 
wasted plus stunted; 83.3% had normal growth, and 16.7% were 
stunted. In the 36-47mo. old children,(n=40), 87.5% were of normal 
growth and 12.5% stunted. In the oldest children,ie those above 
60mo.(n=65), 66.2%, 3.1%, 26.6% and 4.1%, respectively were normal, 
wasted, stunted and wasted plus stunted. The corresponding figure 
was 83.1%, 5.6%, 11.2%, and 0% for the 48-60mo. old group.
Fig 2b depicted the trends in the skinfolds thicknesses 
(triceps and subscapular) and the mid- upper arm circumference. The 
mean triceps skinfolds, for the 200 cohorts, was 8 . 2 + 1 .8mm with 
values ranging from 7.2 + 1.0mm in New Achimota to 8 . 8 + 1.8mm in 
Ashalley Botwe. Subscapular skinfolds was 5.7 + 1.2mm, with values 
ranging from 5.2 + 0. 8mm in New Achimota to 6.0 + 1. 4mm in Ashalley 
Botwe. The mean MUAC was 16.1 + 1.0cm, ranging in value, from
16.1 + 1.0cm in Ashalley Botwe to 16.2 + 1.0cm in New Achimota.
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Pe
rc
en
ta
ge
 
(%
)
Fig 1a Age distribution of cohorts in the various communities
Ashalley Botwe 0  Kwabenya ED Dome ESI New Achimota
Fig 1b Leve| ° f  Malnutrition (wasting, stunting, wasting plus stunting) 
among preschoolers in communities studied
Wasted
70- —
50
Normal
~*~AII areas 
+  Ashalley Botwe 
96 Kwabenya 
■ Dome
^  New Achimota
Stunted Wasted & Stunted
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Fig 2a NUTRITIONAL STATUS IN THE VARIOUS AGE GROUPS
Fig 2b Skinfold and mid-upper arm circumference 
measurements of cohorts
TRICEPS(TSF,mm) SUBSCAP(SS,mm) MUAC(om)
r t * ! i  AREAS F I a s HALLEY BOTWE IIDkWABENYA CSdOME E In EW  ACHiMOTA
55
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The biochemical data of the cohorts were matched for the 
various types of growth/nutritional status to indicate whether 
there were any significant differences in the biochemical data that 
could explain differences in the growth/nutritional status. These 
values are shown in table 2 and figs. 3a-6b.
When the mean values of the indicators of zinc nutriture 
(hair zinc, rbc zinc, plasma zinc, and alkaline phosphatase 
activity) were compared to the reference values, (table 2 ), it was 
seen that they all fell within their reference range except for the 
rbc zinc for the stunted groups of Kwabenya and Dome and the wasted 
plus stunted group in New Achimota, where they fell slightly below 
their reference values. No significant differences (p>0.05) were 
however found between them and those of the normal groups. This 
indicated the absence of zinc deficiency in all cohorts studied.
The analyses of the data for the "All areas combined" group, 
using Duncan's and LSD multiple comparison tests indicated that 
there were significant differences (p< 0.05) in the weight,
height,triceps and mid upper arm circumference (MUAC) levels in the 
various types of growth; however, there was no significant 
difference (p>0.05) in the case of proteins, albumin, alkaline 
phosphatase activity (ALP), rbc zinc levels, plasma zinc, hair zinc 
or A/G ratios.
The trend was basically the same when the data were broken 
down into communities. Thus, in Ashalley Botwe, Dome and New
4.3. Biochemical data based on nutritional status:
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Achimota, there was a significant difference (p<0.05) in the 
weight, height, triceps, and MUAC levels, while no significant 
differences existed in the case of proteins, albumin, A/G ratios, 
ALP, rbc zinc, plasma zinc, and hair zinc levels.
In the determination of the various biochemical parameters, 
the accuracy of the analytical procedures were evaluated using 
basically two approaches. For hair, plasma and rbc zinc 
determinations, a standard bovine liver ( # 1577b; kindly donated 
by the National Institute of Standards and Technology, USA) was 
analyzed alongside these analyses anytime analyses were done. The 
average zinc level in this sample was 130+ 5ppm which compared 
favourably with certified value of 127+16 ppm.
For the protein and albumin, percentage recoveries were 
carried out. Recovery ranged from 95-101% for protein and 96-100% 
for albumin. For the precision, the coefficient of variation , (cv) , 
of 10 replicate determinations was carried out. The respective 
values for protein, albumin, hair zinc,plasma zinc, rbc zinc and 
alkaline phosphatase activity were 9.4%, 7.7%, 5.1%, 8.1%, 7.0%, 
and 1 0 .0%.
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Pa
rt
s/
m
il
li
on
 
(P
PM
)
Fig 3a Trends in the mean plasma protein, albumin, and A/G 
ratios in the various communities
13ALL AREAS S aSHALLEY BOTWE Q kWABENYA 0DOME EUnEW ACHIMOTA
F ig  3b  P la sm a  z in c  a n d  r b c  z in c  le v e ls  in  th e  v a r io u s  c o m m u n i t ie s
14
Plasma  ^inc(ppm) RBC z inc(ppm)
. r ,  b o t w e  D k WABENYA  Q d OME E lNEW ACHIMOTA
58
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rig 4a The mean Albumin/Globulin ratio in the different nutritional states
2
1.5
.2
03
CC !
o
0.5
0
M NORMAL □  WASTED □  STUNTED □  WASTED & STUNTED
Note the absence of wasting and wasting plus stunting in Kwabenya 
Fig 4b Albumin levels in the different nutritional states
ALL AREAS ASHALLEY BOTWE KWABENYA DOME NEW ACHIMOTA
5 3 NORMAL P ]WASTED □  STUNTED ^WASTED & STUNTED
59
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Pl
as
ma
 
zin
c 
lev
els
 
(P
PM
) 
Al
ka
lin
e 
ph
os
ph
at
as
e 
ac
tiv
ity
 
(U
/L
)
5a Trends in the mean alkaline phosphatase activity in the different nutritional states
E3NORMAL Q w ASTED Q STUNTED  H wASTED & STUNTED
Fig 5b Plasma zinc levels in the various nutritional states 
in the communities
S'WASTED □  STUNTED H  WASTED & STUNTED
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ig 6a TRENDS IN THE MEAN HAIR ZINC IN VARIOUS NUTRTIONAL STAl ES IN THE COMMUNITIES,
400 ---------------------------------- -----------------
„ s 300
2
CL
CL,
<D
>
0) 200
O
c
Kj
(0
T 100
ALL AREAS ASHALLEY BOTWE KWABENYA NEW ACHIMOTA
E3 NORMAL 0WASTED □STUNTED ISlWASTED & STUNTED 
Fig 6b MEAN HEIGHT AND ALKALINE PHOSPHATASE ACTIVITY IN THE COMMUNITIES
600 200
150
100
ALL AREAS ASHALLEY BOTWE KWABENYA DOME NEW ACHIMOTA
H mpight  B alkal ine  PHOS.
61
Me
an
 
he
igh
t 
(c
m
)
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4.4. Correlation among some indicators of zinc nutriture and 
anthropometric parameters.
1. The equation for regression line between hair zinc concentration 
and height of subjects for the various nutritional states was:
Y = 0.008x + 99.612, r=0.092
2. The equation for regression line between plasma zinc 
concentrations and height of subjects for the various nutritional 
states was: Y= 23.222x + 76.400, r=0.675
3 . The equation for regression line between rbc zinc concentrations 
and height of subjects for the various nutritional states was:
Y= 2.569X + 67.790, r=0.615
4. The relationship between hair zinc levels and
(i) triceps, Y =0.007x +6.264, r=0.297
(ii) subscapular, Y =0.017x +2.711, r=0.699
(iv) MUAC. Y =0.019x +11.393, r= 0.850
62
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CHAPTER FIVE
DISCUSSION OF RESULTS
The present study was undertaken essentially to ascertain 
whether or not zinc deficiency occurs within preschool children in 
the selected communities. If it does, whether the zinc nutritional 
status relates in any way to the various anthropometric states of 
the cohorts. The 4 communities studied were Ashalley Botwe, 
Kwabenya, Dome and New Achimota. They were chosen because of easy 
access by road from the research station and for the fact that they 
represent typical Ghanaian set-ups desired in the study. Thus 
Ashalley Botwe and Kwabenya villages represent typical southern 
rural villages, Dome represents a typical periurban cosmopolitan 
township while New Achimota is almost urban.
In Ashalley Botwe and Kwabenya, over 90% of the children 
within the desired age group (3-5 years) attend the nursery school 
in which the study was conducted. For Dome and New Achimota, other 
nurseries existed so the percentage of the cohorts assessed in 
these two settings was about 50% in each case.
The socioeconomic strata, based on the number of sandcrete 
houses, zinc/aluminium roofed houses, and general infrastructural 
developments such as availability of places of convenience, health 
posts, and potable water are as follows: New Achimota> Dome >
Ashalley Botwe > Kwabenya.
In the study, age, anthropometric measurements (weight, 
height, triceps, subscapular and MUAC) and biochemical indicators 
of protein and zinc nutritional status( total plasma protein,
63
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albumin, A/G ratio, plasma zinc, rbc zinc, alkaline phosphatase 
activity and hair zinc) were determined in the 200 cohorts. Their 
ages ranged from 35 to 62mo.
Based on a cut-off point of -2SD on the NCHS reference 
standard, 16.5% of the 200 cohorts screened were chronically 
malnourished,(stunted); 3.5% were acutely malnourished (wasted), 
1.5% were chronically and acutely malnourished ( wasted plus 
stunted), while 78.5% were of a normal nutritional status (fig. 
la). The wasted plus stunted group represents cohorts which have 
acute ( i.e. recent ) malnutrition Superimposed on a background of 
Chronic malnutrition.
The Ghana National Demographic Health Survey of 198825 on 
preschool children reported a chronic malnutrition rate of about 
28% and wasting rate of 7.4%. A 1987-1988 Agroecological nutrition 
survey117 based on z-score cut-off point of -2SD. indicated that 
malnutrition was particularly high in the savannah agroecological 
zone,ie northern sections of Ghana. The rates were 36% (chronic 
malnutrition) and 9.5% (acute malnutrition). The Greater Accra 
region was said to have the lowest levels of both chronic and acute 
malnutrition, being 22% and 6.5%, respectively.
When these figures are compared with the values obtained in 
the present study, (ie. 22% versus 16.5% for chronic malnutrition, 
and 6.5% versus 3.5%, for acute malnutrition), using tests of 
proportion, indicated that there is no significant difference 
(p>0.05) between the levels of chronic or acute malnutrition 
obtained in 1988 and the present study. It means that both forms of
64
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malnutrition are still with us and of the same magnitude as in 
1988. This is rather sad in view of numerous intervention 
programmes mounted by various agencies.
It is evident ( from fig.lb) that in Ashalley Botwe, Dome and 
New Achimota, the cohorts suffer from both acute and chronic 
malnutrition, while in Kwabenya, they suffer from chronic 
malnutrition only.
Based on the percentage of cohorts of normal nutritional 
status, the nutritional adequacy in the four communities can be 
ranked as follows. New Achimota> k!wabenya> Dome > Ashalley Botwe. 
The differences in the nutritional states are however not 
significant (p>0.05), except in the case of Ashalley Botwe. 
Analysis using test of proportions (table 1), indicated that the 
percentage of cohorts with normal nutritional status was 
significantly greater (p< 0.05) in New Achimota than in Ashalley 
Botwe. Also the percentage stunting in Ashalley Botwe was 
significantly higher (p< 0.05) than in either Dome or New Achimota.
In order to ascertain whether there were any significant 
differences in the zinc level in the various nutritional states, 
the biochemical data of the cohorts have been grouped based on 
anthropometric classification. (table 2). In this way, the 
biochemical indicators, such as protein, albumin, A/G ratio, 
plasma, rbc and hair zinc are compared in the various nutritional 
states. It must be noted that this type of comparison is valid 
because cohorts in the age-group of 35-60mo. are regarded as a 
homogenous group with virtually similar biochemical values.
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When the mean biochemical values were compared with standard 
values (table 2 ) , it was seen that only the plasma zinc in the 
wasted plus stunted group in Ashalley Botwe fell below the accepted 
level. The value was 0.63ppm as against a standard quoted value of
0.70 ppm67'76. Considering the fact that the normal plasma zinc range 
( determined by AAS ) is 0.50-1.50ppm118, the level of 0.63ppm is 
acceptable.
The mean plasma alkaline phosphatase activity obtained in this 
work is 447+131 U/L (with a range of 243-879 U/L) as compared to a 
normal range of 164-984 U/L119. The mean + SD plasma and hair zinc 
values are 1.07 + 0.33 ppm ( range of 0.50-2.25 ppm) and 206.4 + 
101.6 ppm ( range of 73.4-632.7 ppm), respectively as compared to 
normal levels of 0 .50-1. 50ppm118 and >70 ppm20'67, respectively. The 
mean + SD for rbc zinc is 13.0 + 2.6 ppm, with a range of 11.8-14.2 
ppm. as compared to a normal range of 12-14 ppm17.
It must be noted that even though the accepted lower limit for 
hair zinc is 70ppm17,20,76 , the upper limit has not been established. 
However, literature values indicate values up to 426ppm for elderly 
black Americans76. It might be expected that zinc levels in hair 
samples of normal African preschool children will be higher than 
the level in elderly people due to faster growth rate. Thus, using 
both plasma zinc (indicator of acute zinc status) as well as hair 
and rbc zinc (indicators of chronic zinc status), as indicators of 
zinc nutritional status, the mean values obtained in this study, 
are within the normal physiological range.
The analysis of the biochemical data in the normal, wasted,
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stunted and wasted plus stunted groups (table 2), using Duncan's 
and LSD multiple comparison tests indicate that there is no 
significant difference among any of the biochemical indicators in 
the various nutritional states. This also indicates that the 
protein nutritional status was adequate.
A visual inspection of the hair zinc values in the various 
groups (table 2 ) would suggest that values for the wasted plus 
stunted groups are significantly less than that for the other 
groups. The lack of any statistical significance is due to the 
rather large standard deviations, reflecting the differences in the 
respective hair zinc values in the cohorts.
It is therefore clear from the analyses that there is no 
significant difference (p>0.05) in the indicators of zinc nutriture 
(plasma zinc, hair zinc, rbc zinc and alkaline phosphatase 
activity) in the various anthropometric states. In other words, 
there is no zinc deficiency in the cohorts and also the zinc 
status does not relate in any way to the various anthropometric 
states, including stunting.
It must be noted that even though growth retardation is one of 
the earliest manifestations of zinc deficiency66, studies have shown 
that low zinc levels are not a characteristic feature of short 
stature per se. In a typical study30 , hair zinc concentrations were 
determined in 40 children, aged 4-16 years, referred to the 
pediatric endocrine clinic at Colorado General Hospital, USA, for 
investigation of severe growth retardation. Final diagnosis 
included growth hormone deficiency, familial short stature,
67
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primordial short stature, constitutional short stature, and 
hypothyroidism. The mean hair zinc levels did not differ from that 
of the control group and only one child, diagnosed as having 
constitutional short stature, had levels less than 70 ppm30. The 
results of the present study also show that hair zinc levels can be 
adequate even in the case of stunted growth, hence low zinc levels 
are not a characteristic feature of short stature per se.
The only work reported in the literature on the zinc status in 
preschool children in Ghana was by Ferguson et al.120- In their work, 
hair zinc levels in 76 preschool children, consuming cereals or 
starchy foods, were assessed in 2 rural southern villages. It was 
found that 83% of the children in one village (Sleper) and 39% in 
another village (Gidantuba), had 'low' hair zinc of <1.68^mol/g 
(109.8/xg/g) . This study clearly shows that there can be very wide 
variations in the percentage of subjects affected by 'low' levels 
of zinc within the various preschool populations in different 
communities in Ghana.
It is also noteworthy that the subjects used in the present 
study consumed mixed diet, as compared to the cereal/starchy diet 
in Ferguson's work, hence the absence of zinc deficiency in the 
present cohort, is not surprising. It is well known that cereals 
are a rich source of phytic acid, which is the most potent dietary 
inhibitor of zinc absorption in the small intestine120.
The starchy diets (as in Fergeson's work) will be a fairly 
rich source of fibre, which is also a potent inhibitor of zinc 
absorption. It is therefore apparent that the results of the two
68
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primordial short stature, constitutional short stature, and 
hypothyroidism. The mean hair zinc levels did not differ from that 
of the control group and only one child, diagnosed as having 
constitutional short stature, had levels less than 70 ppm30. The 
results of the present study also show that hair zinc levels can be 
adequate even in the case of stunted growth, hence low zinc levels 
are not a characteristic feature of short stature per se.
The only work reported in the literature on the zinc status in 
preschool children in Ghana was by Ferguson et al.120- In their work, 
hair zinc levels in 76 preschool children, consuming cereals or 
starchy foods, were assessed in 2 rural southern villages. It was 
found that 83% of the children in one village (Sleper) and 39% in 
another village (Gidantuba), had 'low' hair zinc of <1.68/imol/g 
(109.8^tg/g) . This study clearly shows that there can be very wide 
variations in the percentage of subjects affected by 'low' levels 
of zinc within the various preschool populations in different 
communities in Ghana.
It is also noteworthy that the subjects used in the present 
study consumed mixed diet, as compared to the cereal/starchy diet 
in Ferguson's work, hence the absence of zinc deficiency in the 
present cohort, is not surprising. It is well known that cereals 
are a rich source of phytic acid, which is the most potent dietary 
inhibitor of zinc absorption in the small intestine120.
The starchy diets (as in Fergeson's work) will be a fairly 
rich source of fibre, which is also a potent inhibitor of zinc 
absorption. It is therefore apparent that the results of the two
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studies can not be compared, due to some differences in the 
experimental design.
Equations for regression between hair, plasma and rbc zinc and 
the height for the various nutritional states, ie. normal, wasted, 
stunted and wasted plus stunted groups, indicate correlation 
coefficients, r , ranging from 0.092, for hair zinc and the 
corresponding heights to an r value of 0.675 for regression line 
between rbc zinc and the corresponding heights. This indicates a 
fairly strong positive correlation between height and either plasma 
zinc (r=0.675), or rbc zinc (r=0.615), but a low correlation 
between the height and hair zinc (0.092).
Even though there was no statistical difference between 
plasma, hair, and rbc zinc levels in the various nutritional states 
for the 200 cohorts ( due to the large standard deviations, 
resulting from differences in the individual values), the 
regression equations between height and these indicators ( which do 
not take account of standard deviations) show that there is a 
positive linear correlation between plasma, hair or rbc zinc levels 
and heights in the various nutritional states.
Although these results showed that there is no zinc 
deficiency in these cohorts, this does not necessarily mean that 
zinc metabolism is normal in the cohorts. Studies have shown 
beneficial effects of zinc supplementation even though plasma zinc 
levels are normal121,122. Beside, circulating and tissue levels of 
zinc do not necessarily reflect zinc status; for example, high hair 
zinc has been found in both replete and severe deficiency states3.
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Perhaps, this vindicates the fact that we are yet to find sensitive 
indicators for assessing zinc nutritional states.
70
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CHAPTER SIX 
CONCLUSIONS AND RECOMMENDATIONS
6.1. CONCLUSIONS.
The results of the present study indicated quite a high level 
of chronic malnutrition (stunting) 16.5% (33/200) and a lower
level of acute malnutrition (wasting)-3.5% (7/200) among the
preschoolers studied. As discussed earlier, these figures are 
comparable to those found in the 1987-1988 agroecological survey 
carried out in the Greater Accra region117.
The present study has also shown that there is no zinc 
deficiency in these cohorts, based on the parameters used, and that 
the zinc status did not differ significantly (p>0.05) in the 
various t^ le anthropometric states, even in the case of stunting.
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6.2. RECOMMENDATIONS FOR FUTURE WORK.
1. The study should be repeated in other areas, especially in the 
northern sector of the country, where malnutrition is generally 
higher than in the south.
2. Since zinc deficiency is also associated with excessive maternal 
bleeding at birth, and hypertension during pregnancy66, both of 
which are common in Ghana, a proper study should be designed to 
measure the zinc status of pregnant women at regular intervals, 
especially in areas or populations where these conditions are 
prevalent.
3. Zinc supplementation trials should be done, with appropriate 
controls, in normal and malnourished preschool children and other 
vulnerable groups,such as the elderly, to test its efficacy in our 
populations and to demonstrate any deficiencies in zinc nutrition 
in various populations.
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7. APPENDIX
(a) Determination of micro molar absorptivity ie. e
Mean absorbance reading = 0.731 ; e =A4Np((K04mmol/L)x25xlO'3 
= 0 .731x25xl0'3 
=18 .28xl0~3
Substituting this value into equation 1, we have 
Catalytic activity ( U L"1) = 3.027x 10~3
------------------------  x (
(18 . 28xl0'3) xlx 0 . 027xl0‘3
= 6133 x ( & A / ^ t )  2
Equation 2 was used to calculate the catalytic(enzyme) 
activity of all the plasma.
(b) Sample calculation:
From measurement of a plasma sample, the change in 
absorbance/ min. ie.(k.A/^t) = 0.052
Substituting this value into equation 2 above, the enzyme 
activity of this plasm sample = (0.055 x 6133) U L ' 1
= 337 U L ' 1
73
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TABLE 1. ANTHROPOMETRIC CLASSIFICATION OF THE COHORTS
GROWTH COMMUNITY MALE FEMALE TOTAL
All areas n % n % n %
combined
Normal 87a 43 . 5 70a 35.0 157 78.5
Wasted 5b 2.5 2b 1 . 0 7 3.5
Stunted 21c 10.5 12c 6 .0 33 16.5
Wasted
&stunted 2d 1 . 0 ld 0.5 3 1. 5
Total 115 57.5 85 42.5 200 100.0
Ashalley Botwe
Normal 23 41. 0 16 28.6 39e 69.6
Wasted 2 3.6 0 0 2f 3.6
Stunted 7 12.5 7 12.5 14s 25.0
Wasted &
stunted 0 0 1 1 . 8 lh 1 . 8
Total 32 57.1 24 42.9 56 100.0
Kwabenya
Normal 14 43.8 12 37.5 26e 81.2
wasted 0 0 0 0 0f 0
Stunted 6 18.8 0 0 69 18.8
Wasted &
stunted 0 0 0 0 0h 0
Total 20 62.5 12 37.5 32 100.0
74
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Table 1 continued
Wasted 
Stunted 
Wasted & 
stunted 
Total
Dome
30 44.1 
2 2.9 
4 5.9
1 1.5 
37 54.4
25 36.8 
1 1.5 
5 7.4
0 0 
31 45.6
55' 80.9 
3k 4.4 
9l 13.2
lm 1.5 
68 100.0
Normal
Wasted
Stunted
Wasted &
stunted
Total
New Achimota
20 45.5 
1 2.3 
4 9.1
1 2.3 
26 59.1
17 38.6 
1 2.3 
0 0
0 0
18 40.9
37' 84.1 
2k 4.5 
4L 9.1
lm 2.3 
44 100.0
Normal = weight-for-height and height-for-age are both >- 2SD of NCHS reference standards 
Uasted = weight-for-height is <-2 SD of NCHS reference standard.
Stunted = Height-for-age < -2SD of NCHS reference standard;
Uasted £ stunted = weight-for-height and height-for-age are both <-2SD of NCHS reference standard.
Analysis using test of proportion indicated that numbers marked with the same superscript are not significantly different (p>0.05).
75
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TABLE 2. MEAN VALUES OF THE ANTHROPOMETRIC AND BIOCHEMICAL DATA OF THE COHORTS
Growth/
Community
Weight
(kg)
Height
(cm)
Triceps
(mm)
Sub­
scapular
(nrn)
Mid 
upper 
arm circ 
(cm)
Protein 
( g/dL)
Albumin 
( g/ dL)
Albumin/
globulin
ratio
Alkaline
phosphat
ase
activity 
( U/L)
Plasma
zinc
(ppm)
Hair
zinc
(ppm)
RBC
zinc I 
(ppm)
Reference
ranges
5.8-8.3 3.5-5.5 1-2.5 164-984 0.50-
1.50
>70 12-14
All areas 
coobined
Normal
n=157 15.7
(1.7)
102.9
(5.5)
8.2
(1.7)
5.7
(1.1)
16.3
(0.9)
6.6a
(0.7)
3.9”
(0.3)
1.6°
(0.5)
473”
(129)
1.13'
(0.35)
247.7f 
(101.6)
13.1®
(2.6)
Wasted
n=7
13.4
(1.5)
104.6
(5.5)
6.4
(0.9)
4.4
(0.5)
14.6
(0.8)
6.9“
(1.1)
S.?
(0.5)
1,2*
(0.3)
386d
(82)
1.16*
(0.37)
200.9f 
(65.2)
14.0g 
(2.6)
Stunted
n=33
14.3
(1-7)
97.1
(6.1)
8.1
(2.1)
5.6
(1.4)
15.9
(1.1)
6.7”
(0.6)
3.9b
(0.3)
1.6*
(0.5)
485d
(152)
1.04*
(0.23)
220.0*
(83.8)
12.9s
(2.4)
Stunted &
Hasted
n=3
12.6
(0.6)
100.4
(2-1)
7.8
(0.3)
4.8
(0.4)
14.7
(0.5)
7.1a
(0.3)
3.8b
(0.5)
1.3°
(0.7)
444“
(114)
0.95*
(0.29)
157.6f 
(40)
12.1E
(1.0) *
Ashalley
Botwe
Normal
n=39
15.5
(1.4)
101.6
(5.8)
8.6
(1.6)
6.0
(1.3)
16.3
(0.7)
7.2
(0.9)
3.8
(0.4)
1.2
(0.4)
452
(126)
1.10
(0.28)
213.5
(56)
13.5
(3.1)
Wasted 
n=2 j
13.0
(0)
101.3
(1.5)
6.4
(1.4)
4.6
(1.1)
14.2
(1.1)
7.0
(0.9)
3.6
(0.3)
1.2
(0.2)
325
(50)
1.06
(0.20)
162.5
(10)
16.2
(3.1)
Stunted
n=14
13.6
(1.8)
93.1
(6.7)
9.7
(2-3)
6.5
(1.6)
15.9
(1.3)
6.9
(0.7)
3.9
(0.4)
1.4
(0.5)
418
(125)
1.03
(0.24)
250.6
(71.5)
13.0
(3.6)
Wasted &
stunted
n=1
12.0
(0)
101.3 8.0 4.6 15.0 7.4
(0.3)
3.6
(0.3)
1.0
(0.2)
370 0.63
(0.10)
13.4
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Table 2 continued
Kwabenya
Normal
n=26
15.2
(1.6)
101.1
(6.7)
8.6
(1.5)
5.8
(1.0)
16.3
(0.7)
6.5
(0.8)
3.9
(0.3)
1.7
(0.7)
530
(180)
1.15
(0.38)
353.7
(139.1)
12.5
(2.9)
Uasted
n=0
0 0 0 0 0 0 0 0 0 0 0 0
stunted
n=6
13.3
(1.5)
94.6
(5.6)
8.0
(2.0)
6.0
(0.8)
15.6
(1.1)
6.1
(0.5)
3.8
(0.3)
1.9
(0.5)
659
(199)
1.06
(0.08)
295.8
(121.3)
10.9
(1.1)
Masted &
stunted
n=0
0 0 0 0 0 0 0 0 0 0 0 0
Dome
Normal
n=55
15.8
(1.9)
104.0
(4.8)
8.2
(2.0)
5.7
(1.2)
16.3
(1.1)
6.3
(0.6)
3.8
(0.4)
1.7
(0.5)
466
(108)
1.14
(0.41)
200.7
(66.3)
13.0
(2.3)
Uasted
n=3
13.7
(2.1)
105.4
(6.1)
6.7
(0.8)
4.4
(0.3)
15.0
(0.5)
6.7
(0.6)
3.7
(0.5)
1.2
(0.2)
482
(33)
1.19
(0.51)
270.6
(43.5)
12.2
(2.4)
Stunted
n=9
14.2
(0.9)
98.4
(3.6)
8.0
(1.5)
5.5
(0.9)
16.0
(0.6)
6.6
(0.5)
3.9
(0.2)
1.6
(0.4)
428
(86)
0.87
(0.21)
202.6
(64.9)
11.9
(4.6)
Uasted
Sstunted
n=2
13.3
(0.4)
101.6
(0.8)
7.6
(0.3)
5.4
(0.8)
14.3
(0.4
7.2 4.0
(0.1)
1.4 575 1.04 185.9 12.1
(0.1)
Reference
ranges
5.8-8.3 3.5-5.5 1-2.5 168-984 0.50-1.5 >70 12-14
77
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Table 2 continued
Hew
Achimota
Normal
n=37
16.1
(1.8)
104.9
(4.4)
7.3
(1.0)
5.4
(0.8)
16.4
(0.9)
6.6
(0.5)
4.0
(0.3)
1.7
(0.4)
444
(115)
1.16
(0.32)
220.1
(109.5)
13.3
(2.0)
Uasted
n=2
13.5
(2.1)
106.7
(8.3)
5.9
(0.6)
4.2
(0.3)
14.4
(1.1)
7.1
(0.6)
3.9
(0.8)
1.3
(0.4)
352 1.19 167.9
(32.9)
13.6
(2.0)
Stunted
n=4
15.9
(2.4)
102.2
(1.6)
6.7
(1.4)
4.4
(0.5)
16.0
(1.2)
7.3
(0.3)
4.1
(0.2)
1.4
(0.2)
435
(76)
1.10
(0.32)
171.7
(49.6)
15.9 j 
(4.9)
Uasted
&stunted
n=1
12.5 98.3 7.8 4.4 15.1 6.7 3.8 1.5 387 1.19 129.3 10.9
Reference
ranges
5.8-8.3 3.5-5.5 1-2.5 164-984 0.50-1.5 >70 12-14
Values are means for both sexes. SD values are shown in parenthesis. Values with the same superscript are not significantly different (P>0.05). Test for 
significance was done using LSD and Duncan's multiple range test. Sources of reference ranges: Reference # 36,111 & 118
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8. REFERENCES
1. Atinmo T. Food, Nutrition and health issues in Africa. In: 
Abstracts of the 14th international congress of nutrition. 
August 1989 (s47-4) p. 303.
2. FAO monogram. Food and nutrition at the turn of millennium,
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