9

1990 by the Humana Press, Inc.

All rights of any nature, whatsoever, reserved.
0163-4984/90/2401-0083 $02.00

Zinc Acutely and Temporarily Inhibits

Adrenal Cortisol Secretion in Humans
A Preliminary Report
J. B R A N D / ~ O - N E T O , * " B. B. DE M E N D O N ~ A , 2 T. SHUHAMA,3
J. S. MARCHINI,' W. P. PIMENTA,' AND M. T. T. TORNERO1
1Department of Internal ,a'ledicine, Faculdade de ,a,ledicina de
Botucatu, Universidade Estadual Paulista, 18610, Botucatu, SP;
2Department o f Internal ,a'ledicine, Faculdade de ,a'ledicina,
Universidade de S~o Paulo, 05403, S~o Paulo, SP; and 3Department
o f Physical Chemistry, Faculdade de Ci6ncias Farrnac6uticas de
Ribeir~o Preto, Universidade de S~o Paulo, I4.049,
Ribeir~o Preto, SP, Brazil

ABSTRACT
Hypo- and hyperzincemia has been reported to cause alterations
in the adrenal secretion. To determine the acute effect of zinc on
cortisol levels, we studied 27 normal individuals of both sexes aged
20-27 y after a 12-h fast. The tests were initiated at 7:00 AM when an
antecubital vein was punctured and a device for infusion was installed and maintained with physiological saline. Zinc was administered orally at 8:00 AM. Subjects were divided into an experimental
group of 13 individuals who received doses of 25, 37.5, and 50 mg of
zinc and a control group of 14 individual who received 20 mL of
physiological saline. Serial blood samples were collected over a period of 240 rain after basal samples ( - 3 0 and 0 min). We detected an
acute inhibitory effect of zinc on cortisol secretion during 240 min of
the study period in the experimental group.
Index Entries: Acute hyperzincemia; zinc supplement, effects
of on cortisol secretion; plasma zinc and cortisol levels; normal
individuals.
Author to whom all correspondence and reprint requests should be addressed.
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B r a n d , o-Nero et al.

INTRODUCTION
The adrenals of vertebrates are known to be rich in zinc (1), but few
data are available in the literature about the effects of zinc on cortisol
synthesis and secretion. The adrenal cortex may be affected by zinc in
some aspects. For example, rats with zinc deficiency have high cholesterol and ascorbic acid levels in their adrenals in relation to control
animals and increased plasma 11-hydroxysteroid levels under basal conditions or when stimulated with ACTH (2). However, excess zinc has an
opposite effect on plasma steroid levels, since it significantly inhibits
plasma 11-hydroxysteroids in rats (3).
Conversely, the effect of glucocorticoids on zinc has been detected
both in humans and in experimental animals. Patients with adrenal
insufficiency and Cushing syndrome, respectively, show increased and
decreased serum zinc levels (4). These results have also been obtained for
adrenalectomized cats (4) and rats supplemented with dexamethasone
(5), thus corroborating the existence of an inverse correlation between
zinc and glucocorticoids.
On the basis of these considerations, the objective of the present
study was to determine the possible existence of an acute effect of zinc on
basal cortisol secretion in humans, a phenomenon that has not yet been
reported in the literature.

MATERIAL AND METHODS

We studied 27 normal individuals (medical students) aged 20-27 y
after obtaining informed consent in writing, according to the recommendations of the Declaration of Helsinki. The individuals had no history of
endocrinopathy, used no type of drugs, and had ideal weights. The test
.was started at 7:00 AM after a 12-h fast. A no. 19 infusion device was
initially inserted into an antecubital vein. Basal blood samples were
collected at 7:30 and 8:00 AM ( - 30 and 0 min) and all individuals maintained resting decubitus from the beginning to the end of the test. The
control group consisted of 14 subjects of both sexes who received 20 mL
physiological saline, p.o. Serial blood samples were collected from 8 of
these individuals at the following time points: 30, 60, 90, 120, 150, 180,
210, and 240 min, and at 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110 and 120
rain from the remaining 6. Heptahydrate zinc sulfate diluted in 20 mL
deionized water was administered to the experimental group after the
basal collection. Three men and 4 women ingested 25 mg zinc and 3
women ingested 37.5 mg zinc. Blood samples were collected from each
subject at 30-min intervals for 240 rain. Three other women received 50
mg zinc and blood samples were collected from them at 10-min intervals

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Zinc and Cott/sol Secretion

85

for 120 min. The objective of this reduction in time scale was to guard
against the possibility that some hormonal alteration may go undetected.
Plastic syringes and tubes and zinc-free EDTA were used to obtain
plasma. Plasma samples were stored at -20~ for zinc and cortisol
determination. All tubes and pipets used were previously washed with
acetone, 3 N hydrochloric acid, 0.1% EDTA, and deionized water.
Zinc was measured by atomic absorption spectrophotometry (6).
Intrassay variation was 2.5% and sensitivity 0.02 ~g/mL. Samples were
measured in triplicate and those showing hemolysis were excluded.
Cortisol was measured by radioimmunoassay by the double-antibody
method using kits of Diagnostic Products Corporation, US. Intra- and
interassay variation was 6.4 and 7.4%, respectively, and sensitivity was
1.0 ~,g/dL. Samples were measured in duplicate.
The basal levels of zinc and cortisol were defined as the mean
concentration of the - 3 0 and 0 values. Data were analyzed statistically
by the Sign test, by the t-test for independent samples, and by the MannWhitney test (7).

RESULTS
Since the individuals in the control group showed plasma cortisol
levels of 3.4-18.5 p,g/dL during the basal period, we defined individuals
within this range as the sample for the experimental group. Thus, we
had a total of 14 individuals in the control group and 12 in experimental
group (1 individual showing a basal level of 24.1 p,g/dL was excluded
from the study). Plasma zinc and cortisol are presented in Figs. 1, 2,
and 3.
For data analysis, the direction and magnitude of the basal differences (basal cortisol minus cortisol level at time t) were observed, each
individual serving as his/her own control. Direction was analyzed by the
Sign test. The number of individuals with a fall in cortisol levels was
statistically significant at all times, in the control group, except for 210
min, and in the experimental group (Table 1). Comparison of the magnitudes of the differences in cortisol levels between the control and the
experimental group showed that the mean difference was statistically
higher (t-test) in the the experimental group at all times analyzed (Table
2). The maximum difference in cortisol level within each individual was
also measured, regardless of the occasion when the maximum and minimum peaks had occurred. The comparison of the magnitude of the
maximum difference in cortisol between the two study groups revealed
that the maximum difference was statistically higher (Mann-Whitney
test) in the experimental group up to 120 rain, thus corroborating the
data in Table 1.

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Brand&o-Neto et aL

86

16

4
N

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r

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-

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N

BASAL :30

60

90

120 150 180 210 240

TIME ~ rain

Fig. 1. Plasma cortisol in the control group (o-o) and experimental group
(o--o), and plasma zinc in the experimental group (nt), according to oral zinc
dose (25 mg) and collection time. The results are reported as means (~) ___
standard deviation (SD).

DISCUSSION
The acute inhibitory effect of zinc on cortisol secretion in vivo was
demonstrated in the present experiment. The fall in cortisol in the experimental group in relation to the control group occurred during the 240
min of the study period. The intrinsic mechanism of this phenomenon is
still to be elucidated. At the moment, evaluation of the interrelationship
between this loss of cortisol and the different zinc doses was made
difficult by the small sampling in each subgroup.
Very little is known about the effects of zinc on the physiological
processes of cortisol secretion. The fact that zinc inhibits basal cortisol
secretion in humans may be related to a direct blockade of cortisol
synthesis and secretion in the adrenal cortex. This cause-effect relationship has been supported by reports of other authors who observed
adrenal hyperplasia and a decrease in plasma 11-hydroxysteroids in rats
on zinc-rich diets. (3). This trace element is known to be important for the
structural and functional integrity of the cell membrane, since, by acting
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1/ol. 24, 1990

Zinc and Cortisol Secretion

87

16

A
J

12
a

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BASAL 3 0

60

90

120 150 180

T I M E ~ rain

210

24.0

Fig. 2. Plasma cortisol in the control group (o--o) and experimental group
(o--o), and plasma zinc in the experimental group (B-N), according to oral zinc
dose (37.5 mg) and collection time. The results are reported as means (2) +
standard deviation (SD).
on the sulfhydryl group, it can inhibit several membrane enzymes,
among them ATPase and adenyl cyclase (8), and, through this effect, it
may block cortisol secretion by the fasciculated and reticular cells of the
adrenal cortex.
Moreover, many plasma membrane receptors have been reported to
have the function of transmitting hormonal information to the interior of
the cell (9). We know that zinc, by acting on membrane receptors for
mast cells and platelets, can inhibit histamine and serotonin release,
respectively (10,11). In this respect, it is plausible to postulate an inhibitory effect of zinc also on ACTH-receptor sites.
A study is currently underway in our laboratory to determine the
site(s) of action and the intrinsic mechanisms of these processes.

SUMMARY
The ability of the adrenal cortex to decrease cortisol secretion in
response to ingestion of zinc was studied in 13 normal individuals of
both sexes aged 20 to 27 y. Plasma cortisol levels fell significant in all
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Vol. 24, 1990

Brand&o-Neto et aL

88
16

k\

,.-.-12

~-

""

=k

~8
I.-0

(3

BASAL I0

20

30

40

50
60 70
TIME ~ rain

80

90

I00

II0

120

Fig. 3. Plasma cortisol in the control group (e--i) and experimental

group (o--o), and plasma zinc in the experimental group ( ! t ) , according to
oral zinc dose (50 mg) and collection time. The results are reported as
means (Y) + standard deviation (SD).
Table 1
Number of Individuals Showing Decreased Cortisol Levels
(X) During Basal Time, Total Number of Individuals (N),
and Test of Significance~ for the Different Collection Times
for the Control and Experimental Groups
Control
Time, min
120
150
180
210
240

X
13"*
7*
7*
5 NS
7*

(N)
(14)
(8)
(8)
(8)
(8)

Experimetal
X
(N)
12"*
(12)
9**
(9)
9**
(9)
9**
(9)
9**
(9)

'Sign test for Ho: P [Yo > Yt] = 89vs H,: P [Yo > Yt] > 89 and NS P
[X < X] > 5%.
*Indicates 1% P [X < X] < 5% and ** indicates P [X < X] < 1%.
individuals in relation to the control g r o u p t h r o u g h o u t the study. These
results d e m o n s t r a t e that acute h i g h blood zinc levels w e r e able to inhibit
basal cortisol levels in n o r m a l individuals.

ACKNOWLED~NTS
This w o r k was s u p p o r t e d b y FAPESP (Proc. no. 86/2511-3; 88/0950-5)
a n d F U N D U N E S P (Proc. no. 289/88, DFP). We a c k n o w l e d g e the technical assistance of Maria Nice da S. Tavares.
Biological Trace Element Research

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89

Zinc and Cortisol Secretion

Table 2
Mean Difference (d) in Cortisol Level During Basal Time,
Number of Individuals (N), and t-Test" Comparing the Control
and Experimental Groups for the Various Collection Times
Control
Time, rain
120
150
180
210
240

d
2.76
3.00
2.60
3.15
2.05

(N)
(14)
(8)
(8)
(8)
(8)

Experimental
d
(N)
8.09
(12)
7.12
(9)
8.04
(9)
8.96
(9)
7.41
(9)

t-Test
**
*
**
**
**

9t-Test for H0: u~ = uE vs Hi: Uc< uE. **Indicates P [T < tl < 1%, and * indicates 1% <
P IT < t] < 5%.

REFERENCES
1. E.J. Underwood, Trace Elements in Human and Animal Nutrition, 4th ed., E. J.
Underwood, ed., Academic, NY, 1977, pp. 196-242.
2. J. Quarterman, Proc. Nutr. Soc. 31, 74-A (1972).
3. J. Quarterman, Trace Element Metabolism in Animals-2, W. G. Hoekstra, J. W.
Suttie, H. E. Ganther, and W. Mertz, eds., University Park Press, Baltimore,
1974, pp. 742-744.
4. R. I. Henkin, Trace Element Metabolism in Animals-2, W. G. Hoekstra, ]. W.
Suttie, H. E. Ganther, and W. Mertz, eds., University Park Press, Baltimore,
1974, pp. 647-651.
5. K. R. Etzel, S. G. Shapiro, and R. J. Cousins, Biochem. Biophys. Res. Commun.
89, 1120 (1979).
6. M. M. Parker, F. L. Humoller, and D. J. Mahler, Clin. Chem. 13, 40 (1967).
7. S. Siegel, Nonparametric Statistics for the Behavioral Sciences, 1st ed., S. Siegel,
ed., McGraw-Hill, NY, 1956, pp. 61-158.
8. W. J. Bettger and B. L. O'dell, Life Sci. 28, 1425 (1981).
9. K. J. Catt and M. L. Dufau, Ann. Rev. Physiol. 39, 529 (1977).
10. W. Kazimierczak and C. Maslinski, Agentes and Actions 4, 203 (1974).
11. M. Chvapil, Med. Clin. North America 60, 799 (1976).

Biological TraceElementResearch

Vol.24, 1990