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oral contraceptive& 2
Women using estrogen-containing oral con- indices of vitamin B6 nutrition measured were
traceptives excrete elevated amounts of trypto- reported previously (5). In brief, 9 healthy control
women (average age 22.3 1.9 years), and 15 women
phan metabolites after a tryptophan load test (23.2 3.1 years) who had used oral contraceptive
compared with women not taking oral contra- agents for at least 6 months were given a diet that
ceptives (1-3). Elevated levels of 3-hydroxy- contained only 0.19 mg of pyridoxine equivalents per
anthranilic acid were also observed in basal day (6). Oral contraceptive users started the diet on
day 1 1 of a 21-day pill sequence. Control subjects
urines from such subjects (4). When pyridoxine
started 14 days after onset of the previous menses. For
was administered to such subjects the excretion the first 4 days, while baseline studies were made, the
of tryptophan metabolites was decreased diet was supplemented daily with 0.8 mg of pyri-
toward normal levels, but in some subjects large doxine hydrochloride (PN-HC1). This supplement was
amounts of the vitamin may be required (3). then withdrawn and both groups of subjects consumed
only the deficient diet for 28 days. After this
These data have been widely interpreted as
depletion period, subjects were supplemented with
indicating that the use of oral contraceptive either 0.8, 2.0, or 20 mg/day of PN-HCI for another
drugs causes an increased requirement for 28 days. Before release from the study, subjects were
vitamin B6, although other explanations for the supplemented for a final 4 days with 100 mg
PN-HC1/day. During the baseline period, and at
altered tryptophan metabolism may be pos-
weekly intervals throughout the study, several indices
sible. To evaluate the effect of oral contracep- of vitamin B6 nutrition were measured in each subject.
tive usage on the requirement for vitamin B6, a The indices measured included urinary tryptophan
variety of indices of vitamin B6 nutrition were metabolites before and after a 2.0 g oral load of
measured to determine the rate at which L-tryptophan (7, 8), urinary cystathionine after a load
control and oral contraceptive using women
became depleted of this vitamin while ingesting From the Division of Clinical Oncology, Uni-
versity of Wisconsin Medical School and Department
a diet low in vitamin B6. The same indices were
of Nutritional Sciences, University of Wisconsin
used to measure the rate at which these subjects College of Agricultural and Life Sciences, Madison,
became repleted when supplemented with Wisconsin 53706.
physiological levels of pyridoxine. Supported in part by Wisconsin College of
Agricultural and Life Sciences, Contract NIH, HICHD
72-2782 with the National Institute of Child Health
Methods and materials
and Human Development, and Grant No. CA-13302
The experimental details of these studies con- from the National Cancer Institute, Public Health
cerning the selection of subjects, diets used, and Service, Bethesda, Maryland 20014.
The American Journal of Clinical Nutrition 28: MAY 1975, pp. 535-541. Printed in U.S.A. 535
536 LEKLEM El AL.
of 3.0 g of L-methionine (9, 10), urinary 4-pyridoxic At comparable times of depletion and at
acid (5), plasma pyridoxal phosphate (5), and erythro- equal levels of PN -HC1 supplementation, the
cyte activities of alanine arninotransferase and aspar-
mean excretion of tryptophan metabolites was
tate aminotransferase (5). In order to study metabo-
lism along the kynurenine pathway and to circumvent consistently greater than that of controls, and
any variations in activity of tryptophan oxygenase, suggests that the requirement for vitamin B6
loading doses of L-kynurenine sulfate (200 mg/dose) may be slightly greater in oral contraceptive
were given initially, at the time of maximum
users than in controls. However, it should be
deficiency, and after repletion with pyridoxine for 4
weeks. pointed out that because of large individual
variations and limited numbers of subjects,
Results many of these differences do not achieve
Measurement of urinary tryptophan metabo- statistical significance. Details of the excretion
lites after tryptophan loading prior to starting of individual tryptophan metabolites are pub-
the deficient diet showed that the oral contra- lished elsewhere (8).
ceptive users excreted elevated levels of kynure- The urinary excretion of cystathionine after
nine , acetylkynurenine , 3-hydroxykynurenine a 3.0 g oral load of L-methionine is shown in
and xanthurenic acid compared with the con- Fig. 2. The pattern is different from that of the
C.,J
both groups (5).
To evaluate metabolic effects of oral contra-
Ui
-J
0 ceptive use on the tryptophan metabolic path-
way independent of any effects on the activity
Ui
z of tryptophan oxygenase, 200 mg loads of
z L-kynurenine sulfate monohydrate were given
0
I before deficiency, at peak deficiency, and after
4I-
pyridoxine supplementation. The excretions of
U)
CONTROLS O.C.
the first 2 weeks of repletion with PN-HC1 the 0.8 #{149}- 0.8 0--- .7
#{149} 2.0 #{163}- 2.0 0---
excretion by oral contraceptive users remained
above that of controls at comparable times.
.5
-----
Excretion of urinary 4-pyridoxic acid de-
creased at similar rates in both groups of - ------- - i
-
6.- PLP 1 and controls were found initially and activity of
CONTROLS OC. both groups decreased similarly during the
0.8 0---
0.8.-
2.0- 20 0---
depletion period. Daily supplements of 0.8 mg
I 2 , of PN-HC1 slowly increased the activity but not
a.
U)
0 to predepletion levels. With 2.0-mg supple-
i0-
a. ments, the activity of both groups had risen to
8
approximately predepletion levels. The final
x0
0 6-
points shown were after 3 or 4 days of
supplementation with 100 mg PNHC1/day.
0. 4-
4
This level of supplement resulted in normal or
In2 supernormal levels.
4,
-J
a. _L.- L i The above erythrocyte preparations were
0 2 3 4 5 6
7 8 also assayed after fortification in vitro with
WEEK OF STUDY
-
4---
FIG. 4. Concentration
- ---Be
of plasma
-
pyridoxal
-64
phos-
- -, saturating
stimulation
progressed,
levels
and
of PLP (Fig.
by PLP increased
did so to
6). The percent
extent the in
as the
same
deficiency
q 23
23 23 23 23
Period of Study
aminotransferase , and the findings paralleled which time all other indices were within normal
those of the alanine enzyme although the control ranges. This suggests that the oral
relative decreases induced by vitamin B6 de- contraceptives may have some relatively spe-
ficiency were not as great. Details of these cific effect on the metabolism of tryptophan
studies have been presented elsewhere (5). which is independent of vitamin B6 levels. This
effect may be primarily on the activity of
Discussion tryptophan oxygenase, since studies in rats have
The excretion of tryptophan metabolites by shown direct as well as adrenal-mediated effects
oral contraceptive users, after the tryptophan of estrogens on the activity of this enzyme
load test, was significantly different from ( 11). However, the present observations in
similarly loaded controls prior to induction of subjects given small kynurenine loads, which
vitamin B6 deficiency.This confirmed previous bypass any tryptophan oxygenase effects, sug-
observations of altered tryptophan metabolism gest that the usage of contraceptive hormones
in oral contraceptive users (1-3). Other indices also may have an effect elsewhere in the
of vitamin B6 nutrition measured before vita- pathway of kynurenine metabolism. Previous
mm B6 depletion were not clearly different in studies (12, 13) indicate that steroids or steroid