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Effects of tryptophan-rich breakfast and light exposure during the daytime on melatonin secretion at night

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The purpose of the present study is to investigate effects of tryptophan intake and light exposure on melatonin secretion and sleep by modifying tryptophan ingestion at breakfast and light exposure during the daytime, and measuring sleep quality (by using actigraphy and the OSA sleep inventory) and melatonin secretion at night. Thirty three male University students (mean +/- SD age: 22 +/- 3.1 years) completed the experiments lasting 5 days and 4 nights. The subjects were randomly divided into four groups: Poor*Dim (n = 10), meaning a tryptophan-poor breakfast (55 mg/meal) in the morning and dim light environment (<50 lx) during the daytime; Rich*Dim (n = 7), tryptophan-rich breakfast (476 mg/meal) and dim light environment; Poor*Bright (n = 9), tryptophan-poor breakfast and bright light environment (>5,000 lx); and Rich*Bright (n = 7), tryptophan-rich breakfast and bright light. Saliva melatonin concentrations on the fourth day were significantly lower than on the first day in the Poor*Dim group, whereas they were higher on the fourth day in the Rich*Bright group. Creatinine-adjusted melatonin in urine showed the same direction as saliva melatonin concentrations. These results indicate that the combination of a tryptophan-rich breakfast and bright light exposure during the daytime could promote melatonin secretion at night; further, the observations that the Rich*Bright group had higher melatonin concentrations than the Rich*Dim group, despite no significant differences being observed between the Poor*Dim and Rich*Dim groups nor the Poor*Bright and Rich*Bright groups, suggest that bright light exposure in the daytime is an important contributor to raised melatonin levels in the evening. This study is the first to report the quantitative effects of changed tryptophan intake at breakfast combined with daytime light exposure on melatonin secretion and sleep quality. Evening saliva melatonin secretion changed significantly and indicated that a tryptophan-rich breakfast and bright light exposure during the daytime promoted melatonin secretion at this time.
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O R I G I N A L A R T I C L E Open Access
Effects of tryptophan-rich breakfast and light
exposure during the daytime on melatonin
secretion at night
Haruna Fukushige
1
, Yumi Fukuda
2
, Mizuho Tanaka
1
, Kaoru Inami
3
, Kai Wada
4
, Yuki Tsumura
5
, Masayuki Kondo
6
,
Tetsuo Harada
4
, Tomoko Wakamura
1
and Takeshi Morita
2*
Abstract
Background: The purpose of the present study is to investigate effects of tryptophan intake and light exposure
on melatonin secretion and sleep by modifying tryptophan ingestion at breakfast and light exposure during the
daytime, and measuring sleep quality (by using actigraphy and the OSA sleep inventory) and melatonin secretion
at night.
Methods: Thirty three male University students (mean ± SD age: 22 ± 3.1 years) completed the experiments
lasting 5 days and 4 nights. The subjects were randomly divided into four groups: Poor*Dim (n = 10),
meaning a tryptophan-poor breakfast (55 mg/meal) in the morning and dim light environment (<50 lx)
during the daytime; Rich*Dim (n = 7), tryptophan-rich breakfast (476 mg/meal) and dim light environment;
Poor*Bright (n = 9), tryptophan-poor breakfast and bright light environment (>5,000 lx); and Rich*Bright
(n = 7), tryptophan-rich breakfast and bright light.
Results: Saliva melatonin concentrations on the fourth day were significantly lower than on the first day in the
Poor*Dim group, whereas they were higher on the fourth day in the Rich*Bright group. Creatinine-adjusted melatonin
in urine showed the same direction as saliva melatonin concentrations. These results indicate that the combination of
a tryptophan-rich breakfast and bright light exposure during the daytime could promote melatonin secretion at night;
further, the observations that the Rich*Bright group had higher melatonin concentrations than the Rich*Dim group,
despite no significant differences being observed between the Poor*Dim and Rich*Dim groups nor the Poor*Bright
and Rich*Bright groups, suggest that bright light exposure in the daytime is an important contributor to raised
melatonin levels in the evening.
Conclusions: This study is the first to report the quantitative effects of changed tryptophan intake at
breakfast combined with daytime light exposure on melatonin secretion and sleep quality. Evening saliva
melatonin secretion changed significantly and indicated that a tryptophan-rich breakfast and bright light
exposure during the daytime promoted melatonin secretion at this time.
Keywords: Bright light exposure, Circadian rhythm, Melatonin, Sleep, Tryptophan
* Correspondence: morita@fwu.ac.jp
2
Department of Environmental Science, Fukuoka Womens University, 1-1-1,
Kasumigaoka, Higashi-ku, Fukuoka 813-8529, Japan
Full list of author information is available at the end of the article
© 2014 Fukushige et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the
Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public
Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this
article, unless otherwise stated.
Fukushige et al. Journal of Physiological Anthropology 2014, 33:33
http://www.jphysiolanthropol.com/content/33/1/33
Background
Melatonin is a hormone secreted by the pineal gland in
the brain. Its secretion is controlled by the suprachias-
matic nucleus in the hypothalamus, the central biological
clock of circadian rhythms in humans. Daily exposure to
light resets the phase of the clock in the suprachiasmatic
nucleus [1]. Melatonin secretion usually shows a peak at
midnight, and it is often used as an indicator of sleep qual-
ity and phase advances or delays of the sleep-wake cycle
[2,3]. In the clinical field, clinical doctors and researchers
use melatonin to improve sleep quality: to treat in-
somnia and depression and to reduce jet lag, and it is
recommended by some clinicians as a preventive agent for
breast cancer [4-9].
Tryptophan is one of the essential amino acids and is
contained in proteins sourced from milk, eggs, meat,
grains, and beans. It can cross the bloodbrain barrier
and is transformed into serotonin in the brain, subse-
quently converted to melatonin. The biosynthesis of
melatonin is rate-limited by the activity of arylalkylamine
N-acetyltransferase [10]. Zawilska et al. reported that
arylalkylamine N-acetyltransferase activity is affected by
light; its activity declines in the photoperiod (while mela-
tonin secretion also decreases) and increases in the sco-
toperiod (when melatonin secretion also increases) [11].
Therefore, tryptophan intake and the timing of light
exposure must be considered together if the effect of
tryptophan upon melatonin secretion and sleep quality
is to be maximized.
Hudson et al. [12] reported that using protein as a
source of tryptophan before sleep improved its quality at
night by amounts that were comparable with the intake
of pharmaceutical-grade tryptophan. Markus et al. [13]
examined the effects of tryptophan intake (from different
sources) at breakfast on the plasma tryptophan/large
neutral amino acids (TRP/LNAA) ratio and mood. The
results showed that intake of a tryptophan-rich source from
hydrolyzed protein had significantly greater effects on the
plasma TRP/LNAA ratio and improvement of mood than
did pure tryptophan and alpha-lactalbumin whey protein.
In addition, Markus et al. [14] measured depressive mood
and perceptual-motor and vigilance performance in
subjects under stress (who had high or low chronic stress
vulnerabilities) after they had ingested tryptophan-rich
egg protein hydrolysate in the morning. The results
revealed that egg protein hydrolysate improved the
depressive mood in all subjects and perceptual-motor
and vigilance performance in those subjects who had
low chronic stress vulnerability. These experiments were
performed with the aim of examining the effects of trypto-
phan intake in the morning on serotonin synthesis, which
improves mood and mental activity. However, an import-
ant factor for this aspect of tryptophan function light
intensity during the daytime was not considered,
and effects on sleep and melatonin secretion were not
investigated.
Nakade et al. [15] reported the relationship between
tryptophan intake and sleep quality (in children aged 2
to 6) in a survey of daily breakfast composition, morning
light exposure, and sleep. They found that tryptophan
intake at breakfast, coupled with morning light exposure,
was associated with higher melatonin secretion and easier
onset of sleep the following night. There is also evidence
that tryptophan intake in the morning and light exposure
at night affect the following sleep; Wada et al. [16] reported
that tryptophan-rich breakfasts and low-color temperature
light sources at night increased saliva melatonin concen-
tration in University soccer club members. Nevertheless,
these investigations were carried out without regulation of
several aspects of the individualslifestyle, including over-
all diet and daytime light exposure.
The purpose of the present study is to investigate ef-
fects of tryptophan intake and light exposure on sleep
and melatonin secretion, by modifying tryptophan inges-
tion at breakfast and light exposure during the daytime,
and measuring sleep quality and melatonin secretion at
night.
Methods
Subjects and group setting
The experiment was performed during September 17th
to 21st, 2012 (lasting 5 days and 4 nights) on 40 male
University students, none of whom had any mental dis-
orders (Cornell Medical Index), food allergies, or were
taking drugs. The data from 33 of them (mean ± SD age:
22 ± 3.1 years, height: 173 ± 5.5 cm, weight: 61.3 ± 6.7 kg,
BMI: 20.5 ± 2.3) were analyzed as three subjects with-
drew and four subjectssaliva samples were too small to
be analyzed. The subjects were randomly divided into
four groups: Poor*Dim (n = 10), tryptophan-poor break-
fast (55 mg/meal) in the morning (between 07:30 h and
08:00 h) and dim light environment (<50 lx) during the
daytime (between 07:30 h and 18:00 h); Rich*Dim (n = 7),
tryptophan-rich breakfast (476 mg/meal) and dim light
environment; Poor*Bright (n = 9), tryptophan-poor
breakfast and bright light environment (>5,000 lx); and
Rich*Bright (n = 7), tryptophan-rich breakfast and bright
light environment. It is more convenient and practical for
healthy young people living a normal, busy lifestyle if the
tryptophan in their diet is derived from common food-
stuffs; therefore, the tryptophan-poor breakfast consisted
mainly of vegetables, such as salad and orange juice,
whereas the tryptophan-rich breakfast was loaded with
proteins from food sources such as salmon and natto (fer-
mented soybeans; a Japanese traditional food) rather than
from nutritional supplements. The experimental condi-
tions and subjectsphysical characteristics for the four
groups are shown in Table 1. Subjectsage, height, weight,
Fukushige et al. Journal of Physiological Anthropology 2014, 33:33 Page 2 of 9
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and BMI were not significantly different between the four
groups (one-way ANOVA). Table 2 shows the estimated
intake of energy and basic nutrients, including tryptophan,
at breakfast, lunch, and supper. The breakfast menus were
designed to differ in tryptophan content (TRP-Poor and
TRP-Rich breakfasts) but have minimal changes in other
nutrients and energy. For each group, breakfast, lunch, and
supper were the same on all experiment days. The energy
and nutrient intakes of the controlled food were calculated
according to the Standard Tables of Food Composition in
Japan (5th Revised and Enlarged Edition). Lunch and sup-
per were controlled with contents that were recommended
as appropriate for healthy young males by the Ministry of
Health, Labour and Welfare of Japan.
The experimental protocol was conducted in accord-
ance with the Declaration of Helsinki and conformed to
international ethical standards, and was approved by the
Ethics Committee at Fukuoka Womens University. Sub-
jects gave prior written, informed consent and appropriate
compensation was given to them after they had completed
the experiment.
Experimental procedure
Figure 1 shows the experimental protocol. For one week
prior to the experiment, subjects were asked to eat a
vegetable-based breakfast (for tryptophan depletion) and
to keep to a regular sleep-wake cycle (retiring at 00:00 h
and rising at 07:00 h). Throughout this week, subjectsad-
herence to these requirements was checked by objective
measurement of activity with an Actiwatch (Mini-Mitter
Co. Inc., Oregon, USA) and requiring the subjects to fill in
a diary detailing sleep and meals eaten.
The subjects entered individual rooms at 13:00 h on
the first day and stayed under a dim light condition
(<50 lx, 3,000 K). A curtain was closed and ceiling lights
were filtered for the condition. They continuously wore
the Actiwatch and their sleep (including sleep efficiency
and sleep latency) was monitored throughout the ex-
periment. During days 2 to 4, food and drink intake
was regulated (see above), and subjects were required
to refrain from watching any video display monitors
(cell phone, computer, or TV) in order to regulate
light exposure and sensitivity to light. They spent all
their time in the chair between 07:0000:00 h except
for using a bathroom. All subjects were allowed to
read a book or magazine and/or listen to music, but not
to take naps. Breakfast was served between 07:30 h and
08:00 h, lunch between 12:10 h and 12:40 h, and supper
between 19:10 h and 19:40 h. Subjects had 200 mL of
water with each meal and were free to drink only water
between meals.
Table 1 The four experimental groups
Group Poor*Dim Rich*Dim Poor*Bright Rich*Bright
Number of subjects 10 7 9 7
Age (years)*
1
22.1 (4.6) 21.4 (2.5) 21.0 (2.4) 21.4 (2.1)
Height (cm)*
1
170.9 (6.1) 175.1 (5.1) 173.1 (4.0) 174.3 (6.3)
Weight (kg)*
1
57.7 (5.6) 62.1 (7.4) 64.1 (7.1) 62.0 (5.8)
BMI*
1
19.9 (2.3) 20.3 (2.9) 21.2 (1.9) 20.6 (2.5)
Tryptophan content of breakfast Poor*
2
Rich*
3
Poor*
2
Rich*
3
Lighting condition during the daytime Dim*
4
Dim*
4
Bright*
5
Bright*
5
*
1
Mean (SD) is shown for age, height, weight, and BMI.
*
2
Tryptophan content of 55 mg.
*
3
Tryptophan content of 476 mg.
*
4
< 50 lx between 07:30 h and 18:00 h.
*
5
> 5,000 lx between 07:30 h and 18:00 h.
Table 2 Energy and nutrient intake at breakfast, lunch,
and supper
Energy and nutrients Breakfast Lunch Supper
TRP-Poor TRP-Rich
Energy*
1
(kcal) 378 579 810 723
Tryptophan (TRP) (mg) 55 476 285 318
Protein (g) 5.6 36.2 25.6 27.4
Lipid (g) 7.7 17.5 23.1 16.4
Carbohydrate (g) 71.7 65.2 121.6 113.9
Sodium (mg) 813 1118 1742 941
Vitamin B6 (mg) 0.22 0.61 0.49 0.7
Isoleucin (mg) 162 1649 1021 1084
Leucin (mg) 311 2855 1826 1964
Phenylalanine (mg) 200 1731 1028 1096
Tyrosine (mg) 149 1388 774 898
Valine (mg) 223 1936 1177 1298
LNAA*
2
(mg) 1045 9559 5826 6340
TRP/LNAA (mol/mol) 0.036 0.034 0.033 0.034
*
1
All energy and nutrient intakes of the controlled food in the experiment
were calculated according to the Standard Tables of Food Composition in
Japan, and the coefficients for each food constituent were different from
Atwaters coefficients.
*
2
LNAA: Large, neutral amino acids (isoleucin, leucin, phenylalanine, tyrosine,
and valine).
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For days 2 to 5, subjects rose at 07:00 h and answered
the OSA sleep inventory [17] after a urine sample
had been collected. Subjects in groups Poor*Dim and
Rich*Dim remained under dim light between 07:30 h and
18:00 h whereas subjects in groups Poor*Bright and
Rich*Bright stayed under bright light (>5,000 lx) with
natural sunlight by opening curtain and artificial light by
having a lighting device (Bright Light Me, Solartone Ltd.,
Tokyo, Japan) placed in front of them. Under the bright
light condition, natural light changed color temperature ac-
cordingly. It was not compulsory for subjects to gaze at the
lighting devise. Vertical illuminance and color temperature
at eye level were checked for dim and bright conditions
by using an illuminometer (Photo Recorder PHR-51,
T & D Co., Nagano, Japan) and an illuminance spec-
trophotometer (CL-500A, Konica Minolta, Inc., Tokyo,
Japan) respectively.
Saliva was collected into collection tubes with a pure
cotton swab (Sarstedt AG & Co., Tokyo, Japan) every
hour between 18:00 h and 00:00 h on days 1 to 4. Saliva
samples were immediately centrifuged at 1,500 gfor
5 min after the collection, and the collection tubes with-
out the cotton swab were then frozen at 30°C until
analysis. Subjects retired at 00:00 h after emptying their
bladder and their urine was collected between 00:00 h and
07:00 h. The urine samples were also stored at 30°C.
Subjects were free to leave after breakfast on day 5. All
urinary and salivary analyses were carried out in duplicate
and the mean values of the duplicates were used for statis-
tical analyses.
Analysis method
The following factors were analyzed: i) sleep efficiency
and sleep latency from the actigraphs analyzed by the
Actiware-Sleep software, as measures of objective sleep
quality; ii) subjective estimates of sleep quality on rising,
using the OSA sleep inventory (sleepiness, sleep main-
tenance, worries, integrated sleep feeling, and sleep initi-
ation); and iii) melatonin concentrations in the rising
urine sample and saliva samples taken in the 6 hours be-
fore retiring. Creatinine-adjusted melatonin, which was
not metabolized and was excreted in urine was repre-
sented as nighttime melatonin secretion in urine.
Urine samples were analyzed with RIA kits (RK-MEL2
200 tests, Bühlmann Laboratories AG, Schönenbuch,
Switzerland). The mean coefficients of variation (CV) of
the intra- and inter-assay precision were 7.9% and 11.7%,
respectively. The limit of detection (LoD) was 0.3 pg/mL
and the limit of quantification (LoQ) was 0.9 pg/mL. Re-
garding creatinine analysis, Creatinine kits (DeterminerL
CRE, Kyowa Medex Co. Ltd., Tokyo, Japan) were used.
The mean CVs of the intra- and inter-assay precision
were <5% and <3%, respectively. The LoD was 0.04 mg/
dL and the LoQ was 0.1 mg/dL. Saliva samples were ana-
lyzed with RIA kits (RK-DSM2 200 tests, BÜHLMANN
Laboratories AG, Schönenbuch, Switzerland). The mean
CVs of the intra- and inter-assay precision were 7.9% and
9.8%, respectively. The LoD was 0.2 pg/mL and the LoQ
was 0.9 pg/mL.
The dim light melatonin onset (DLMO) was determined
by linear interpolation between adjacent saliva samples
Figure 1 Experimental protocol. Breakfast was served between 07:30 h and 08:00 h, lunch between 12:10 h and 12:40 h, and supper between
19:10 h and 19:40 h. For days 2 to 5, subjects rose at 07:00 h and answered the OSAsleep inventory after their urine sample had been collected
(indicated by a white arrow underneath the time scale). Subjects in groups Poor*Dim and Rich*Dim continuously stayed under dim light (<50 lx)
whereas subjects in groups Poor*Bright and Rich*Bright stayed under bright light (>5,000 lx) between 7:30 h and 18:00 h. Saliva was collected
every hour between 18:00 h and 00:00 h (black arrows) and subjects retired at 00:00 h after emptying their bladder.
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using a fixed threshold of 3 pg/mL [18]. For five subjects,
their melatonin levels did not rise above the absolute
threshold (3 pg/mL) during the sampling period (18:00
00:00 h). Therefore, their data were excluded from DLMO
analyses. Phase shift of saliva melatonin secretions was
considered by the difference of the DLMOs on days 1 and
4(Day1Day 4) and statistically analyzed by using one-
way ANOVA.
Two-way ANOVA for repeated measurements (Group
and Day, Time and Group, or Time and Day) was used
for sleep efficiency and sleep latency from the actigraphs,
the OSA sleep inventory, and saliva and urine melatonin
concentrations; in addition, for saliva melatonin concen-
trations, three-way ANOVA for repeated measurements
(Group, Day, and Time) was also used. Differences in main
effects were investigated using Bonferroni corrections.
The analyses were performed useing SPSS (Ver. 19,
IBM, Tokyo, Japan), and a Pvalue of <0.05 was consid-
ered to be statistically significant.
Results
There were no significant differences (between days 2
and 5 and the four groups) or interactions (between Group
and Day) with regard to sleep efficiency and sleep latency,
as assessed from the actigraphs (two-way ANOVA, Group
and Day; Figure 2). However, even though there were no
significant differences between scores for the individ-
ual groups (multiple comparisons using the Bonferroni
method), ANOVA indicated that there were statistically
significant differences between the four groups overall
in the scores for sleepiness (P<0.01), sleep maintenance
(P<0.05), and worries (P<0.05) from the OSA sleep
inventory (Group and Day; Figure 3). Main effects showed
that sleepiness and worries scores differed significantly
with Day (sleepiness, F (1, 33) = 6.0, P<0.05; worries,
F (1, 33) = 10.9, P<0.01) when they were compared on
the second and fifth days. The sleepiness score was 27.3 ±
1.0 (mean ± SE) on the second day and 29.3 ± 1.0 on the
fifth day, and the worries score was 27.7± 0.9 on the sec-
ond day and 30.2± 1.0 on the fifth day, which means that
sleep quality was better (estimated as the result of sleepi-
ness) and they were less worried on the fifth day com-
pared to the second. Furthermore, although there were no
significant differences between the scores on days 2 and 5,
in Rich*Bright, the mean scores of sleepiness, sleep main-
tenance, worries, integrated sleep feeling, and sleep initi-
ation on day 5 were higher than on day 2.
Regarding saliva melatonin concentration, there was a
significant interaction between Group and Day (F (3, 29) =
6.2, P<0.01), but no interaction was found in relation to
Time (three-way ANOVA; Group, Day, and Time). Saliva
melatonin concentrations showed significant differences
according to the time of the evening (F (6, 174) = 49.6,
P<0.01).
In a further analysis of the effects of tryptophan and
light condition upon saliva melatonin, concentrations
duringtheeveningsofdays1and4wereseparately
assessed by ANOVA for differences between Group and
Time (Figure 4). On day 1, there were no significant dif-
ferences between the four groups whereas, on day 4,
the concentrations in the groups differed significantly
(P<0.01). Multiple comparisons showed that, on day 4,
there were significant differences between Poor*Dim and
Rich*Bright (P<0.01) and Rich*Dim and Rich*Bright
(P<0.01). Furthermore, saliva melatonin concentrations
during the evenings of days 1 and 4 were compared by
ANOVA for differences between Time and Day (Figure 5).
There were significant differences between the concentra-
tions on days 1 and 4 in the groups Poor*Dim (F (6, 54) =
5.0, P<0.05) and Rich*Bright (F (6, 36) = 2.9, P<0.05), but
not in Rich*Dim and Poor*Bright. Compared with the first
day, saliva melatonin secretion on the fourth day was
lower in Poor*Dim but higher in Rich*Bright.
On days 1 and 4, the DLMO was considered for phase
shifts of melatonin secretion. Table 3 shows the DLMOs of
the four groups. There were significant differences between
the shifts of DLMOs (Day 1Day 4) in the four groups
(one-way ANOVA, Group, P<0.01). Multiple compari-
sons showed that there were significant differences be-
tween Poor*Dim and Poor*Bright (P<0.01), Poor*Dim
Figure 2 Mean score (and SDs) for sleep efficiency (a) and latency (b) of the four groups on days 2 (white bars) and 5 (filled bars).
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and Rich*Bright (P<0.01), Rich*Dim and Poor*Bright
(P<0.01), and Rich*Dim and Rich*Bright (P<0.01). The
DLMOs were significantly advanced by bright light expos-
ure but not by a tryptophan-rich breakfast.
Creatinine-adjusted melatonin in urine in the rising
samples taken on days 2 and 5 are shown in Figure 6.
In groups Poor*Dim and Rich*Dim, the mean values
of the concentrations were lower on day 5, whereas,
in Poor*Bright and Rich*Bright, they were higher on
day 5. These results showed the same direction as saliva
melatonin secretion. However, there were no statisti-
cally significant differences between the days, and no
significant interaction between the Group and Day
(two-way ANOVA).
Figure 3 Mean score (and SDs) for scores of the OSA sleep inventory in the four groups on days 2 (white bars) and 5 (filled bars).
(a) Sleepiness, (b) Sleep maintenance, (c) Worries, (d) Integrated sleep felling, and (e) Sleep initiation.
Figure 4 Average saliva melatonin concentrations during the evenings of day 1 and day 4.
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Discussion
The effects of a tryptophan-rich or tryptophan-poor
breakfast and bright or dim light exposure during daytime
upon sleep efficiency and latency (using actigraphy), five
subjective aspects of sleep (from the OSA sleep inventory),
and saliva and urine melatonins have been examined.
Although there were no significant group differences
between saliva melatonin concentrations on the first day,
significant differences were present on the fourth (Figure 4).
The significant differences were between the Poor*Dim
and Rich*Bright groups and between Rich*Dim and
Rich*Bright groups (in both cases being higher in the
Rich*Bright group). These results indicate that a tryptophan-
rich breakfast and bright light exposure during daytime
could promote saliva melatonin secretion at night; further,
the observations that the Rich*Bright group had higher
melatonin concentrations than the Rich*Dim group des-
pite no significant differences between Poor*Dim and
Rich*Dim nor Poor*Bright and Rich*Bright, suggests that
bright light exposure in the daytime is an important con-
tributor to raised melatonin levels in the evening, as has
previously been found by others [19-22]. In addition, the
DLMOs of saliva samples were significantly advanced by
bright light exposure (Table 3). It is reported that bright
Figure 5 Average saliva melatonin concentrations (and SDs) during the evenings of days 1 and 4 in each group. Note that the data
show the same as those in Figure 4.
Table 3 DLMOs of the four groups on days 1 and 4
Group Number of
subjects
DLMO*
1
Day 1 Day 4 Day 1Day 4 (min)
Poor*Dim 8 21:53 (54) 22:12 (58) 19 (13)
Rich*Dim 6 21:30 (60) 22:36 (47) 66 (14)
Poor*Bright 7 21:23 (64) 19:57 (75) 86 (32)
Rich*Bright 7 21:38 (60) 20:07 (52) 90 (53)
*
1
Mean time of day and SDs are shown for DLMO on days 1 and 4.
Figure 6 Mean creatinine-adjusted melatonin concentration
(and SDs) of the four groups in the rising sample taken on days
2 (white bars) and 5 (filled bars).
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light exposure (5,000 lx) in daytime (11:0017:00 h) raised
the following melatonin secretion at night and advanced
its phase shift [19]; the results herein were in agreement
with this report.
As shown in Figure 5, comparisons of saliva melatonin
concentrations on the first and fourth days showed sig-
nificant differences in groups Poor*Dim and Rich*Bright
but no significant differences in groups Rich*Dim and
Poor*Bright. The differences between days 1 and 4 were
in the opposite direction in groups Poor*Dim and
Rich*Bright; in the Poor*Dim group, melatonin secre-
tion on the fourth day was lower than on the first day,
whereas it was higher on the fourth day in Rich*Bright. It
remains to be investigated if this change in the time-
course of melatonin secretion observed in the evening
continues until the following morning that is, if there
are phase advances/delays in the secretory profile and/
or if there are changes in the total amount of melatonin
secreted during the course of the whole night. How-
ever, in urine melatonin secretion, compared with the
second day, the mean melatonin concentration on the
fifth day was lower in Poor*Dim but higher in Rich*Bright
(Figure 6). Although not statistically significant, these re-
sults show the same direction as found in saliva concen-
tration (Figure 5). Therefore, the changes in melatonin
secretion in saliva could indicate an increase of melatonin
secretion and/or a phase shift of melatonin secretion. This
point will be investigated by measuring saliva melatonin
concentrations after midnight in a further study. Further-
more, only in groups Poor*Dim and Rich*Bright there
were significant differences between saliva melatonin
concentrations on days 1 and 4, and no significance was
found in groups Rich*Dim and Poor*Bright (Figure 5).
These results indicate that a combined effect of trypto-
phan intake and bright light exposure an increase of sal-
ivary melatonin after a tryptophan-rich breakfast was
augmented by bright light exposure during the daytime
and vice versa provides information that can be used to
promote a healthy lifestyle.
Increased melatonin secretion raises the issue of changes
in sleep. Although no significant differences were found
for objective measures of sleep efficiency and latency, sub-
jective feelings of sleepiness and worries were significantly
improved on the fifth day compared to the second. In
addition, only in Rich*Bright, all mean scores (sleepiness,
sleep maintenance, worries, integrated sleep feeling, and
sleep initiation) on day 5 were higher than on day 2. Al-
though there were no significant differences between the
scores on days 2 and 5, it is possible that tryptophan in-
take and bright light exposure improve subjective sleep
quality. However, since there were no significant differ-
ences between the four groups, it is also possible that
there was a general improvement in sleep over the course
of the experiment, unrelated to the treatments. This
improvement might result from the subjects becoming ac-
customed to the conditions in the experimental chamber.
Furthermore, effects of light and tryptophan might depend
upon the age and sensitivity of subjects [13,23]. Since the
subjects were young and healthy, the tryptophan and light
effects might not be strong enough to cause clear sleep
changes, although our bright condition (>5,000 lx) was
bright enough to occur only rarely in most individuals
daily lives. The generality of the present findings remains
to be elucidated; for example, whether these results apply
to females, older subjects, and non-Japanese populations
whose basic diet is different.
It should also be considered why a tryptophan-rich
breakfast and daytime exposure to bright light had no
effects on objective (actigraphic) measures of sleep effi-
ciency and latency. One reason might be due to limita-
tions of the Actiwatches, which, although convenient for
the subject, do not assess sleep architecture, they overesti-
mate sleep latency, total sleep time, and sleep efficiency,
and underestimate intermittent awakenings [24,25]. An-
other reason might be the sample size used in this study; a
larger sample size would help increase the accuracy of
data analysis and the small group sizes might have led to
the present negative results (a Type 2 error). In addition,
Silber and Schmitt [23] suggested that, in healthy adults,
tryptophan intake during the daytime had a relaxing and
calming effect whereas taking it at night might have only a
minimal effect on the subsequent sleep [23]. Therefore,
the effects of the timing of tryptophan intake upon mood
and sleep require further study.
The study design is also limited with regard to an ab-
sence of complete control of the food eaten at breakfast;
although the TRP/LNAA ratio was controlled, as shown
in Table 2, other nutrients and calorie intake differed. It
is possible that these other factors affected the OSA
scores and melatonin secretion. A stricter regulation of
the subjectsdiet would be required to investigate these
effects, even though complete control of the diet would
be difficult to achieve. One way to investigate an effect
of tryptophan independent of other nutrients is to ad-
minister pure tryptophan without food manipulation, as
was done by Markus et al. [13,14]. Their results showed
that pure tryptophan and tryptophan-rich protein hy-
drolysate had greater effects on the plasma TRP/LNAA
ratios and brain tryptophan availability, and thus caused
mood and performance improvements. The effects of
pure tryptophan or tryptophan-rich protein hydrolysate
on melatonin secretion and sleep will be investigated in
our next study.
Conclusions
This study is the first to report quantitative effects of
changed tryptophan intake at breakfast combined with
daytime light exposure on melatonin secretion and sleep
Fukushige et al. Journal of Physiological Anthropology 2014, 33:33 Page 8 of 9
http://www.jphysiolanthropol.com/content/33/1/33
quality. Evening saliva melatonin secretion changed sig-
nificantly and indicated that a tryptophan-rich breakfast
and bright light exposure during the daytime promoted
melatonin secretion at this time.
Abbreviations
CV: Coefficients of variation; DLMO: Dim light melatonin onset; LoD: Limit of
detection; LoQ: Limit of quantification; Poor*Bright: Tryptophan-poor
breakfast and bright light environment; Poor*Dim: Tryptophan-poor breakfast
and dim light environment; Rich*Bright: Tryptophan-rich breakfast and bright
light environment; Rich*Dim: Tryptophan-rich breakfast and dim light
environment; TRP/LNAA: Tryptophan/large neutral amino acids ratio.
Competing interests
The authors declare that they have no competing interests.
Authorscontributions
The authors alone are responsible for the content and writing of the paper.
All authors read and approved the final manuscript.
Author details
1
Department of Human Health Sciences, Graduate School of Medicine, Kyoto
University, 53, Shogoin, Kawahara-cho, Sakyo-ku, Kyoto 606-8507, Japan.
2
Department of Environmental Science, Fukuoka Womens University, 1-1-1,
Kasumigaoka, Higashi-ku, Fukuoka 813-8529, Japan.
3
Department of Living
Environmental Science, Fukuoka Womens University, 1-1-1, Kasumigaoka,
Higashi-ku, Fukuoka 813-8529, Japan.
4
Laboratory of Environmental
Physiology, Graduate School of Integrated Arts and Sciences, Kochi
University, 2-5-1, Akebonocho, Kochi 780-8520, Japan.
5
Department of Food
and Nutrition, Junshin Junior College, 1-1-1, Chikushigaoka, Minami-ku,
Fukuoka 815-8510, Japan.
6
Comprehensive Housing R & D Institute, Sekisui
House Ltd, 6-6-4, Kabutodai, Kizugawa-city, Kyoto 619-0224, Japan.
Received: 21 May 2014 Accepted: 23 October 2014
Published: 19 November 2014
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doi:10.1186/1880-6805-33-33
Cite this article as: Fukushige et al.:Effects of tryptophan-rich breakfast
and light exposure during the daytime on melatonin secretion at night.
Journal of Physiological Anthropology 2014 33:33.
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