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Short communication
The effects of Hericium erinaceus (Amyloban
®
3399) on sleep quality
and subjective well-being among female undergraduate students: A
pilot study
Hisayoshi Okamura, Ph.D.
a
,
*
, Nobuko Anno, Ph.D.
b
, Akira Tsuda, Ph.D.
c
,
Takahiro Inokuchi, Ph.D.
d
, Naohisa Uchimura, M.D., Ph.D
a
,
e
,
Kazutoyo Inanaga, M.D., Ph.D.
f
a
Cognitive and Molecular Research Institute of Brain Diseases, Kurume University, Kurume, Fukuoka, Japan
b
Department of Food and Nutrition, Kyushu Nutrition Welfare University, Kitakyushu, Fukuoka, Japan
c
Department of Psychology, Kurume University, Kurume, Fukuoka, Japan
d
Research Institute of Medical Mass Spectrometry, Kurume University School of Medicine, Kurume, Fukuoka, Japan
e
Department of Psychiatry, Kurume University School of Medicine, Kurume, Fukuoka, Japan
f
Chikusuikai Institute for Neuroinformation, Chikusuikai Hospital, Yame, Fukuoka, Japan
article info
Article history:
Received 27 January 2015
Received in revised form
30 March 2015
Accepted 3 April 2015
Keywords:
Hericium erinaceus
Amyloban
®
3399
Sleep quality
Salivary free-MHPG
1. Introduction
Currently in Japan, one out of every four to five people suffers
from sleep disorders [1,2]. A very high percentage of undergraduate
students, who are at the last stage of adolescence, suffer from sleep
problems because of staying up late at night and sleeping until late
in the morning or maintaining irregular sleep patterns. These be-
haviors lead to disruption of circadian rhythms and deterioration of
quality of life, involving a decline in productivity due to daytime
sleepiness [3].
Compared with students who get adequate sleep (6e8 h per
night), students who habitually sleep in excess (more than 9 h per
night) or inadequately (less than 5 h per night) have a strong self-
awareness of emotional and physical distress as measured by
General Health Questionnaire (GHQ-28). In particular, the levels of
salivary free 3-methoxy-4-hydroxyphenylglycol (free-MHPG) (a
metabolite of central noradrenaline [NA]), and immunoglobulin A
(s-IgA) antibodies (which play a role in the immune system), are
substantially lower among students with excessive sleep. Our
previous study indicated that disturbance of sleep habits is closely
related to decline in subjective well-being, as well as a weakening
of immune functions and NA systems [4].
Hericium erinaceus (Lion's Mane mushroom) has long been used
for culinary and medicinal purposes. Its cognitive benefits have
recently drawn more attention, and studies have investigated the
therapeutic use of this mushroom in patients with mild dementia
[5e7]. In addition, Nagano et al. [8] reported that after 4 weeks of H.
erinaceus administration, participants' feelings of depression, irri-
tability, and fatigue significantly declined compared with those
before administration, suggesting its efficacy in improving mood.
These findings suggest that H. erinaceus may be beneficial for
people who suffer from circadian rhythm disorders and help alle-
viate sleep problems, while improving the quality of life for people
who lead an unhealthy or non-productive lifestyle by habitually
staying up late and sleeping in.
This pilot study evaluated the effects of a 4-week administration
of H. erinaceus (Amyloban
®
3399) on female undergraduate stu-
dents who were likely to have a high incidence of sleep problems.
We assessed changes in sleep quality and subjective well-being
with the GHQ-28 and PSQI (Pittsburg Sleep Quality Index).
Furthermore, we examined the level of salivary free-MHPG after
awakening, which is generally regarded as an accurate index of
chronic stress and depressive symptoms and reflects sympathetic
nervous system activity [9]. Thus, we were able to conduct a
comprehensive analysis of the effects of Amyloban
®
3399 on sleep
quality and subjective well-being of female students.
*Corresponding author. Cognitive and Molecular Institute of Brain Diseases,
Kurume University, 67 Asahi-machi, Kurume, Fukuoka 830-0011, Japan. Tel.: þ81
942 31 7581; fax: þ81 942 31 7911.
E-mail address: okamura_hisayoshi@med.kurume-u.ac.jp (H. Okamura).
Contents lists available at ScienceDirect
Personalized Medicine Universe
journal homepage: www.elsevier.com/locate/pmu
http://dx.doi.org/10.1016/j.pmu.2015.03.006
2186-4950/Copyright ©2015, International Society of Personalized Medicine. Published by Elsevier B.V. All rights reserved.
Personalized Medicine Universe 4 (2015) 76e78
2. Material and methods
2.1. Participants
The participants were eight female undergraduate students
(mean age: 21.7 ±0.4 years), all of whom had a National Dietitian
Exam scheduled within 1 month. None of the participants had a
history of serious illnesses or prescription medicine use.
2.2. Procedure
The participants were asked to visit the lab and were given a
brief description of the experimental procedures and saliva
collection method. A questionnaire sheet consisting of GHQ-28 and
PSQI questions was distributed. In addition to completing the
questionnaires, the participants were asked to fill out their grade
level, gender and age.
2.3. Measurements
2.3.1. Questionnaires
GHQ-28 [10]: This questionnaire assessed the participants'
mental health over the past week. This self-administered ques-
tionnaire consisted of 28 items, divided into four subscales: “so-
matic symptoms,”“anxiety and insomnia,”“social dysfunction,”
and “depression.”Participants responded to the questions using a
four-point scoring system. Each subscale had a seven-point
maximum. Cut-off points were the following: 2e3 points is a
mild, and 4 points and higher is considered a moderate for “somatic
symptoms”and “anxiety and insomnia,”,1e2 point score is mild,
and 3 points and higher is considered moderate symptom for
“social dysfunction”and “depression”.
PSQI [11]: This questionnaire was used to assess sleep habits and
sleep quality over the 1-month period. There were 18 items con-
taining seven components (quality of sleep, sleep duration, sleep
latency, sleep efficiency, sleep disturbance, use of sleep medication,
and excessive daytime sleepiness). Higher scores suggested poor
sleep quality, with a score of 5.5 being the cut-off point.
2.3.2. Saliva collection and free-MHPG measurement
Each participant was asked to collect saliva samples immedi-
ately upon waking. A Spitz device (“Sarisoft”) was used to collect
the samples, which were immediately stored in a 80
C freezer. All
samples were collected at the end of the study. The free-MHPG
level was measured using gas chromatography mass spectrom-
etry (Hitachi-M80B, Hitachi, Japan), as described by Yajima et al.
[12].
2.4. Administration of H. erinaceus
Participants in this study self-administered six tablets per day of
Amyloban
®
3399 (Mushroom Wisdom, Inc., East Rutherford, NJ
USA), divided into 2 or 3 doses. The supplement was taken with
food for a period of 4 weeks. Participants were not explained
anything about Amyloban
®
3399 during registration; they were
just told that “This is a kind of supplement.”
2.5. Ethical considerations
The ethics committee of the university approved this study.
Participants' safety was the priority and the research data were
used for the purposes of this study only. Participants' information
was kept confidential. They were given written and oral explana-
tions before providing their consent.
2.6. Statistical analysis
Data analysis was performed with a Windows version of SPSS
(Statistical Package for the Social Sciences). A t-test was used to
evaluate the mean difference between GHQ-28 subscale scores,
PSQI scores, and average level of salivary free-MHPG before and
after administration of Amyloban
®
3399. In each statistical analysis,
ap-value less than 0.05 was considered statistically significant, and
ap-value less than 0.10 was considered a marginally significant
difference.
3. Results
The average PSQI score before administration of Amyloban
®
3399 was 7.3. Six of the participants scored higher than the cut-off
point (5.5) and two scored below. The scores (mean ±standard
deviation) for the GHQ-28 subscales were as follows: “somatic
symptoms,”3.8 ±2.7 (mild), “anxiety and insomnia,”5±1.9
(moderate), “social dysfunction,”1.9 ±2.1 (mild), and “depression,”
1.8 ±1.3 (mild).
On the “anxiety and insomnia”subscale of the GHQ-28, there
was a declining trend after 4 weeks of supplement administration
(t¼1.86, df ¼14, p<0.10). No significant differences were observed
on the other subscales after the 4-week administration of Amylo-
ban
®
3399. There were no statistically significant differences in
PSQI scores associated with Amyloban
®
3399 use. However, after
the 4-week administration, the average score showed a decline
(pre-administration: 7.25, post-administration: 5.75), and the
number of participants who scored above the 5.5 cut off point
declined from six to four. After 4 weeks of Amyloban
®
3399
administration, levels of salivary free-MHPG significantly increased
compared with those during pre-administration (t¼2.25, df ¼14,
p<0.05; Table 1).
4. Discussion
This study comprehensively evaluated subjective ratings on the
GHQ-28 and PSQI questionnaires, as well as the objective assess-
ment of salivary free-MHPG levels, taken from 8 female under-
graduate students to assess the effects of 4 weeks of administration
of Amyloban
®
3399 on sleep quality and subjective well-being. The
average PSQI score from the eight participants before adminis-
trating Amyloban
®
3399 was 7.3 and was higher than the cut-off
point (5.5). In addition, the average score on the “anxiety and
insomnia”subscale of the GHQ-28 questionnaire before the
administration was 5 points, and the percentage of those exhibiting
moderate symptoms was high as well. These results reflect a
disturbance in sleep habits, and an increase in negative mood and
anxiety levels associated with preparations for the national exam
that all the participants were scheduled to take in about a month.
Table 1
Comparison of before and after Hericium erinaceum intake.
Before Hericium
erinaceum
After Hericium
erinaceum
pvalue
GHQ-28
Somatic symptoms 3.8 ±2.7 3.4 ±1.7 p¼0.745
Anxiety and insomnia 5 ±1.9 3.3 ±1.8 p¼0.084
Social dysfunction 1.9 ±2.1 1.9 ±1.5 p¼1.000
Depression 1.8 ±1.3 1.0 ±1.6 p¼0.319
PSQI 7.3 ±2.6 5.8 ±2.9 p¼0.292
Salivary free-MHPG 5.6 ±1.9 9.5 ±4.0 p¼0.029
Values are represented as mean ±standard error.
The pvalues indicate the difference between before and after Hericium erinaceum
intake.
H. Okamura et al. / Personalized Medicine Universe 4 (2015) 76e78 77
After 4 weeks of Amyloban
®
3399 use, the “anxiety and
insomnia”score decreased. PSQI scores also decreased, although
this difference was not statistically significant. Inanaga [13] has
reported that intake of Amyloban
®
3399 improves negative mood
such as irritability and anxiety, and raises incentive associated with
improved concentration and motivation. In addition, he also re-
ported that Amyloban
®
3399 was effective in improving symptoms
of sleep apnea [13]. These findings suggest that taking H. erinaceus
(Amyloban
®
3399) could improve negative mood and sleep disor-
der symptoms. However, 2 out of 8 participants in this study did not
have sleep problems as assessed by PSQI. Therefore, this study may
have lacked the statistical power necessary to examine the effects
of Amyloban
®
3399 on sleep disturbances. Further studies will be
required to investigate the effects of Amyloban
®
3399 on circadian
rhythm sleep disorders and/or sleep disturbance using a larger
number of participants.
In this study, the level of salivary free-MHPG before adminis-
tration of Amyloban
®
3399 was 5.6 ±1.9 ng/ml. This was low in
comparison with the levels from healthy participants in our pre-
vious studies (9.3 ±1.8 ng/ml) [4]. However, after 4 weeks of
Amyloban
®
3399 use, these levels increased to levels comparable
with those in healthy participants. It is suggested that the levels of
salivary free-MHPG in people with unidentified complaints are low
immediately after awakening. The possible correlation between
changes in salivary levels of free-MHPG after awakening and sub-
jective stress reaction needs to be investigated in further detail.
Shimbo et al. [14] reported that Erinacine A, which is isolated from
Hericium erinaceum, enhanced the synthesis of nerve growth factor
(NGF) by increasing the secretion of NA and catecholamines. Our
study was consistent with these findings with regard to the in-
crease in salivary free-MHPG. Furthermore, an increase in salivary
free-MHPG levels coincided with a trend towards improvement in
anxiety levels and sleep quality. These results indicated that one of
the possible effects of Amyloban
®
3399 could be an ability to bal-
ance out the mind and body. Thus, the administration of H. erina-
ceus (Amyloban
®
3399) could improve mood and circadian rhythm
sleep disorder symptoms, as well as the quality of life, in unhealthy
people (“semi-healthy”people at early stages of illness) who led an
unproductive lifestyle. However, salivary free-MHPG level is known
to be related to depressive mood and anxiety. Increased levels of
free-MHPG in conjunction with increased psychological stress have
also been reported [15,16]. Therefore, further studies would be
required to elucidate the impact of Amyloban
®
3399 on salivary
free-MHPG levels.
5. Conclusion
This pilot study assessed the effects of 4 weeks of administration
of Amyloban
®
3399 on subjective well-being and sleep quality in
female undergraduate students. The results revealed an increase in
salivary free-MHPG, which corresponded to an improvement in
anxiety and quality of sleep. Thus, we conclude that one of the
possible effects of Amyloban
®
3399 is to balance out the mind and
body. In the future, we will need to study the effects of Amyloban
®
3399 on sleep quality and everyday work, using a larger number of
student participants.
Footnotes
Amyloban
®
3399 made based on a proprietary extract called
“Amycenone”was used for this study. It contains standardized
amounts of the following compounds:
1. Hericenone (0.5%) eHericenone stimulates synthesis of nerve
growth factor, which promotes nerve protection [17,18].
2. Amyloban (6%) eFat soluble compound, which reduces the
endoplasmic reticulum stress caused by amyloid beta and helps
increase the survival of nerve cells [7].
Funding
This work was supported by a Grant-in-Aid for Young Scientists
B: 24730613 from Japanese Society for the Promotion of Science to
Hisayoshi Okamura.
Acknowledgments
The author wishes to thank Masaki Shirota, CEO of Mushroom
Wisdom, Inc., E. Rutherford, NJ, USA that developed Amylo-
ban
®
3399 as a dietary supplement, and Tomoko Nakamura, Vice
President of Sun Medica Co., Ltd., Tokyo Japan, who provided
Amyloban
®
3399 for use in this study.
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