Changes in salivary oxytocin after inhalation of clary sage essential oil scent in term-pregnant women: a feasibility pilot study
Tadokoro et al. BMC Res Notes
Changes in salivary oxytocin after inhalation of clary sage essential oil scent in term-pregnant women: a feasibility pilot study
Yuriko Tadokoro 0 1 4
Shigeko Horiuchi 1 2 3
Kaori Takahata 0 1 4
Takuya Shuo 1 6
Erika Sawano 1 5
Kazuyuki Shinohara 1 5
0 St. Luke's International University , 10-1 Akashicho, Chuo, Tokyo 104-0044 , Japan
1 Trial registration The Clinical Trials Registry of University Hospital Medical Information Network in Japan- UMIN000017830. Registered:
2 Ho, Kanagawamachi, Kanazawa , Ishikawa 920-1181 , Japan
3 Graduate School of Nursing Science, St. Luke's International University , 10-1 Akashicho, Chuo, Tokyo 104-0044 , Japan
4 St. Luke's International University , 10-1 Akashicho, Chuo, Tokyo 104-0044 , Japan
5 Graduate School of Biomedical Sciences, Nagasaki University , 1-12-4 Sakamoto, Nagasaki 852-8523 , Japan
6 Hokuriku University
Objectives: This pilot study using a quasi-experimental design was conducted to evaluate the feasibility (i.e., limited efficacy, practicality, and acceptability) of our intervention protocol involving inhalation of the scent of clary sage essential oil by pregnant women and measurement of their preinhalation and postinhalation oxytocin levels. Results: Participants were women of singleton pregnancies between 38 and 40 gestation weeks (N = 11). The experiment group (n = 5) inhaled the scent of clary sage essential oil diluted 50-fold with 10 mL of odorless propylene glycol for 20 min. Regarding limited efficacy, the oxytocin level 15 min postinhalation increased in 3 women and was unmeasurable in 2. The control group (n = 6) inhaled similarly without the 50-fold dilution of clary sage essential oil. Their oxytocin level increased in 2 women, decreased in 2, and was unmeasurable in 2. Uterine contraction was not observed in both groups. Regarding practicality, 3 of the 11 women could not collect sufficient saliva. The cortisol level decreased in both groups postinhalation. The protocol had no negative effects. Regarding acceptability, burden of the protocol was not observed.
Pregnant women; Induction of labor; Complementary and alternative medicine; Aromatherapy; Clary sage essential oil; Inhalation; Salivary oxytocin; Uterine contraction; Salivary cortisol; Feasibility study
Medical induction of labor has been widely used [
However, it occasionally interferes with physiologic
childbirth, necessitating the evaluation of the effects of
complementary and alternative medicine for
stimulating labor [
]. Aromatherapy has been
complementarily used for stimulating and strengthening labor
contraction (e.g., repeated use of the scent of clary sage
essential oil) [
]. However, there are few studies that
have investigated the effects of aromatherapy on labor
Labor involves uterine contractions (UCs) caused by
]. Clary sage essential oil is thought to
stimulate labor by increasing the oxytocin level. However,
studies evaluating changes in the oxytocin level of
pregnant women by any intervention remain scarce. Thus, we
conducted this feasibility study as a basis for future larger
This research was a pilot study to determine the
feasibility of our intervention protocol in which pregnant
women inhaled the scent of clary sage essential oil. Our
primary objective was to evaluate the protocol’s
limited efficacy in terms of the oxytocin level and UC. Our
secondary objective was to evaluate the protocol’s
practicality (i.e., saliva collection ability, cortisol level, and
negative effects on participants) and acceptability (i.e.,
burden of intervention and scent perception).
This research was a feasibility pilot study conducted
using a quasi-experimental design with two arms:
experiment group and control group.
Participants and setting
The inclusion criteria included low-risk pregnant women
between 38 and 40 gestation weeks before labor onset
for planning spontaneous delivery. The exclusion criteria
included age (< 25 or > 35) and aromatherapy allergy. The
Additional file 1 shows details of participants’ criteria
and request to participants until the intervention.
Sample size was not calculated because this pilot study
aimed to provide a descriptive evaluation of the feasibility
of the intervention protocol. Thus, about five participants
for each group were considered appropriate. The first half
of the participants was assigned to the control group and
the last half to the experiment group. The feasibility pilot
study was conducted at a birth center in Tokyo, Japan
between June 2015 and August 2015.
The participants, experimental setting staff, and
biologists measuring the outcomes were masked about the
The inhalation intervention for each participant was
started at 13:00. The participants rinsed their mouth with
water, answered a questionnaire, and drank 100 mL of
water. After 10 min from the water intake, the first saliva
sample was collected [
] and the inhalation was started.
The experiment group inhaled the scent of clay sage
essential oil (Salvia sclarea, Tree of life, Tokyo, Japan)
] for 20 min. The scent was produced by bubbling
using an air pump at 2.0 mL/min in an airtight bottle
containing 10 mL of odorless propylene glycol diluted
with 50-fold of clary sage essential oil and delivered
10 cm away from the nostril [
]. The Additional file 2
provides details of the inhalation intervention.
The control group inhaled similarly using the same
device but without the dilution of clary sage essential oil
with 10 mL of propylene glycol.
For the primary objective, salivary oxytocin level was
measured at four time points: 10 min preinhalation
(baseline), and 15, 30, and 60 min postinhalation [
At each measurement point, at least 1.0 mL of saliva was
collected in a polypropylene tube by passive drool after
pooling saliva in the mouth for 3 min [
the volume was less than 1.0 mL, saliva was recollected
as in the first attempt. The participants drank 100 mL of
water again immediately after saliva collection at 30 min
postinhalation. Salivary samples were frozen in a box
containing dry ice after each collection point, and were
then stored at − 80 °C following completion of each
Oxytocin level was assayed in duplicates by
enzymelinked immunosorbent assay (ELISA; ENZO Life
Sciences, NY, USA) following the protocol of Carter et al.,
but with the addition of 500 KIU/µL aprotinin [
]. The inter-assay and intra-assay coefficients of
variability were reported to be 12.6–13.3 and 11.8–20.9%,
Objective UCs were monitored by
cardiotocography. The frequency of subjective UCs was asked from
the participants at preinhalation, and at 30 and 60 min
For the secondary objective, the volume of the
collected saliva and the number of attempts to collect saliva,
both of which constitute saliva collection ability, were
recorded. To evaluate whether our intervention protocol
can be conducted without causing stress [
], the salivary
cortisol level was measured at the same four time points
of oxytocin level measurement. Saliva was collected in a
separate tube to measure oxytocin and cortisol levels at
each measurement point. The assay was conducted by
ELISA in duplicates (Salimetrics, PA, USA) following the
Negative effects included an abnormal fetal heart rate
pattern during the intervention as monitored by
cardiotocography, and neonatal outcomes (Apgar score < 7
and neonatal intensive care unit admission) were
collected from the medical records. For the assessment of
acceptability, the burden of intervention (inhalation,
saliva and buccal mucosa collection, and participation)
in both groups and the scent perception (preference and
strength) in the experiment group were clarified at the
end of the intervention.
Some basic characteristics which were reportedly
related to a lower oxytocin level were identified [
]. The Additional file 3 shows questionnaires and oral
Outcomes were descriptively analyzed. The changes
between the preinhalation and postinhalation oxytocin
and cortisol levels and UC were compared between
groups. Additionally, changes in oxytocin levels were
statistically compared within groups by the Friedman test
and between groups by the Mann–Whitney U test with
a two-sided 5% level of significance using SPSS version
25.0J for Windows.
Eleven women received the intervention: 5 (E1–5) in the
experiment group and 6 (C1–6) in the control group. The
Additional file 4 shows the participants’ flow chart. Most
of the participants’ characteristics were similar between
the groups (Table 1).
Primary outcomes: limited efficacy
The oxytocin level could be measured at all the
measurement points in 3 women in the experiment group (Fig. 1
E1–3). Their oxytocin levels increased at 15 min and then
decreased at 30 min postinhalation. The oxytocin level
could be measured at all the measurement points in 4
women in the control group (Fig. 1 C1–4). The oxytocin
level at 15 min postinhalation increased in 2 women
and decreased in 2 women. However, there were no
significant differences in the oxytocin level changes within
groups and between groups. Postinhalation objective and
subjective UCs were not observed in both groups.
Secondary outcomes: practicality
For saliva collection, 3 of the 11 women (27.2%) failed to
collect 1.0 mL of saliva even with 4 attempts. The other
women could collect 1.0 mL or more of saliva within 3
For oxytocin level measurement, 44 saliva samples were
collected in both groups. Of the 44 samples, 9 contained
1.5 mL or more of saliva and the oxytocin level could be
measured in all of them. Of the 44 saliva samples, 35
contained less than 1.5 mL of saliva and the oxytocin level
could not be measured in 10 of these 35 samples (28.5%)
owing to the small saliva volume postcentrifugation.
The cortisol level at all the measurement points could
be measured in 3 women in the experiment group (Fig. 2
E1, E2, E4). All their cortisol levels decreased at 15 min
postinhalation. All the postinhalation cortisol levels
remained lower than baseline, except in E2. In the control
group, the cortisol level at all the measurement points
could be measured in all the women (Fig. 2 C1–6). The
cortisol levels at all the postinhalation measurement
points were lower than baseline, except in C5.
Any negative effects on the fetal heart rate pattern
during the intervention and on neonatal outcome were not
Secondary outcomes: acceptability
All the burdens of the intervention were judged as
moderate or light in both groups. All the women in the
experiment group liked the scent of clary sage essential oil. The
strength of the scent was judged as appropriate in E1 and
E3, and as weak in E2, E4, and E5 in which the oxytocin
level could be measured only in E2. The postinhalation
oxytocin level in E2 was higher than the baseline.
Experiment group (n = 5)
Control group (n = 6)
Age [SD] (years)
Gestation weeks [SD]
Education ≥ 12 years
Oxytocin receptor polymorphisms
rs 53576 (GG)
Depression (CES-D ≥ 16)
Oxytocin receptor single nucleotide polymorphism was assayed using buccal mucosa samples collected after completing all saliva collections (TaqMan SNP
genotyping assays, Applied Biosystems, Thermo Fisher, MA, USA)
The characteristics reportedly related to a lower oxytocin level were oxytocin receptor polymorphism (GG in rs53576 and rs2254298, and TT in rs1042778) [
having depression [
29, 30, 32
], anxiety [
], a high body mass index and no children, and being married [
]. Depression and anxiety were assessed using the
Japanese version of CES-D [
] and STAI [
SD standard deviation, BMI Body mass index, CES-D Center for Epidemiologic Studies Depression Scale, A-trait Trait anxiety of State-Trait Anxiety Inventory, A-State
state anxiety of State-Trait Anxiety Inventory
p-value Mann–Whitney U test or Fisher’s exact test for comparisons between groups
Experiment group Control group
Fig. 1 Changes in preinhalation and postinhalation oxytocin levels. Left: Experiment group, n = 5, E4 and E5 are not shown because the oxytocin
levels at all the time points were unmeasurable; Right: Control group, n = 6, C5 and C6 are not shown because the oxytocin level at all the time
points were unmeasurable. Comparison within groups, no significant differences in the oxytocin levels were found between preinhalation and
postinhalation (experiment group: p = 0.241; control group: p = 0.682). Comparison between groups, no significant differences were found in
the preinhalation levels of oxytocin (p = 0.310) and the changes between preinhalation and 15 min (p = 1.000), 30 min (p = 0.190), and 60 min
(p = 0.690) postinhalation
Limited efficacy: oxytocin level and uterine contraction
Inhalation of the scent of clary sage essential oil induced
an increasing trend in the oxytocin level at 15 min
postinhalation which was measured during the
inhalation. This increase, however, could not be maintained at
30 min postinhalation measured at 10 min after
finishing the inhalation. Thus, future studies should measure
the oxytocin level at 15 min postinhalation and during or
immediately after the inhalation of the scent of clary sage
essential oil to evaluate the effects.
Uterine contraction was expected to be induced in
accordance with the increase in the oxytocin level.
However, postinhalation subjective and objective UCs were
not observed in the experiment group. Regarding
oxytocin receptors, their amount increases around the onset
of labor [
] and their polymorphism is related the
duration of labor and delivery [
]. Therefore, UC is affected
by the oxytocin level as well as the amount and
polymorphism of oxytocin receptors. Another possibility is
that the increase in the oxytocin level might be small to
cause UCs. Clary sage essential oil contains sclareol and
is expected to have estrogen-like effects because sclareol
has a structure similar to estrogen [
]. As estrogen
enhances the release of oxytocin [
], it can be expected
that a greater absorption of sclareol into the body would
mean a larger increase in the oxytocin level. The present
study used inhalation as an absorption route of the
essential oil into the body, although skin absorption is also
used in aromatherapy. In future studies, administration
methods that facilitate the simultaneous skin and
inhalation absorption of essential oils such as using a footbath
should be adopted for cumulative effects on oxytocin
Practicality: saliva collection ability, cortisol level,
and negative effects
In all the samples with 1.5 mL or more of saliva, the
oxytocin level could be measured. Notably, 27.2% of women
failed to collect 1.0 mL of saliva even with 4 attempts.
The oxytocin level measurement can be further improved
by collecting a minimum amount of 1.5 mL of saliva.
Moreover, the 27.2% of women with deficient saliva
sample should be estimated in the sample size analysis. To
obtain adequate saliva volumes, any stimuli that promote
salivation consistently at each saliva collection may serve
as a solution [
]. Notably, studies indicating the
required saliva volumes and deficiencies in the
appropriate saliva volume for oxytocin level measurement remain
scarce. Recently, the number of studies on oxytocin has
been increasing, and novel findings can also be useful for
other oxytocin studies.
The cortisol level showed a decreasing trend during
the inhalation intervention in both groups, indicating the
absence of intervention-induced stress. The intervention
protocol had no negative effects.
Acceptability: burden of intervention and scent perception
The intervention protocol was within the acceptance
level of the participants. Although 3 of 5 women judged
the scent as weak, the oxytocin level increased
postinhalation. In future studies, the same strength or a stronger
scent can be considered.
This feasibility pilot study showed that inhalation of
the scent of clary sage essential oil induced an
increasing trend in the oxytocin level, but had no effect on UC.
The intervention protocol showed good acceptability. In
terms of practicality, it can be further adjusted to enable
the collection of a sufficient amount of saliva for optimal
measurement of the oxytocin level.
• The limited efficacy of the inhalation could not be
clearly confirmed owing to some missing oxytocin
• The method of oxytocin measurement did not
include an extraction step, which has bearing on the
reliability of the measurements. Future studies should
consider the inclusion of an extraction step.
• The experiment group could easily guess the
presence of the scent of clary sage essential oil which they
liked. Thus, an increase in the oxytocin level from the
perception of any preferred scent is possible.
Additional file 1. Participants’ criteria and request to participants until
intervention. Details of the inclusion and exclusion criteria of the
participants and requests to the participants to avoid any possible disturbance
of the oxytocin and cortisol measurement by enzyme immunosorbent
assay until the intervention.
Additional file 2. Inhalation intervention. Details of inhalation
Additional file 3. Questionnaires and oral questions. Preinhalation and
postinhalation questionnaires and postinhalation oral questions translated
from Japanese to English.
Additional file 4. Participants’ flow chart. Flow chart of the participants
from recruitment to analysis of outcomes.
ELISA: enzyme-linked immunosorbent assay; UC: uterine contraction.
YT and SH designed the study, analyzed the data, and drafted the manuscript.
YT conducted the experiments and collected the data. YT, KT, and TS
organized the inhalation and saliva collection methods. TS analyzed the oxytocin
receptor polymorphisms. ES and KS arranged the measurements of the
salivary oxytocin and cortisol levels. KT, TS, ES, and KS critically reviewed and
revised the draft of the study plan and the manuscript. All authors read and
approved the final manuscript.
We are grateful to all the participants, the staff at the data collection setting,
and Ms. Kanae Kitagawa (aromatherapist and aromatherapy instructor) for
their professional assistance and advice. We are indebted to Dr. Edward F.
Barroga (http://orcid.org/0000-0002-8920-2607) for his editorial review of our
manuscript. Parts of this study were presented in a poster presentation session
at the 19th East Asian Forum of Nursing Scholars, Chiba, Japan in 2016 and
at the 57th Annual Meeting of the Japan Society of Maternal Health, Tokyo,
Japan. Our study closely adhered to the CONSORT guidelines.
The authors declare that they have no competing interests.
Availability of data and materials
All data generated or analysed during this study are included in this published
Consent for publication
All participants provided written informed consent to publish this study.
Ethics approval and consent to participate
This study was approved by the Research Ethics Committee of St. Luke’s
International University (No. 15-04). The participants provided written informed
consent to participate in the study. This study was registered in the Clinical
Trials Registry of University Hospital Medical Information Network in Japan
This study was financially supported by MEXT/JSPS KAKENHI B (26293475) and
A (17H01613), the Society for Women’s Health Science Research, and Yamaji
Fumiko Nursing Research Fund. These funding sources were not involved in
the design, intervention, data collection and analysis, and publication of this
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
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