Effects of Ylang-Ylang Essential Oil on the Relaxation of Rat Bladder Muscle in vitro and White Rabbit Bladder in vivo

Abstract

Current and primary treatment modality in overactive bladder includes the administration of anticholinergics. The demand for new agents has been rising since anticholinergics have proven to come with many side effects. This study was designed to investigate the effects of ylang-ylang essential oil (YYEO) on the relaxation of urinary bladder muscle in vitro and in vivo. Effects of YYEO were assessed on resting tension, and electrical field stimulation- and various drug-induced contraction in vitro by checking the isometric tension changes of muscle strips and same procedures were repeated in the presence of methylene blue, Nw-Nitro-L-arginine methyl ester hydrochloride (L-NAME), or N-ethylmaleimide, and in vivo. YYEO decreased significantly the contractility of strips. There was no statistically significant difference between the treated group only with YYEO and the pretreated group with YYEO and methylene blue or L-NAME. When N-ethylmaleimide was employed, there was a statistically significant decrease in the rate of contraction. In vivo studies showed the same results compared with in vitro study. The results of this study indicate that YYEO has a relaxing effect on the bladder, and such mechanism is thought to be brought about by a pathway mediated by c-AMP.

Full text

INTRODUCTION
In overactive bladder, especially neurogenic detrusor over-
activity that arises in patients with spine disorder above sacral
micturition center, there are many bladder irritative symp-
toms due to low bladder compliance, neurogenic detrusor
overactivity and high intravesical pressure (1-3). In those
cases, there are many complications including recurrent uri-
nary tract infection and vesicoureteral reflex causing upper
urinary tract injury, eventually a renal failure. Thus the pur-
poses of the management in the overactive bladder is to de-
crease the intravesical pressure to appropriate levels and to
protect the upper urinary tract. The current and primary
treatment modalities include the administration of anti-
cholinergics alone or with the intermittent catheterization
(4, 5). However, despite such methods are effective, the
administration of anticholinergics may often be ceased due
to the side effects of anticholinergics such as flushing, drowsi-
ness, constipation, erectile dysfunction, dry mouth or in high
dose, central nervous system symptoms and cardiovascular
symptoms. Also the administration of anticholinergics are
not recommended in cases of hyperthyroidism, congestive
heart failure, liver disease, kidney disease, myasthenia gravis,
and megacolon or closed angled glaucoma. Therefore the
demand for new agents with less adverse effects has been
rising accordingly (6).
Aromatherapy is often used to manage the variable symp-
toms by abnormal regulation of autonomic nervous system
such as anxiety, insomnia or immune system. Aromatherapy
may be used by smelling, inhalation, footbath, or massage,
only with a few side effects such as hypertension or diarrhea.
Ylang-Ylang essential oil (YYEO) (Cananga odorata) is used
worldwide as a perfume. It can also be used as intestinal anti-
spasmodics (7, 8). Recent studies indicate the some compo-
nents in YY oil have vasodilator properties and relaxing effects
on smooth muscle (9-11).
This study was designed to investigate the effects of YYEO,
which is currently used in aromatherapy on the relaxation of
urinary bladder muscle in vitro and in vivo.
METERIALS AND METHODS
Preparation of tissue strips for in vitro study
Male Sprague-Dawley rats weighing 250 to 300 g were used
and anesthetized by intraperitoneal sodium pentobarbital (5
mg/kg) and exsanguinated via cervical dislocation. Following
*
Departments of Urology, Anesthesiology
*
and
Phamacology
, Dankook University College of
Medicine; Department of Chemistry
, Dankook
University College of Natural Science, Cheonan;
Department of Radiology
, Miz-medi Hospital,
Seoul, Korea
Hyung Jee Kim, M.D.
Department of Urology, Dankook University
College of Medicine, 29 Anseodong, Cheonan,
Korea
Tel : +82.41-550-6630, Fax : +82.41-556-0524
E-mail : bluesky@dku.edu
*
The research was conducted by the research
fund of Dankook University in 2001.
409
J Korean Med Sci 2003; 18: 409-14
ISSN 1011-8934
Copyright
The Korean Academy
of Medical Sciences
Current and primary treatment modality in overactive bladder includes the admin-
istration of anticholinergics. The demand for new agents has been rising since anti-
cholinergics have proven to come with many side effects. This study was designed
to investigate the effects of ylang-ylang essential oil (YYEO) on the relaxation of
urinary bladder muscle in vitro and in vivo. Effects of YYEO were assessed on
resting tension, and electrical field stimulation- and various drug-induced contrac-
tion in vitro by checking the isometric tension changes of muscle strips and same
procedures were repeated in the presence of methylene blue, Nw-Nitro-L-arginine
methyl ester hydrochloride (L-NAME), or N-ethylmaleimide, and in vivo. YYEO
decreased significantly the contractility of strips. There was no statistically signif-
icant difference between the treated group only with YYEO and the pre-treated
group with YYEO and methylene blue or L-NAME. When N-ethylmaleimide was
employed, there was a statistically significant decrease in the rate of contraction.
In vivo studies showed the same results compared with in vitro study. The results
of this study indicate that YYEO has a relaxing effect on the bladder, and such
mechanism is thought to be brought about by a pathway mediated by c-AMP.
Key Words : Oils, Volatile; Neurogenic Bladder; Muscle Hypertonia; Cholinergic Antagonists
Received : 10 September 2002
Accepted : 23 January 2003

410
H.-J. Kim, H.-M. Yang, D.-H. Kim, et al.
a suprapubic midline incision, bladder was isolated and lon-
gitudinal detrusor muscle strips measuring approximately 1
10 mm were obtained within the Krebs’ solution with room
temperature airing 95% O
2 and 5% CO2 after bladder body
was dissected. Each strip was mounted in organ-bath contain-
ing 20 mL Krebs’ solution with the following composition
(mmol/L): NaCl, 118.1; NaHCO
3, 25.0; KCl, 4.6; KH2PO4,
1.2; CaCl
2, 2.5; MgSO4, 1.2; glucose, 11.0, and equilibrated
at 37 with 95% O
2 and 5% CO2 for resulting pH 7.4. After
the 90-min equilibration with frequent washout using fresh
Krebs’ solution, strips of detrusor muscle were loaded with
resting tension of 1 g, and optimal isometric tension was ob-
tained as evidenced by two consecutive contraction to bethane-
chol (5
10
-4
M) that differed by less than 10%.
Changes in isometric tension were measured by Grass FT03
transducer (Grass Instruments, Quincy, MA, U.S.A.) and
recorded by Grass 79E Polygraph (Grass Instruments).
Experimental procedure for in vitro study
The effects of YYEO on basal tension are reported as the
tension at the end of the 10-min incubation in the presence
of 0.05 mL YYEO. Electrical field stimulation (EFS) was deliv-
ered via platinum electrodes set on either side of the muscle
strip in each organ bath. Intramural nerves were stimulated
with a Grass 88 field stimulator (Grass Instruments) deliver-
ing biphasic square wave pulse of 50 V, 1.0 ms wide at 1 to
16 Hz with a 2-min interval between the stimulation. After
the 30-min incubation with fresh Krebs’ solution, 0.05 mL
YYEO was added to each organ bath. The tissue was incubat-
ed for 10 min and then EFS was repeated.
In separate experiments, the effects of YYEO on bethane-
chol, ATP and KCl-stimulated contractions were investi-
gated. After the 60-min incubation at 1 g tension, bethane-
chol (0.01 mM) was added to each organ bath. Peak response
was recorded, and the bath was washed out to be refilled
with Krebs’ solution and equilibrated for 30 min. Subse-
quently, after the 10-min incubation in 0.05 mL YYEO,
the stimulation with bethanechol (0.01 mM) was repeated.
The effects were also recorded. The effects of YYEO on stim-
ulation by ATP (2 mM) and KCl (127 mM) were assessed
in the same way with other strips.
In order to investigate which drugs were effective in block-
ing the YYEO-induced relaxation, other separate experiments
were performed. First, after the 60-min equilibration at 1 g
tension, the effects of YYEO on electrical field stimulation
(EFS)-, bethanechol-, ATP- and KCl-stimulated contractions
were investigated with the above method. Baths were then
washed out to be refilled with Krebs’ solution and equilibrat-
ed for 30 min. Subsequently, after the 10-min incubation in
0.001 mM methylene blue (guanylate cyclase inhibitor), the
stimulations with EFS, bethanechol, ATP and KCl were re-
peated. The effects were also recorded. The effects of L-NAME
(Nw-Nitro-L-arginine methyl ester hydrochloride, nitric
oxide synthase inhibitor, 0.001 mM) and N-ethylmaleimide
(c-AMP nonspecific inhibitor, 0.001mM) on relaxation by
YY oil were assessed in the same way in other strips to eval-
uate which mechanism was related to relaxation by YYEO.
Preparation of animal for in vivo study
Male New Zealand white rabbits weighing about 2.5 kg
were used for this investigation. After the sedation with an
intramuscular injection of ketamine (10 mg/kg), the rabbits
were anesthetized and maintained with enflurane (2.5-3.5%).
The animals breathed spontaneously. The rabbits were then
placed supine position, and the body temperature was main-
tained at 37
using a heating pad and lamp. The polyethy-
lene catheter (Clay Adams PE-50, NJ, U.S.A.) was introduced
to bladder through the urethra. The intravesical pressure was
measured by pressure transducer (Grass Instruments, Quincy,
MA, U.S.A.) and recorded by Grass 79E Polygraph (Grass
Instruments). The catheter was connected to a three-way stop-
cock, thus permitting the injection of 37
normal saline (10
mL/min). Intravesical pressure and vesical volume were then
checked, when spontaneous bladder contraction was occurred.
After half of full volume was introduced to bladder with 37
normal saline, changes of intravesical pressure to YY oil,
bethanechol and ATP-induced stimulation were investigated.
The femoral artery on one side was cannulated with polyethy-
lene catheter (Clay Adams PE-10) and continuous monitor-
ing of arterial pressure and heart rate were measured by pres-
sure transducer (Grass Instruments) and recorded by Grass
79E Polygraph (Grass Instruments), when YY oil was admin-
istered through the vein of ear. The femoral catheter was con-
nected to a three-way stopcock, thus permitting the injection
of various drugs. Catheter was filled with heparinized saline
(50 IU/2 hr) to prevent clotting. The femoral artery on other
side was ligated to obtain the maximum effect of given con-
centrated drug.
Experimental procedure for in vivo study
YYEO (0.01, 0.02, 0.05 mL) was injected intravenously,
and the change of arterial pressure and heart rate were mea-
sured with 1-hr interval. The effects of YYEO on bethanechol-
and ATP-stimulated bladder contractions were investigated.
After a minimum of 1 hr of resting, bethanechol (0.01 mM)
was injected intra-arterially and peak response of intravesical
pressure was recorded. After the 60-min equilibrium, subse-
quently YY oil (0.0001-0.01 mL) was injected intra-arteri-
ally, and following the 1-min incubation, the stimulation with
bethanechol (0.01 mM) was repeated. The effects were record-
ed. The effects of YYEO on stimulation by ATP (2 mM) were
assessed in the same way with other rabbits.
Contractile responses were expressed as gram tension per 100
mg of tissue in in vitro study and as a percentage of the active
muscle tone, compared with control in in vivo study. YYEO

Ylang-Ylang Oil in Relaxation of Bladder Muscle
411
was given as a gift from the company (Aroma Newtech, Cheo-
nan, Korea). Bethanechol chloride, ATP, KCl, L-NAME, N-
ethylmaleimide and methylene blue were purchased from Si-
gma Co. (St Louis, MO, U.S.A.). Data were expressed as the
mean (SEM), and differences were assessed using Student’s
t-test, with p<0.05 considered to indicate significance.
RESULTS
In vitro study
YYEO had no significant effect on the resting tone of smo-
oth muscle strips. The response to EFS (1, 2, 4, 8 and 16 Hz)
was significantly suppressed by YYEO from 5.3
0.6, 5.8
0.7, 11.2 1.3, 18.7 2.1, and 26.5 2.6 g/100 mg to
3.1
0.5, 3.4 0.7, 7.0 1.3, 12.0 2.5 and 16.0 3.1 g/
100 mg, respectively (p<0.05, n=19) (Fig. 1). Response to
bethanechol, ATP and KCl were significantly suppressed by
YYEO from 7.2
0.5, 4.4 0.5, and 8.6 0.8 g/100 mg to
3.5
0.4, 2.1 0.3 and 3.5 0.5 g/100 mg, respectively
(p<0.01, n=11, 22, 21, respectively) (Fig. 2). YYEO-induced
suppressions of all stimulation-induced contractility were not
significantly changed in the presence of methylen blue or L-
NAME (data not shown). All stimulation-induced contrac-
tility suppressed by YYEO was significantly restored in the
presence of N-ethylmaleimide from 1.7
0.3, 1.2 0.3 and
1.1
0.2 g/100 mg (bethanechol, ATP and KCl, respective-
ly), and 1.0
0.2, 1.0 0.2, 2.1 0.4, 3.7 0.6 and 5.8
1.0 g/100 mg (EFS) to 4.1 0.8, 2.6 0.5 and 1.8 0.2 g/
100 mg, and 1.8
0.3, 1.8 0.3, 4.1 0.8, 6.1 0.8 and
8.3
0.1 g/100 mg, respectively (p<0.05, n=10, 10, 11, 11,
Contractility (g/100 mg of tissue)
35
30
25
20
15
10
5
0
0 2 4 6 8 10 12 14 16 18
Frequency (Hz)
Control
Ylang-Ylang
Fig. 1. Frequency-contractility curves from bladder muscle strips
in male Spraque-Dawley rats. Pre-treatment with ylang-ylang oil
(0.05 mL) significantly decreases the maximum tension under
the control.
*
p<0.05 indicates significant difference from respec-
tive control, n=19 in each group.
*
*
*
*
*
Contractility (g/100 mg of tissue)
10
8
6
4
2
0
Bethanechol ATP KCl
Control
Ylang-Ylang
Fig. 2. Changes of the contraction induced by bethanchol, ATP,
and KCI after pre-treatment with ylang-ylang oil. Bethanechol, ATP,
and KCI-induced contractions are decreased by pre-treatment
with ylang-ylang oil.
*
p<0.05 indicates significant difference from
respective control (n=11, bethanechol; n=22, ATP; n=21, KCI).
*
*
*
g/100 mg of tissue
BE 0.01 mM ylang-ylang 0.05 mL
60 min
7.6
6.0
00
0
BE
g/100 mg of tissue
ATP 2 mM ylang-ylang 0.05 mL
60 min
4.2
1.7
ATP
g/100 mg of tissue
KCI 126 mM ylang-ylang 0.05 mL
60 min
8.3
0.7
KCI
Contractility (g/100 mg of tissue)
10
8
6
4
2
0
Bethanechol ATP KCl
Control
Ylang-Ylang
Ylang-Ylang with N-ethylmaleimide
Fig. 3. Changes of the contraction induced by bethanechol, ATP
and KCI after pre-treatment with ylang-ylang oil. Bethanechol,
ATP and KCI-induced contractions are decreased by pre-treat-
ment with ylang-ylang oil and N-ethylmaleimide.
*
p<0.05 indi-
cates significant difference from ylang-ylang induced reduction
of contractions (n=8, bethanechol; n=6, ATP; n=6, KCI).
*
*
*

412
H.-J. Kim, H.-M. Yang, D.-H. Kim, et al.
respectively) (Fig. 3, 4).
In vivo study
Intravesical pressure was 16.6 2.5 mmHg, when spon-
taneous bladder contraction was occurred.
During the injection of YYEO (0.01, 0.02 and 0.05 mL)
intravenously, the heart rate was not significantly changed,
but mean arterial pressure was decreased transiently (decrease;
0, 15 and 27.5 mmHg, respectively) (Fig. 5). Intravesical
pressures induced by bethanechol and ATP in the presence
of YY oil (0.0001-0.01 mL) were significantly suppressed
by the amount of the above 0.001 mL YY oil from basal to
110.1
11.4, 55.9 3.8 and 32.7 3.1%, (p<0.01, n=9)
and 90.3
7.8, 37.7 5.8 and 36.1 9.3%, respectively
(p<0.01, n=7) (Fig. 6).
DISCUSSION
YYEO is internationally used as prime component in high
quality perfumes. Forty-nine different components have been
identified by gas chromatography-mass spectrometry (GC-
MS). Major components to list are linalool, linalyl acetate,
eugenol, methyleugenol and methyl benzonate (9-11). In
Brazil, methyleugenol is used in folk medicine as a stomachic
and intestinal antispasmodic; it suggests YYEO makes smooth
muscle of gastrointestinal system relax (7). The main com-
ponent of YYEO, linalool, is used as an antibiotics as well
as an antispasmodics used to treat epilepsy. Eugenol has also
been widely used for many years as a topical, temporary treat-
ment for the pain of pulpitis (12-14). Effects of YYEO on
skeletal muscle and smooth muscle lead us to study the effect
Contractility (g/100 mg of tissue)
20
18
16
14
12
10
8
6
4
2
0
0 2 4 6 8 10 12 14 16 18
Frequency (Hz)
Control group
Ylang-Ylang
Ylang-Ylang with N-ethylmaleimide
Fig. 4. Changes of the contraction induced by electrical field
stimulation (EFS) after pretreatment with ylang-ylang oil. Bethane-
chol, EFS-induced contractions are decreased by pre-treatment
with ylang-ylang oil. Reduction of contractions are not attenuat-
ed by pre-treatment with ylang-ylang oil and N-ethylmaleimide.
*
p<0.05 indicates significant difference from ylang-ylang induced
reduction of contractions, n=11 in each group.
*
Fig. 5. Representative figure showing the effects of intra-arterial ylang-ylang (0.01-0.05 mL) on blood pressure. Administration of ylang-
ylang decreased the amplitude of blood pressure transiently.
BP (mmHg)
100
50
5 mm/sec 0
60 min
ylang-ylang 0.01 mL 0.02 mL 0.05 mL
60 min
12 sec
Intravesical pressure (percentage)
120
100
80
60
40
20
0
Bethanechol ATP
Group
Control
0.0001 mL
0.001 mL
0.01 mL
Fig. 6. Effects of pre-treated ylang-ylang oil on the response of
bethanechol, and ATP.
*
p<0.01, significant difference from
respective control (n=9, bethanechol; n=7, ATP).
*
*
*
*
a b c d
BE 0.01 mM
ylang-ylang 0.0001 mL 0.001 mL 0.01 mL
60 min 60 min 60 min
1
0.5
0
a b c d
ATP 2 mM
ylang-ylang 0.0001 mL 0.001 mL 0.01 mL
60 min 60 min 60 min
1
0.5
0

Ylang-Ylang Oil in Relaxation of Bladder Muscle
413
of it on detrusor muscle contractibility.
In this study, in vitro studies using rat bladder were per-
formed in order to investigate the basic effect of YY oil. Be-
cause of rat’s a little body volume, a relative large amount of
YYEO which was diluted in 0.1% ethanol 1 mL and 0.1%
ethanol itself, in vivo studies using the bladder of rats result-
ed in large errors on the change of intravesical pressure and
were impossible to check the change of blood pressure dur-
ing the study. Thus trials by larger animals were needed to
obtain error-free results and to observe the adverse effect on
the cardiovascular system.
In the results of this study, YYEO inhibited all stimula-
tion by EFS and all drugs to contract the bladder. The results
were the same in in vitro as well as in in vivo studies. When
large amounts, up to 0.5 mL of YY oil was administrated
intravenously, only transient and mild decrease of blood pres-
sure was found. This suggests that the side effect on the car-
diovascular system did not lead to a decrease of intravesical
pressure.
Many experiments have been carried out to determine the
effect of this oil. Elizabetsky et al. (15) used linalool in white
rats to show depression of motor neuron stimulation as well
as affecting the central nervous system (CNS) directly. The
mechanism of this effect is that the linalool combines with
glutamine, CNS excitatory neurotransmitter and inhibits
its action. Also in Buchbauer’s study (16), essential oils have
a selective effect on tissues, but as it has a nonspecific toxic
effect on cell membranes, it has no vital activity. However,
in different studies, effects on smooth muscle relaxation were
investigated. Hume (17, 18) exerts its effect by [K+]
O on L-
type Ca
2+
channels. Eugenol works as Ca
2+
-antagonist. Eugenol
depressed the constrictor response to norepinephrine (NE),
histamine and transmural nerve stimulation. Since NE and
histamine act on different and specific receptors, it is proba-
ble that dilator effect of eugenol was not mediated by the
competition for specific receptor site, but it inhibited one or
more stage of the post-receptor excitation-contraction sequence
in smooth muscle (19). Recently, it is seen that the eugenoldilol,
derived from natural eugenol made a role as a stimulator of
non-selective -adrenoreceptor blocker and -adrenorecep-
tor blocker or as a 2-adrenoreceptor selective partial ago-
nist (17, 20-22). In this experiments, we used crude oil because
we had have the purpose that this oil will be used in aro-
matherapy treatment applied via the skin through massage
or add to bath water or by inhalation. There is some degree
of absorption of the individual components into the blood,
which was calculated at between 4 and 25% when applied
to the skin of a rhesus monkey or man (23). This would be
approximately 8
10
-6
and 1 10
-5
g/mL of an essential oil
in carrier oil applied.
The smooth muscle relaxation was made by c-AMP relat-
ed by activation of adenylate cyclase and c-GMP related to
activation of guanylate cyclase in nitric oxide pathway (24).
Lis-Balchin and Hart’s study (25) showed that smooth mus-
cle relaxation by lavender essential oils was occurred by the
elevated c-AMP levels within the cell. In this study, the action
mechanism of YYEO in bladder contractility reduction is
thought to increasing c-AMP level because the c-AMP non-
specific inhibitor, N-ethylmaleimide only inhibited the re-
duction of bladder contractility by YYEO, but inhibitors
related to nitric oxide pathway did not.
In summary, YYEO is shown to have a relaxing effect on
bladder smooth muscle. The action mechanism of YYEO is
thought through the c-AMP pathway; however the exact me-
chanism is still unclear. YYEO have several key constituents,
one of which would be believed to produce its therapeutic
effect. Therefore, to clarify the YYEO in the clinical appli-
cation and make the products without the loss of its natural
scent, further research is needed to determine the effects of
each key components of the YYEO.
REFERENCES
1. Graham SD. Present urological treatment of spinal cord injury
patients. J Urol 1981; 126: 1-4.
2. Nanninga JB, Wu Y, Hamilton B. Long term intermittent catheteri-
zation in the spinal cord injury patient. J Urol 1982; 128: 760-3.
3. Levin RM. Overview of nerves and pharmacology in the bladder.
Adv Exp Med Biol 1999; 462: 237-40.
4. Moisey CU, Stephenson TP, Brendler CB. The urodynamic and
subjective results of treatment of detrusor instability with oxybuty-
nine chloride. Br J Urol 1980; 52: 472-5.
5. Thuroff JW, Bunke B, Ebner A, Faber P, De Geeter P, Hannappel J,
Heidler H, Madersbacher H, Melchior H, Schaffer W, Schwenzer
T, Stockle M. Randomized, double-blind, multicenter trial on treat-
ment of frequency, urgency and incontinence related to detrusor
hyperactivity: oxybutynin versus propantheline versus placebo. J
Urol 1991; 145: 813-7.
6. McEvoy GK. Genitourinary smooth muscle relaxant. In: Kathy L
editor, Drug Information, Vol. II. Chapter 86. Bethesda: American
society of Health-system pharmacists Inc, 1995: 2504-5.
7. Craveiro AA, Andrade CHS, Matos FJA, Alencar JW, Dantas TNC.
Fixed and volatile constituents of Croton aff. Nepetaefolius. J Nat
Prod 1980; 43: 756-7.
8. Magalhaes PJC, Criddle DN, Tavares RA, Melo EM, Mota TL, Leal-
Cardoso JH. Intestinal myorelaxant and antispasmodic effects of the
essential oil of Croton nepetaefolius and its constituents cineole,
methyleugenol and terpineol. Phytother Res 1998; 12: 172-7.
9. Stashenko E, Torres W, Martinez J. A study of compositional varia-
tion in the essential oil of ylang-ylang (Cananga odorata) during
flower development. J High Resol Chromatogr 1995; 18: 101-4.
10. Stashenko E, Martinez J, Macku C, Shibamoto T. HRGC and GC-
MS analysis of essential oil from Colombian ylang-ylang (Cananga
odorata). J High Resol Chromatogr 1993; 16: 441-4.
11. Olivero J, Gracia T, Payares P, Vivas R, Diaz D, Daza E, Geerlings
P. Molecular structure and gas chromatographic retention behav-
ior of the components of Ylang-Ylang oil. J Pharm Sci 1997; 86:

414
H.-J. Kim, H.-M. Yang, D.-H. Kim, et al.
625-30.
12. Maudsley F, Kerr KG. Microbiological safety of essential oils used
in complementary therapies and the activity of these compounds
against bacterial and fungal pathogens. Support Care Cancer 1999;
7: 100-2.
13. Ostrander FD. Accepted dental therapeutics 1969-1970. A histori-
cal review. J Am Dent Assoc 1970; 80: 589-92.
14. Lis-Balchin M, Hart S. Correlation of the chemical profiles of essen-
tial oil mixes with their relaxant and stimulant properties in man and
smooth muscle preparations in-vitro. Processing of 27th International
symposium essential oils. Vienna, Austria 1997: 24-8.
15. Elisabestsky E, Marschner J, Souza DO. Effects of linalool on glu-
tamatergic system in the rat cerebral cortex. Neurochem Res 1995;
20: 461-5.
16. Buchbauer G. Molecular interaction: biological effects and modes
of action of essential oils. Int J Aroma 1993; 5: 11-4.
17. Hume WR. Effect of eugenol on constrictor responses in blood ves-
sels of the rabbit ear. J Dent Res 1983; 62: 1013-5.
18. Hume W, Veaco N, Becker R. Concentration-dependent depression
of cellular respiration by eugenol. J Dent Res 1982; 61: 202.
19. Pohto M, Scheinin A. Microscopic observations on living dental
pulp. The effect of oil of clove and eugenol on the circulation of the
pulp in the rat's lower incisor. Int Dent J 1960; 5: 405.
20. Lin YT, Wu BN, Horng CF, Huang YC, Hong SJ, Lo YC, Cheng
CJ, Chen IJ. Isoeugenol: a selective beta1-adrenergic antagonist
with tracheal and vascular smooth muscle relaxant properties. Jpn
J Pharmacol 1999; 80: 127-36.
21. Nishijima H, Uchida R, Kameyama K, Kawakami N, Ohkubo T,
Kitamura K. Mechanism mediating the vasorelaxing action of eugenol,
a pungent oil, on rabbit arterial tissue. Jpn J Pharmacol 1999; 79:
327-34.
22. Chang PC, van Brummelen P, Vermeij P. Acute vasodilator action
of pindolol in humans. Hypertension 1985; 7: 146-50.
23. Broaugh RL, Wester RC, Bucks D, Maibach HI, Sarason R. In vivo
percutaneous absorption of fragrance ingredients in rhesus monkey
and humans. Food Chem Toxicol 1990; 28: 369-73.
24. Steers WD. Physiology and pharmacology of the bladder and urethra.
In: Walsh PC, Retik AB, Vaughan ED, Wein AJ eds, Campbell’s Urol-
ogy, 7th edn, Vol. I. Chapter 26. Philadelphia: WB Saunders, 1998:
870-906.
25. Lis-Balchin M, Hart S. Studies on the mode of action of the essen-
tial oil of lavender (Lavandula angustifolia P. Miller). Phytother
Res 1999; 13: 540-2.