��

The effect of Psidium guajava Linn leaf extract on Candida albicans 
adherence and the transversal strength of acrylic resin

amiyatun naini1 and Sherman Salim2
1 Department of Prosthodontic, Faculty of Dentistry Jember University, Jember - Indonesia 
2 Department of Prosthodontic, Faculty of Dentistry Airlangga University, Surabaya - Indonesia

abstract 
Denture	stomatitis	is	an	inflammation	of	oral	cavity	due	to	removable	denture	wearing.	Prevention	of	denture	stomatitis	can	be	

effectively	done	by	using	mouth	rinsing.	Currently,	Indonesian	government	is	actively	promoting	traditional	herbal	medicine	as	an	
alternative	medicine	such	as	Psidium	guajava	Linn	leaf	which	has	an	anti	bacterial	and	anti	fungal	ability.	The	purpose	of	this	study	
was	to	know	the	effective	concentration	and	soaking	duration	to	reduce	Candida	albicans	without	lowering	transversal	strength	of	
acrylic	resin.	This	experimental	laboratory	study	was	using	heat	cured	acrylic	resin	plate	without	surface	polishing.	The	concentration	
of	Psidium	guajava	Linn	leaf	extract	used	in	this	study	were	32%,	34%,	36%,	and	38%	respectively	with	15	minutes,	30	minutes,	1	
hour	and	8	hours	soaking	duration.	The	transversal	strength	was	measured	in	the	same	concentration	with	2	days,	10	days	and	60	days	
soaking	duration.	Sterile	aquadest	was	used	as	control.	Two	direction	ANOVA	and	LSD	test	were	used	in	data	analysis.	The	result	
showed	 significant	 difference	 in	 the	 number	 of	 Candida	 albicans	 colony	 among	 concentrations	 and	 soaking	 durations.	 Significant	
difference	 was	 also	 found	 in	 transversal	 strength	 among	 concentrations	 and	 soaking	 durations.	 It	 is	 concluded	 that	 the	 extract	 of	
Psidium	guajava	Linn	leaf	in	38%	concentration	with	8	hours	soaking	duration	will	lower	the	Candida	albicans	colony,	whereas	38%	
concentration	with	60	days	soaking	duration	will	lower	the	transversal	strength	but	it	is	still	above	the	standard	value.

Key words:	Psidium	guajava	Linn,	acrylic	resin,	Candida	albicans,	transversal	strength

Correspondence: Amiyatun Naini, c/o: Bagian Prostodonsia, Fakultas Kedokteran Gigi Universitas Jember. Jln. Kalimantan no. 37 
Jember 68121, Indonesia. E-mail: amiyatunnaini_drg@yahoo.co.id

introduction 

In dentistry, up to the present time, denture base acrylic 
resin is still widely used besides denture base metal due to 
the relatively cheap price, easily manipulated, non toxic, 
easily reparation process when it is broken, and the small 
dimensional changes,1,2 while the disadvantages of acrylic 
resin are: porous, water absorption ability, easily fracture 
and broken.1 

Denture is a good place for food debris accumulation. 
Denture base acrylic resin can be the place of stain, tartar 
and plaque accumulation. Accumulative plaque and food 
debris will increase the number of Candida	 albicans, 
therefore the endotoxin product can penetrate into mucous 
membrane and cause inflammation which is called denture 
stomatitis.3,4 It is found that 65% of elderly population are 
wearing denture, two thirds of 65% of denture users suffer 
from denture stomatitis.5-7

Prevention of denture stomatitis including pathogenic 
Candida	albicans	decreasing is very important. Candida	
albicans infection can be prevented by good cleaning. 
Removable denture cleaning can be done in two ways i.e 
mechanically and chemically. Mechanically cleaning can 
be done by brushing or using ultrasonic cleaner. Chemically 
cleaning can be done by soaking the denture into cleaning 
solution. The soaking duration in cleaning solution can be 
done for the whole night, 1 hour, 30 minutes or 15 minutes 

depending on the type of material.3 Denture cleaning 
solution is classified into five kinds i.e: alkaline peroxide, 
alkaline hypochlorite, inorganic acid, disinfectant and 
enzyme.8

 Due to relatively expensive price of chemical cleaning 
solution, traditional herb can be used as alternative cleaning 
solution for denture base acrylic resin. Some traditional 
herbal medicines such as leaf of Psidium	guajava	Linn9 can 
be used as denture cleaning solution and has an antiseptic 
and disinfectant function. The simple name of Psidium	
guajava	Linn is Psidii folium. The chemical contents of the 
leaf are tannin, atsiri oil, cuercetine, 3–arabino piranosida, 
guayaverin, leukocianidine, anritoxide, avicularin, galate 
acid. The data taken from fitochemical laboratory showed 
that the mean concentration of galate acid was equivalent 
to 7.71%, and 2.33% cuercetine of 96% ethanol extract. 
The result of pharmacological study showed that extract 
of Psidium	guajava	Linn	leaf has an antimicrobial effect 
against Staphylococcus sp.10 The use of tannin is as an 
astringent, whereas atsiri oil and other active material 
as an antibacterial substance. Galate acid functions as 
antibacterial, antiviral, antifungal, anti inflammation, anti 
tumor, anti anaphilactic, anti mutagenic and it is usually 
used as astringent.11 Galate acid is including phenol group. 
Phenol can kill vegetative cell, fungi and bacteria forming 
spores by protein denaturation and lowering the surface 
tension, so that bacterial permeability will increase.12 If 



�� Dent. J. (Maj. Ked. Gigi), Vol. 41. No. 1 January-March 2008: 25-29

phenol group is used for acrylic resin soaking, either as 
cleaning or anti bacterial solution, the physical characteristic 
of denture base acrylic resin will be changed resulting in 
transversal strength decrease.13 

According to Department of Health, Republic of 
Indonesia, extract of Psidium	 guajava	 Linn leaf in 30%, 
40%, 50%, 60% concentration will inhibit the growth of 
S.	typhi, S.	paratyphi A and S.	paratyphi B.14 Up to now 
there is no study on mycological effect of extract of Psidium	
guajava	 Linn leaf against Candida	 albicans, meanwhile 
acrylic resin can be adversely affected by phenol. Based 
on the above issue, a study on soaking duration of acrylic 
resin plate in various concentrations of Psidium	guajava	
Linn leaf extract against Candida	albicans and transversal 
strength is necessarily done.

material and method 

This study was experimental laboratory using the post 
test–only control group design. The samples were: 10 ×  
10 × 1 mm acrylic resin plate for Candida	 albicans test 
and 65 × 10 × 2.5 mm for transversal strength.15 The 
study was taken place in Laboratorium Dasar Bersama 
(LDB), Laboratory of Dental Technology, Fitochemical of 
Pharmacy Faculty, Laboratory of Microbiology Faculty of 
Dentistry Airlangga University.

The acrylic resin plates were prepared using 10 ×  
10 × 1 mm stippled wax. Samples for Candida	 albicans 
test were not polished. Brass plate master model, 65 × 10 ×  
2.5 mm in size was prepared for transversal strength test, 
then hard gypsum mold was made by mixing 100 gr of 
gypsum powder and 24 ml of water (based on manufacturer's 
recommendation). The mixture was manually steered for  
15 seconds and put into the cuvet on the vibrator. Stippled 
wax was put on the gypsum and left for 15 minutes. 
The gypsum surface was covered by vaseline, then the 
upper cuvet was filled by gypsum on the vibrator. The 
wax was removed by pouring hot water until it was 
clean. The next step, mould was filled by acrylic resin 
i.e powder and acrylic resin liquid in 5.75 gr : 2.5 ml 
(based on manufacturer's recommendation) was prepared 
in porcelain bowl and steered at room temperature until 
it reached dough phase. The mould which was already 
covered by cold mould seal filled by acrylic resin dough. 
Cuvet was closed and pressed by hydrolic press in  
22 kg/cm2 Hg pressure and then put into the curing unit. The 
process was done at 75° C for 90 minutes and continued at 
100° C for 30 minutes. The curing process was completed 
and it was left to let it cool, finally the sample was removed 
from the cuvet.

The leaves for making Psidium	 guajava	 Linn extract 
were taken from a ten year old tree, located in Jl. Kawi 8, 
Jenggawah, Jember, and identified at the Laboratory of 
Botany Pharmacy–Pharmacognocy of Pharmacy Faculty, 
Airlangga University. First, fresh Psidium	guajava	Linn leaf 

was cleaned and dried in the room. Weighed 500 gr and put 
into a crusher and given 1000 ml of ethanol, being maserated 
for 72 hours and then filtered using Buchener funnel. The 
filtrate was evaporated using vacuum evaporator. Finally, 
75 gr extract was obtained and this was 100% extract. 
The extract of Psidium	guajava	Linn leaf was weighed in  
3.2 gr, 3.4 gr, 3.6 gr, 3.8 gr then dissolved in 10 ml of 
sterile aquadest until 32%, 34%, 36%, 38% concentration 
of extract leaf was reached. 

The acrylic resin plates were washed in running water 
for 48 hours, sterilization was done using autoclave at 
121° C for 18 minutes, soaked in sterile saliva for 1 hour 
and washed by PBS twice, then, soaked in extract of 
Psidium	guajava	Linn leaf in 4 kinds of soaking duration  
(15 minute, 30 minute, 1 hour, 8 hour). In every 
concentration the soaking was repeated 7 times and 
aquadest was as control. The samples were washed by PBS 
twice, put into Sabouraud’s broth followed by 30 seconds 
vortexed. Incubation was done in Saboroud’s dextrose at 
37° C for 48 hours, continued by counting the number of 
Candida	albicans colony (cfu/ml).

For transversal strength test, all samples of acrylic resin 
plates (65 × 10 × 2.5 mm in size) soaked in aquadest for  
48 hours, followed by soaking in extract of Psidium	guajava	
Linn leaf 32%, 34%, 36%, 32%, 38% concentration for  
2 days, 10 days, 30 days and 60 days. The replacement of 
soaking solution was done in every 24 hours, then washed 
by aquadest. The samples were dried before transversal 
strength test was done using autograph AG-10Te in cross 
head speed 1/10 mm/second. The distance between the two 
holders was 50 mm. Statistical analysis was done using 
Two-Way ANOVA test, followed by LSD test. 

result 

Table 1 shows the occurrence of Candida	albicans on 
the surface of acrylic resin plate after being soaked in 32%, 
34%, 36%, 38% concentration of Psidium	guajava	Linn leaf 
extract for 15 minutes, 30 minutes, 1 hour and 8 hours.

Kolmogorof Smirnov test showed p > 0.05 which 
means the data had normal distribution. Levene test score 
was 1.411 and level of significance = 0.134 (p > 0.05) for 
concentration and soaking duration, which means the data 
was homogenous, therefore the test was continued by Two-
Way ANOVA test.

ANOVA test showed a significant difference among 
the number of Candida	 albicans colony in acrylic 
resin plate which was soaked in 32%, 34%, 36%, 38% 
concentration of Psidium	guajava	Linn leaf extract in 15 
minutes, 30 minutes, 1 hour and 8 hours (p < 0,05). There 
is an interaction of significant difference in the number 
of Candida	 albicans colony between concentration and 
soaking duration. To know further about the difference 
among the sample groups, LSD test was done and the result 
can be seen in table 2 and 3.



��Naini and Salim: The effect of Psidium guajava Linn leaf extract

There is significant difference among every concentration 
(32%, 34%, 36%, 38% and control). The increase of 
concentration will influence the number of Candida	
albicans colony and also the increase of soaking duration 
will show significant difference. The result of transversal 
strength acrylic resin plate will be shown in table 4.

Kolmogorof Smirnov test showed p > 0.05 which 
means the data had normal distribution. Levene test score 
was 1.477 with level of significance 0.106 (p > 0.05) for 
concentration and soaking duration. It means the data was 
homogeneous. Therefore data analysis was done using 
Two-Way ANOVA test.

ANOVA test showed significant difference in 
transversal strength of acrylic resin plate soaked in 32%, 

table 2.  LSD test on the number of Candida	 albicans 
on the acrylic resin plate after being soaked 
in 32%, 34%, 36%, 38% concentration of 
Psidium	 guajava	 Linn leaf extract and control  
(cfu/0.1 ml)

Group 32% 34% 36% 38% Control

Control
32%
34%
36%
38%

S
S
S
S
-

S
S
S
-

S
S
-

S
-

-

Note:  S  = significant  

table 3. LSD test on the number of Candida	 albicans 
on the acrylic resin plate after being soaked in 
Psidium	guajava	linn leaf extract for 15 minutes, 
30 minutes, 1 hour, 8 hours (cfu/0.1 ml) 

Group
15 

minutes
30 

minutes
1  hour 8 hours

 8 hours
 1 hour
30 minutes
15 minutes

S
S
S
-

S
S
-

S
-

-

Note:  S  = significant  

table 5.  LSD test on transversal strength of acrylic resin 
soaked in 32%, 34%, 36%, 38% concentration 
of Psidium	guajava	Linn leaf extract and control 
(N/mm2)

Group 32% 34% 36% 38% Control

Control
32%
34%
36%
38%

S
S
NS
NS
-

S
NS
NS
-

S
NS
-

NS
-

-

Note: S = Significant, NS =  Not Significant

table 1.  The mean of Candida colony on the surface of acrylic resin plate after being soaked in extract of Psidium	
guajava	Linn	leaf (cfu/0.1 ml)

Group

Soaking Duration

15 minutes 30 minutes 1 hour 8 hours

X SD X SD X SD X SD

Control
32%
34%
36%
38%

803.29
776.57
663.71
545.57
396.43

1.98
1.90
2.29
3.90
2.82

975.71
629.71
524.14
400.86
247.14

2.56
4.19
2.73
4.26
5.84

1021.43
565.00
496.29
281.27
126.29

5.22
3.21
1.49
3.40
3.82

1392.29
67.33
52.71
36.86
8.86

3.39
2.99
2.87
5.55
2.60

Note: X   = Mean, SD = Standard deviation

table 4.  Mean of transversal strength of acrylic resin plate soaked in Psidium	guajava	Linn leaf extract (N/mm2)

Group

Soaking Duration

2 days 10 days 30 days 60 days

X SD X SD X SD X SD

Control
32%
34%
36%
38%

182.31
176.06
170.06
168.94
168.26

9.76
14.13
7.36
8.89
9.92

167.14
166.71
166.03
164.91
163.80

4.17
10.96
5.89
9.48
6.37

163.80
162.69
161.57
159.34
158.23

6.36 
11.42
10.77
8.84
9.78

159.34
158.23
157.11
154.89
151.54

9.92
8.68

11.42
10.49
7.61

Note: X = Mean, SD = Standard deviation



�� Dent. J. (Maj. Ked. Gigi), Vol. 41. No. 1 January-March 2008: 25-29

table 6.  LSD test on transversal strength of acrylic 
resin plate soaked in Psidium	guajava	Linn leaf 
extract for 2 days, 10 days, 30 days, and 60 days 
(N/m2)

Group 2 days 10 days 30 days 60 days

60 days
30 days
10 days
 2 days

S
S
S
-

S
S
-

S
-

-

Note: S = Significant

34%, 36%, 38% concentration of Psidium	 guajava	 Linn 
leaf extract and control with soaking duration 2 days,  
10 days, 30 days and 60 days (p < 0.05). There is interaction 
between concentration and soaking duration and LSD test 
was done (Table 5). 

Significant difference was found among groups of 32%, 
34%, 36%, 38% concentration and control. The increase 
of concentration of Psidium	 guajava	 Linn leaf extract 
would decrease the transversal strength and also significant 
difference was found in soaking duration. The increase of 
soaking duration would decrease the transversal strength.

discussion 

In this study, we used one of family herbal medicine 
plants which is frequently planted in the garden and easily 
grows i.e. Psidium	guajava	Linn (guava) which contains 
galate acid and atsiri oil as anti fungal.16 One of the forces 
received by acrylic denture base is transversal strength. 
Soaking of acrylic resin plate either in cleaning or anti 
bacterial solution can alter the basic character of denture 
base acrylic resin such as transversal strength.14 

Statistical analysis Two-Way ANOVA test followed 
by LSD test shows that the higher the concentration of 
the extract and the longer the soaking duration resulting 
significant decreasing of the number of Candida	albicans. 
On the contrary the lower concentration and the shorter 
the soaking duration, the number of Candida	 albicans 
will significantly increase. It is because extract of Psidium	
guajava	Linn contains 96% of mean total of 7.71% galate 
acid. By the increase concentration, galate acid content 
would also increase including phenol in the extract 
solution so the anti Candida	 albicans will also increase. 
The duration of soaking was 15 minutes, 30 minutes,  
1 hour, and 8 hours in extract of Psidium	guajava	Linn leaf 
will also decrease the number of Candida	albicans colony 
because the increase of contact duration will increase 
the affectivity of anti microbial agent. The affectivity of 
material will be influenced by the concentration, duration 
and temperature.17 Galate acid including phenol group is 
used as anti bacterial, anti fungal, and can also be used as 
astringent.11 Chemical substance of galate acid has anti 
fungal activity against Candida	albicans.16 Based on anti 

microbial function, phenol can kill cell of vegetative fungi 
and bacteria forming spore by protein denaturation and 
decrease the surface tension so the permeability of bacteria 
and fungi will increase.12

Transversal strength of acrylic resin plate resulted 
from Two-Way ANOVA and LSD test shows the higher 
the concentration and the lower the soaking duration, the 
transversal strength of acrylic resin plate will significantly 
decrease. On the contrary, the lower the concentration and 
the shorter the soaking duration, the transversal strength 
of acrylic resin plate will significantly increase. Long 
polyester polymer consisting of repeated methyl metacrilate 
unit with high polarity is the possible cause of acrylic resin 
degradation resulting in transversal strength decrease. Ester 
in acid environment will be hydrolyzed. If polymer is 
hydrolized, degradation will occur resulting in transversal 
strength decrease.2 Phenol content in guava leaf will cause 
stretching of intra molecular bind in which sooner or later 
can break intra molecular bind, so the transversal strength 
will decrease.18 The mean of the lowest transversal strength 
is in 60 day–soaking duration (151.54 N/mm2). This value 
is much higher than recommended transversal strength of 
heat cured acrylic resin that is not less than 55 N/mm2.16 
The acrylic 2 days, 10 days, 30 days, and 60 days soaking 
duration of the acrylic resin plate is equal to 6 days, 1 month, 
3 months, and 6 months use.

It is concluded that 38% concentration and 8 hour 
soaking duration will decrease the number of Candida	
albicans	 in acrylic resin plate. Thirty eight percent 
concentration and 60 days - soaking duration will decrease 
the transversal strength of acrylic resin plate but it was still 
above the standar value that is not less than 55 N/mm2.

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