JRSSEM 2021, Vol. 02, No. 02, 260 – 278

E-ISSN: 2807 - 6311, P-ISSN: 2807 - 6494

DOI : 10.36418/jrssem.v2i2.296

Formulation of Lemongrass Extract Mouthwash

(Cymbopogon Citratus) as a Non-Pharmacological

Effort in Inhibiting The Growth of Bacteria That

Cause Dental Caries

Purnama Fitri

1*

Rasipin

2

Ari Suwondo

3

Poltekkes Kemenkes Semarang, Indonesia

1,2,3

*

e-mail: purnamafitr[email protected]

1

, rasipin@gmail.com

2

, ari.suwondo@yahoo.co.id

3

*Correspondence: Purnama Fitri

Submitted

: September 2022,

Revised

: September 2022,

Accepted

: September 2022

Abstract: Caries is a dental disease characterized by damage ranging from enamel, dentin, and

pulp. Caries is caused by bacteria that stick to the teeth. The purpose of this study was to prove

the effectiveness of lemongrass in inhibiting the growth of bacteria that cause dental caries. The

pretest-posttest laboratory and field experiment consisted of a lemongrass mouthwash

intervention group with concentrations of 33%, 36%, 39% and a chlorhexidine control group,

each subject gargled for 2 minutes. Saliva was collected before and after gargling. The variables

studied were the number of colonies and bacterial inhibition. The paired test of 36% (p=0.017)

and 39% (p=0.006) concentration intervention groups was significantly different, while the

chlorhexidine control group (p=0.091) was not significantly different. The organoleptic test

results of the 36% lemongrass mouthwash had a dark brown color, slightly thick, the distinctive

smell of lemongrass stems was still present, and the taste of mint. Based on this test, the most

acceptable lemongrass mouthwash concentration is 36%. Giving a 39% lemongrass mouthwash

formulation for 2 minutes effectively reduces the number of bacterial colonies that cause dental

caries and the inhibition is better than the chlorhexidine control, while the 36% concentration is

not significantly different from chlorhexidine.

Keywords: Antibacterial, Lemongrass Extract Mouthwash, Caries-Causing Bacteria

Purnama Fitri, Rasipin, Ari Suwondo | 261

DOI : 10.36418/jrssem.v2i2.296

INTRODUCTION

Oral health is very important for every

individual. Problematic oral health can

cause impaired speech function, chewing

function, and aesthetic function which will

definitely have an impact on one's

activities. According to Federation

Dentaire International (FDI), around 90%

of the world's population experiences oral

health problems ranging from dental

caries, dental supporting tissue disease

and even oral cancer (Hontong et al.,

2016).

The oral cavity is one of the places in

the body that contains microorganisms

with the highest diversity compared to

other places. The most abundant

microorganism in the oral cavity is

Streptococcus sp which plays a role in the

beginning of the dental caries process. In

addition, the bacterial colonies found at

the beginning of plaque formation are

Streptococcus mutans bacteria which have

an important role in the development of

dental caries in animals and humans

(Juliantoni & Wirasisya, 2019).

Teeth are hard organs found in the

mouth that are used to process food when

eating, with that teeth have a function to

tear, chew and smooth a food before the

food enters the throat. Therefore, it is

necessary to care for dental care so that

the teeth are maintained and intact

(Hafizah, 2021).

If you don't care about dental health, it

can cause diseases that can damage the

tooth coating, namely dental caries. Dental

caries is a disease that affects many

children and adults, both in milk teeth and

permanent teeth. Dental caries is a dental

tissue disease characterized by tissue

damage, starting from the tooth surface,

namely from enamel, dentin, and

extending towards the pulp (Afrinis et al.,

2020).

Dental caries is an infectious disease

that affects almost 95% of the population

in the world. The dental morbidity rate

ranks as the 6th most common disease.

Oral disease in Indonesia caused by dental

caries ranks 10th highest with a prevalence

of 45.68%. Dental caries is the most

common disease in the oral cavity

(Darsono, 2020).

Cariogenic bacteria are bacteria that

have the ability to cause dental caries.

These bacteria include Actinomyces,

Lactobacillus, Streptococcus mutans,

Streptococcus sanguinis, etc. The main

bacteria causing dental caries are

Streptococcus mutans and Lactobacillus

acidophilus. S. mutans is a normal

inhabitant of the oral cavity that can turn

into a pathogen when the living

environment of the bacteria is favorable

and there is an increase in population.

Streptococcus mutans and Lactobacillus

acidophilus in the oral cavity produce

lactic acid from fermented sugars, causing

the pH of the plaque to decrease, if the

decrease in pH occurs continuously, it will

cause demineralization of the tooth root

surface. These bacteria are resistant to

acid, therefore they can survive in a

medium with a high level of acidity. The

bacteria attach to the tooth surface which

can metabolize carbohydrates to produce

organic acids which cause a decrease in

oral pH, causing demineralization of tooth

enamel. Based on the results. Based on the

results (Riskesdas, 2018) the prevalence of

Purnama Fitri, Rasipin, Ari Suwondo | 262

dental caries in Indonesia is still relatively

high at 57.6% and the proportion of

cavities in Indonesia is 45.5% and in

Central Java province is 43.4%.

A healthy oral cavity without plaque

accumulation can be achieved by

mechanical means such as brushing teeth.

Cleaning actions by brushing teeth are

often unable to reach the entire surface of

the teeth, so other efforts are needed such

as utilizing antibacterial materials

(Pujoraharjo & Herdiyati, 2018).

Mouthwash can be used to help clean the

oral cavity from plaque and

microorganisms that can cause damage to

teeth and supporting tissues. The

antibacterial properties of mouthwash are

determined by the content of its active

ingredients. Chlorhexidine is a gold

standard mouthwash from the bisbiguanid

group that can be effective as an anti-

plaque and anti-gingivitis ingredient.

Chlorhexidine has the ability to inhibit S.

mutans bacteria (Fauzia et al., 2021).

urrently, chlorhexidine is routinely used by

doctors in the treatment of patients with

periodontal cases, orthodontia and

maxillofacial surgery. Chlorhexidine is

available as mouthwash, soap, gel, spray,

toothpaste and varnish in different

strengths. Lower strengths have been used

as mouthwashes (0.12%, 0.2%, 0.1%).

Many studies have shown a correlation of

efficacy and the strength of chlorhexidine

used, unfortunately higher concentrations

show an increase in side effects i.e.

staining of teeth and restorations and

decreased taste. Long-term use of

chlorhexidine can cause several side

effects such as causing tooth

discoloration, impaired taste, and is an

allergen if in constant contact14. These

effects will occur if routine use is carried

out continuously for more than 2 weeks

within 2 times a day and in the long term

for more than 2 years or if the use does

not follow the correct rules. This is in

accordance with previous research that

after using chlorhexidine for 3-6 months,

many extrinsic steins were found on the

tooth surface. In cases of periodontitis,

chlorhexidine is one of the antibiotics used

in the treatment of periodontal disease,

but there is a problem of bacterial

resistance when antibiotics are used

continuously. Microorganism resistance to

antibiotics has an impact on high medical

costs, infections are now back as a

problem in the world of medicine. Efforts

to overcome this resistance are carried out

by finding new alternatives in inhibiting

microorganisms.

Oral health is a fundamental part of

general health and affects overall well-

being. Having optimal oral health can help

a person maintain the function of their

oral cavity, and can make a person feel

better and more confident. Oral health is

considered very important because the

oral cavity is a place for bacteria to grow, if

the oral cavity is not cleaned properly,

food debris will stick to the surface of the

teeth and will form plaque. This plaque is a

gathering place for bacteria that cause

dental and oral diseases (Fatmawati, 2015).

Oral health is one of the factors that

support a healthy paradigm and is a

national development strategy to realize

health development for socially and

economically productive human resources.

Therefore, every individual must have the

awareness, willingness, and ability to

263 | Formulation of Lemongrass Extract Mouthwash (Cymbopogon Citratus) as a Non-

Pharmacological Effort in Inhibiting The Growth of Bacteria That Cause Dental Caries

improve their health to the highest level

(Susilawati & Damayanti, 2020).

Maintenance of oral hygiene is one of

the efforts in improving health. The mouth

functions not only for the entrance of food

and drink but more than that and not

many people realize the magnitude of the

role of the mouth for health and well-

being. Therefore, oral health plays a very

important role in supporting one's health

(Ratih & Yudita, 2019). Oral health is

sometimes the umpteenth priority for

some people, even though oral and dental

diseases have a serious impact on general

health, because teeth and mouth are the

entry points for germs and bacteria so that

they are likely to interfere with the health

of other organs.

MATERIALS AND METHODS

The conceptual framework in this study

consists of independent variables, namely

lemongrass extract mouthwash

formulations, dependent variables, namely

the growth of bacteria that cause dental

caries.

The type of research used is laboratory

and field experiments, pure experiment

with pretest-posttest with control group

design consisting of 2 (two) groups,

namely the intervention group and the

control group. The sampling method used

was purposive sampling.

The population used was junior high

school students. While the research

sample was 35 students

maximum age of 15 years who have

caries teeth. The sampling technique in

this study was non probability sampling,

which is a sampling technique that does

not provide equal opportunities for each

member of the population to be selected

as a sample on the basis of sample

selection considerations according to the

researcher's judgment. The non probability

sampling method used in this study is

purposive sampling.

The tools used are beaker glass,

maceration vessels, porcelain cups,

funnels, glassware, and glassware.

porcelain, funnel, measuring cup, Hot

plate, needle ose, filter paper, Laminar air

flow, microscope spirit lamp, mortar,

stamper, analytical balance (Sarltorius),

glass object, wooden clamp, test tube,

stirrer, glass, petri dish, erlenmeyer,

volume pipette, stir bar, scales, test tubes,

Durham tubes, tube racks, autoclave,

transpipettes, dropper pipettes, inkas,

incubator, oven, parchment paper, cotton

wool, pH meter, vernier, knife, plastic pot,

horn spoon, Vacum Rotary Evaporator,

Brookfield viscometer, waterbath and

mouthwash container.

Research on lemongrass extract

mouthwash formulations, and bacteria

that cause dental caries was conducted at

the Microbiology and Parasitology

Laboratory of the Faculty of Medicine,

UNISSULA Semarang to test the

effectiveness of lemongrass extract

mouthwash formulations against the

growth of bacteria that cause dental

caries, this research was conducted in June

2022.

RESULTS AND DISCUSSION

The study was conducted in June 2022

at SMP Negeri 12 Semarang and the

Laboratory of the Faculty of Medicine

Purnama Fitri, Rasipin, Ari Suwondo | 264

Unissula Semarang which is located at

Jalan Raya Kaligawe Km. 4, Semarang,

Central Java. The materials used were

lemongrass extract (Cymbopogon

citratus), 90% ethanol, glycerin, benzoic

acid, xylitol, oleum menthe, and distilled

water. The research process began with

the extraction of lemongrass plants and

the formulation of mouthwash

preparations carried out at the Laboratory

of the Faculty of Medicine, Unissula

Semarang. The tools used were blender,

laboratory glassware (Pyrex), test tubes

(Pyrex), digital scales, oven, viscometer,

drop pipette, spatel, stirring rod and rotary

evaporator. The lemongrass stems that

have been collected are cleaned of dirt

then washed thoroughly using running

water then chopped and dried using an

oven. The research subjects were obtained

from the saliva of respondents who

experienced dental caries at SMP Negeri

12 Semarang, then the saliva was taken to

the Laboratory of the Faculty of Medicine,

Unissula Semarang for culture and began

the research process of the number of

bacterial colonies and bacterial inhibition

tests. The bacterial culture process is

carried out on nutrient media to obtain

bacteria that cause dental caries. After the

bacteria grow on the media, gram staining

is done to identify gram-positive and

gram-negative bacteria. After gram

staining, the type of bacteria is known,

then proceed with planting bacteria with

agar media to find bacterial species.

The cultured bacteria are then

rejuvenated or purified by inoculating 1

ose of pure culture on Nutrient Agar

media and then incubated at 37°C for 48

hours. In this research sample, the bacteria

that cause dental caries found are

Streptococcus mutans bacteria.

The concentrations of lemongrass

extract mouthwash formulations used in

this research are 33%, 36%, and 39%

concentrations.

Bacterial testing was carried out on

each treatment group using lemongrass

extract mouthwash formulations at

concentrations of 33%, 36%, 39%, and

chlorhexidine mouthwash control.

The incubated bacteria were diluted by

mixing 1 ose of bacteria into a test tube

containing 1 ml of NaCl, then

homogenized using a vortex and the

turbidity was standardized with McFarland

concentration so that the number of

bacteria was eligible for the test. The

growth inhibition test of Streptococcus

mutans bacteria was carried out by

dilution, namely by putting 1 ml of

bacteria and 20 ml of Nutrient Agar in 5

enlenmeyer tubes with a volume of 50 ml

containing lemongrass extract mouthwash

formulations with each concentration of

33%, 36%, and 39% and chlorhexidine

control. All tubes were incubated at 49°C

to observe bacterial inhibition by looking

at the turbidity of the tubes compared to

the chlorhexidine control. Then the tubes

were poured into Petri dishes and planted

on Nutrient Agar, then incubated at 37°C

for 48 hours. The minimum zone of

inhibition formed was measured by

counting bacterial colonies using markers

and lights. Markers are used as stationery

to write markers on the number of

bacteria on petri dishes, lamps are used as

lighting aids when counting bacterial

colonies assisted by a microscope.

1. Data Normality Test

265 | Formulation of Lemongrass Extract Mouthwash (Cymbopogon Citratus) as a Non-

Pharmacological Effort in Inhibiting The Growth of Bacteria That Cause Dental Caries

Normality tests are carried out using

the Shapiro-Wilk statistical test which

shows the distribution of normally

distributed or abnormally distributed data.

Table 1. Table of Normality of Data on the

Number of Bacterial Colonies

Bacteria

Group

Shapiro-

Wilk

Sig.

Streptococcus

mutans

33% concentration

solution

0.337

36% concentration

solution

0.301

39% concentration

solution

0.118

Chlorhexidine

0.082

Based on Table 1, it shows that with the

Shapiro-Wilk normality test, a p> 0.05 value

is obtained so that the ability to reduce the

number of bacterial colonies is normally

distributed.

2. Parametric Test

The ability of the treatment group and

the control group to Streptococcus mutans

bacteria had a normal data distribution so

that this study used a paired parametric

test with the Paired T-Test and an unpaired

test with the One Way Annova test.

a. The Paired T-Test was used to

determine whether there was a

difference in the average killing power

of Streptococcus mutans in each

group.

Table 2.Ability of Each Group in Reducing the Number of Bacterial Colonies

Group

Mean±SD Pretest

Mean±SD

Posttest

Delta±SD

P-

values

Concentration

33%

248.17±185.67

106.33±53.836

141.83±229.40

0.190

Concentration

36%

297.60±176.10

80.80±97.374

216.80±122.34

0.017

Concentration

39%

302.33±147.65

10.50±13.293

291.83±155.33

0.006

Chlorhexidine

364.00±338.31

288.00±257.36

136.00±159.15

0.091

Based on table 2, the lemongrass

extract mouthwash formulation with a

concentration of 36% and a concentration

of 39% has a p value <0.05 indicating that

these concentrations are more capable of

reducing bacteria compared to the

lemongrass extract mouthwash

formulation with a concentration of 33%,

and chlorhexidine which has a p value >

0.05. The highest ability to reduce the

number of bacterial colonies was shown in

the formulation of lemongrass extract

mouthwash with a concentration of 39%.

Based on the table, a graph was created

that illustrated the ability of each

treatment group to reduce the number of

bacterial colonies.

b. One Way Annova test

One Way Annova test was conducted

forlook at the differences between each

group.

Purnama Fitri, Rasipin, Ari Suwondo |266

DOI : 10.36418/jrssem.v2i2.296

Table 3.Intergroup Difference Test in Reducing the Number of Bacterial Colonies

Pretest ± SD

Group

Mean±SD

P-values