ANTIBACTERIAL ACTIVITY OF TEMULAWAK ESSENTIAL OIL EXTRACT

(Curcuma xanthorrhiza Roxb.) AGAINST Porphyromonas gingivalis GROWTH

Periodontal disease is a dental and oral health problem that has a quite high prevalence
in the community with the prevalence of periodontal disease in all age groups in Indonesia is
96.58% ( Alibasyah , 2016 ). Periodontal disease starts from gingivitis which if not treated
further can develop into periodontitis . Periodontitis is an inflammation of the supporting
tissues of the teeth caused by pathogenic bacteria on plaques (Newman et al ., 2015).
Plaque bacteria on early colonization was dominated by Gram-positive bacteria such as
genus Streptococccus, and Actinomyces. Plaque bacteria in secondary colonization occur after
initial colonization, secondary colonization bacteria are dominated by Gram negative bacteria
such as Prevotela intermedia, Fusobacterium nucleatum, and Porphyromonas
gingivalis (Newman et al ., 2015). P. gingivalis is most commonly found in chronic
periodontitis with a prevalence of 53.8% and aggressive periodontitis with a prevalence of
79.6% (Newman et al., 2015).
Periodontitis treatment includes mechanical treatments such as comprehensive plaque
control with scaling root planing , curettage, so as to eliminate inflammation and reduce
pocket depth (Orgendrik, 2012). Additional therapy with antibiotics is also needed to support
mechanical treatment, because of additional therapy of antibiotics are more effective and
accelerate healing than just mechanical treatment (Krismariono, 2009 ; Wijayanto et al .,
2014 ).
Administration of antibiotics in the treatment of periodontal disease can be done locally
or systemically (Setiawan et al., 2013) . One of the commonly used local antibiotics is
the metronidazole gel which is proven effective against negative Gram Anaerobic
bacteria which causes of periodontitis (Wijayanto et al ., 2014). Patients who have
hypersensitivity or are allergic to the metronidazole gel are contraindicated to use this
ingredient (Arunachalam et al ., 2017). Overcoming this, there needs to be a variety of drug
choices with the development of new antibiotics from herbal ingredients.
One of the plants that can be used as medicinal plants is Curcuma
xanthorrhiza Roxb. Based on research by Jeeva et all (2012) the results of analysis of curcuma
essential oil with Gas Chromatography-Mass Spectroscopy (GCMS), the largest active
compound content in curcuma essential oil extract namely xanthorrihizol,
camphene and curcumin which are known to have antibacterial activity.
 The aim of this study was to determine the antibacterial power of curcuma rhizome
( Curcuma xanthorrhiza Roxb.) Essential oil against P. gingivalis .
 RESEARCH METHODOLOGY
Essential Oil Extract Curcuma Rhizome . The curcuma rhizome essential oil extract
was obtained using the Water and Steam destilation method . Curcuma rhizome is harvested at
the age of 7-8 months, obtained from a farmer in Dusun Hamlet RT 19 / RW 07 in Tanggung
Village , Padang Subdistrict , Lumajang Regency. Distillation is carried out for about 5 hours
with a water temperature of 100°C. Furthermore, essential oils and water are separated by a
separating funnel. Oil obtained was placed in a dark bottle sealed and stored at a cool place of
18°C.
Isolate P. gingivalis . Isolate P. g ingivalis used was P. gingivalis ATCC
33277 in providing bacterial cultures which use BHIB ( Brain Heart Infusion) . Suspension P.
gingivalis was made by means of 0.3 ml of bacteria was put into a test tube containing 4ml of
saline solution , then the turbidity is in accordance with the standard McFarland 0.5 (1.5 x
10 8 CFU mL ) .
Antimicrobial Test P. gingivalis . The method used in this study is the agar disc
diffusion test with 4 times repetition. This method is done on Agar Muller-Hinton (MH-
A). Essential oil curcuma rhizome with concentration of 25; 50; 75; 100% taken as much as
20 μl, then dropped on paper discs with a diameter of 5 mm. Then the paper
discs are placed scattered over the surface of the MH-A media that has been inoculated
with P. gingivalis. Petridis were put into desiccator and incubated at 37 ° C for 24 hours. After
24 hours, the measurement of inhibitory zone diameter around the paper discs using a digital
caliper can be done.
RESULTS
The results of the research are shown in the form of inhibitory zone which was
formed around the paper discs in all treatment groups of Curcuma rhizome essential oil
extract ( C. xanthorrhiza Ro xb.). It was shown that there was an inhibitory zone which looked
clear around the paper disc , as seen in Figure 1. The average of inhibitory zone diameter of
curcuma rhizome essential oil extract ( C. xanthorrhiza Roxb.) can be seen in Table 1.
Picture 1 . Inhibitory zone of curcuma rhizome essential oil against P.
gingivalis (orange arrow) .
Table 1 . The average value and standard deviation of the diameter of the inhibitory
zone (mm) of curcuma rhizome ( C. xanthorrhiza Roxb.) Essential oil against P. gingivalis
Information:
N = Number of repetitions
 K- = negative control
K + = positive control
Cx25 = Essential oil of curcuma rhizome concentration of 25%
Cx50 = Essential oil of curcuma rhizome concentration of 50%
Cx75 = Essential oil of curcuma rhizome concentration of 75%
Cx100 = Essential oil of curcuma rhizome 100% concentration.
Data on inhibitory zone diameter were analyzed statistically using the Kruskal Wallis
test showed the data has a significance value of 0.001 (p < 0.05) . This shows that there are
differences in the study group and followed by the Mann Whitney test aims to find out which
groups have significant differences between groups . The Mann Whitney test results can be
seen in Table 2.
Table 2. Mann Whitney Test Results
Description: * = significant value
DISCUSSION
The antibacterial power of Cx essential oil extract is directly related to the active
compounds that make it up. Cx essential oil containing 92% is largely composed
of terpene group compounds , specifically sesquiterpenes , monoterpenes and other
oxygenated derivatives , which generally have a low toxicity level and act as antibacterial
( Male et al ., 2012 ). The terpenes and other oxygenated derivatives compounds are
hydrophobic or lipophilic, which can enter into the membrane structure specifically. This
causes expansion of the membrane so that there is an increase in fluidity which ultimately
causes bacterial cell lysis (Trombetta et al ., 2005; Nazzaro et al ., 2013; Zengin and Baysal,
2014) .
The most active compound in Cx essential oil is Xanthorrizol group of
sesquiterpen as much as 64.38%, followed by other terpenes such as camphene group
of sesquiterpen (8.27%), and curcumin group of monoterpen (5.85%) . The rest in small
amounts, composed of α-Pinene , β-curcumene , zingiberene, α-thujene, β-pinene, Myrcene,
Linalool, β-bisabolol, and ar-curcumene (Rukayadi and Hwang , 2006
The antibacterial power of Cx essential oil can be classified based on the diameter of
the inhibitory zone. Ponce et al . (2003) classified antibacterial power into 4 levels, namely
weak, moderate, strong, and very strong. Antibacterial power is said to be weak if the diameter
of the inhibitory zone < 8 mm, medium 8-14 mm, strong 15-19 mm, and very strong if> 20
mm. The diameter of inhibitory zone of Cx essential oil to the growth of P. gingivalis which
has an average value of inhibitory zone diameter of 8.1mm-10.1 mm . According to the
classification of Ponce et al . (2003), all four concentrations have antibacterial power with
moderate levels on P. gingivalis .
The results of this study are in line with the research conducted by Syafira (2018) which
states that Cx essential oil extract with a concentration of 25%, 50%, 75%, and 100% has
antibacterial power against Enterococcis faecalis bacteria (gram-positive), and Fusobacterium
nucleatum (gram-negative). Essential oils with a concentration of 100% have antibacterial
power greater than the concentrations of 75%, 50%, and 25% in bacteria. This is also in line
with the research conducted by Prabuseenivasan et al . (2006) and Male et al . (2012) which
stated that curcuma rhizome essential oil will work better at higher concentrations.
This study used 12.5% metronidazole gel as a positive control. The diameter of the
metronidazole gel inhibitory zone for the growth of P. gingivalis is in the range 30-31
mm. When referring to the classification of Ponce et al . (2003), the antibacterial power of
12.5% metronidazole gel to the growth of P. gingivalis was very strong.
Further research is needed regarding the antibacterial power of a single active
compound from Cx essential oil to the growth of P. gingivalis and optimization using different
essential oil extraction methods or increasing the combination potentiation with other herbal
essential oils so as to have the same quality with the existing products. By doing so, this
research can be used as a reference for further research, especially in developing periodontal
antibiotic drugs.
THANK YOU
We thank Drg . Melok Aris W., M.Kes. Sp. Perio and drg. Depi
Praharani, M.Kes who have provided many inputs and suggestions in this study.