Characterization Evaluation of Clove Flower Oil (Syzygium aromaticum (L.) Meer) And Pharmacological Properties Of Anthelmintic

Abstract


Background
Clove oil is included in the family Myrtaceae genus Syzygium, the essential oil produced from clove plants (Syzigium aromaticum) is found in Indonesia, India, and Madagascar (Alma et al., 2007). Clove oil can be isolated from leaves (1-4%), stems (5-10%), and clove flowers (10-20%). Essential oils from clove flowers have the best quality and are expensive because they contain eugenol reaching 80-90%, clove flower oil has biological activities which are as anti-microbial, anti-pain, antioxidant, anti-cancer, anti-inflammatory, antiinflammatory, anti-depressant, and antiulcer (Kamataou et al., 2012). Clove oil is highly valued in the medical field as carminative and stimulant and is said to be an anthelmintic. In recent years, it has been recognized as an anesthetic that is effective in calming fish in a number of fisheries management (Mittal et al., 2014). The focus of this review is to provide information about the components of Syzygium aromaticum, and pharmacological activities against anthelmintics.

Materials and Methods Determination of plants
Determination of clove plants is carried out at Herbarium Bogoriense, Botany Field, Bogor Biology Research Center-LIPI. This determination aims to ascertain the truth of the plant species to be used.

Clove flower oil distillation
Clove flowers are distilled using the steam distillation method. The sample used was 4 kg distilled for 8 hours connected to the condenser. During heating the distillation of the oil will evaporate with water vapor through the condenser, then a condensation process occurs that produces essential oils in clove flowers. Destilates are collected and separated from the water contained in them using anhydrous sodium sulfate in a separating funnel (Prianto et al., 2013).

Characterization of the physical properties of clove flower oil
Characterization of physical properties is done by looking at the color, smell, refractive index, and specific gravity measurements. These parameters have standardized values as a condition of trade quality in Indonesia listed in SNI 06-4267-199606-4267- (Prianto et al., 2013. Characterization of clove flower oil components using the Gas chromatography-mass spectrometry (GC-

MS) method
The characterization of clove flower oil components was carried out by injecting 0.05 µL of clove flower oil into the syringe of GC-MS with a capillary column length of 30 m. The stationary phase used is 5% diphenyl or 95% dimethylpolisiloksan while the mobile phase used is helium gas with a flow rate of 53.0 mL / minute at a pressure of 12.0 kPa. Column temperature is 50.0 • C and 300,00 • C injector temperature with split ratio of 93.0. Identification of the components of clove flower oil based on the comparison of standard eugenol characterization (Prianto et al., 2013).

Results and Discussion Determination of plants
Clove plants obtained from Lambaro Seubun Village, Lhoknga Subdistrict, Aceh Besar Regency were conducted at Herbarium Bogoriense, Botany Field, Bogor Biology Research Center-LIPI. The results of the determination showed that the plants used in the study were Syzygium aromaticum (L.) Merr. L.M. & Perry from the Myrtaceae tribe.

Clove flower oil distillation
Clove flower oil distillation of 4 kg was carried out using a steam distillation method for 4 days with a time of 8 hours per day. The distillation yields 163mL of clove flower oil. Purification of clove flower oil was carried out using Sodium Sulphate Anhydrous (Na2SO4). Anhydrous sodium sulfate aims to separate and bind water bound to clove flower oil (Prianto et al., 2013).

Characterization of the physical properties of clove flower oil
Parameters characterized have values that have been standardized on trade quality requirements in Indonesia listed in SNI 06-4267-1996.
The results of the characterization in Table 2. have met the quality requirements of SNI 06-4267-1996, which means meeting the quality standards of essential oil trading in Indonesia. The refractive index of clove oil produced is 1.52 with a specific gravity of 1.05, light yellow and has a distinctive smell of clove oil. Characterization of clove flower oil components using the Gas chromatography-mass spectrometry (GC-
The different types of compounds identified and the percentage of components in clove flower oil are caused by several factors including the differences in the varieties of clove flowers used, preparation of samples before distillation and distillation methods. Geographical location will affect the components of inorganic soil where clove plants grow, so that it will affect the secondary metabolites of clove flower oil produced (Srivastava et al., 2005).

Pharmacological
Properties of Anthelmintics Bhowmik et al., (2012) said that clove essential oil contains around 85-92% eugenol which is very active against microorganisms (classified as bactericidal in nature). Cloves are also said to be anthelmintic, antiviral, antimicrobial, antiseptic, and anti-fungal. Dhanraj et al., (2015) in his research entitled In Vitro study of the effect of Syzygium aromaticum ethanol extract on carbohydrate metabolism Cotylophoron cotylophorum validates the anthelmintic properties of Syzygium aromaticum ethanol extract against C. cotylophorum characterized by inhibition of carbohydrate metabolic enzymes, namely pyruvate kinase, phosphoenolpyruvate carboxykinase, lactatdehydrogenase, malatdehydrogenase, fumaratreductase and succinate dehydrogenase which can reduce ATP production may be fatal for parasites. This study uses five concentrations, namely. Evaluation of the anthelmintic activity of Eugenia caryophyllus alcohol extract with concentrations (5; 10; 15 mg/mL) & Uncaria gambier with concentrations (50; 100; 150mg/mL) on Pheretima posthuma worms in Sonalkar and Sachin (2014) obtained a concentration of 15mg/mL clove extract showed a paralysis response in 1.5 minutes and lysis occurred in the 9th minute. Uncaria gambier at a concentration of 150mg/mL paralysis occurred in the 2.5 minute and lysis occurred in the 62 minutes. Eugenia caryophyllus and Uncaria gambier alcohol extract showed significant anthelmintic activity the greater the concentration the faster the time of death of worms. The comparative evaluation of these two plants shows that Eugenia caryophyllus has a strong anthelmintic activity compared to Uncaria gambier. Patil et al., (2013) used ethanol extract of Syzygium aromaticum with the aim of investigating the potential of anthelmintic on Pheritma posthuma with concentrations (2.5; 5 and 10 mg / mL). Positive control as standard used albendazole 20 mg / mL and distilled water as a negative control. The results obtained by the S. aromaticum ethanol extract concentration of 2.5; 5 and 10 mg/mL show paralysis at 4.267±0.252, 2.433±0.306 and 1.033±0.551 minutes while lysis occurs at 45±2.00, 35±4.35 and 2.333±0.351 minutes. Albendazole (20 mg/ml) was shown to have paralysis at 1185.33±79.02 minutes while lysis at 1608.33±78.57 minutes. This study shows that the ethanol extract of S. aromaticum showed significant anthelmintic activity at higher concentrations as evidenced by bleeding and necrotic patches observed externally on the worm's body. From this it can be concluded that the ethanol extract of S. aromaticum has anthelmintic activity which is useful for fighting worm infections. Kumar and Singh (2014) in his study said that Allium sativum, Ferula asafoetida and Syzygium aromaticum can be used as anthelmintic. Anthelmintic activity was tested in vitro against the liver worms Fasciola gigantica. The results of his research state that ethanol extract is more toxic than other organic extracts. The ethanol extract of S. aromaticum (2h LC50 2.95 mg/mL) was more toxic to F. gigantica than A. sativum (2h LC50 3.48 mg/mL) and F. asafoetida (2h LC50 3.94 mg/mL). The in vitro anthelmintic activity of Pheritima posthuma worms from S. aromaticum (flower buds) and C. zeylanicum (bark) water extracts examined by Kumari et al., (2015) stated that activity depends on the concentration observed in the extract. A concentration of 20 mg/mL of C. zeylanicum showed significant activity compared to S. aromaticum. This study also carried out phytochemical screening of extracts of S. aromaticum and C. zeylanicum revealing the presence of alkaloids, tannins, phenolics, phlobatannins, cardiac glycosides, anthraquinone glycosides, hydrocinone hydrakuinone, flavonoids and terpenoids.

Conclusion
The distillation results obtained 163 mL of clove oil with physical properties that meet the requirements of SNI 06-4267-1996. The results of the analysis using GC-MS showed that clove oil contained 6 components, namely, Eugenol 88.20%, Alpha Copaene 0.49%, Caryophyllene 3.77%, Alpha Humulene 0.48%, Eugenyl acetate 6.76%, and Caryophyllene Oxide 0.30%. Another fact shows that clove flower oil has anthelmintic activity.