INTRODUCTION:

Coral “Betadine” plant (Jatropha multifida Linn.) is a rare plant but is widely used by the people of South Sumatra, Indonesia as an ornamental plant. Apart from being an ornamental plant, betadine plants, especially the sap on betadine leaf stalks, are widely used as wound medicine that can heal wounds and pain due to accident or cut. Jatropha multifida is a member of the Euphorbiaceae family. The genus name Jatropha is originated from the Greek “iatros” (doctor) and “trophe” (nutrition), as it is used in pharmaceuticals¹. Jatropha is a rich source of secondary metabolites with intriguing biological activities (such as pathogen inhibition). In general, secondary metabolites are a significant source with a wide range of structural arrangements and properties². Because of its properties as a wound medicine, this plant is known by the community as a betadine plant (wound medicine).

Previous research showed that the phytochemical test results of betadin leaf ethanol extract contained phenolic compounds, tannins and flavonoids. Liana's study also showed the antibacterial activity of the active fraction of betadine leaves against Staphylococcus aureus bacteria³. This can be caused by the content of active substances flavonoid and phenol. These results are supported by research by Nagaharika, et.al⁴ which showed betadine leaves can be anti-inflammatory, while Sudaryono's research⁵ showed that betadine plant stems were able to increase the blood platelet count of Mus musculus animals. Flavonoids were anti-inflammatory so they can reduce inflammation and help reduce pain, if there was bleeding or swelling in the wound. Phenol compounds have the ability to form complex compound with proteins through hydrogen bonds, so that they can damage bacterial cell membranes⁶. Based on this, the research that had been done were still limited to anti-imflamation and testing of rat blood platelets or the antibacterial activity of Staphylococcus aureus as an antimicrobial using GC-MS (Gas Chromatography–Mass Spectroscopy). Numerous reports have determined the pharmaceutical effectiveness of various Jatropha species^2,7,8; however, information on the Betadine leaves is limited.

The purpose of this experiment was to determine the composition of chemical compounds contained in the methanol extract of betadine (Jatropha multifida Linn) leaves by GC-MS. The results of this analysis can be used as a reference in determining the antimicrobial activity of betadine leaves.

MATERIALS AND METHODS:

Plant material:

The materials used in this study were betadine leaves (Jatropha multifida Linn) from South Sumatra, Indonesia.

Sample Extraction:

Betadine leaves and petioles were separated and washed, then drained and dried in direct sunlight for 4 hours. A total of 50grams of betadine leaf simplicia was extracted with a solvent ratio of 1:10 (w/v). The solvent used was methanol. Extraction was carried out 2 x 24 hours by maceration. The maceration results were filtered, then evaporated with a rotary evaporator at a temperature of 50^oC⁹.

Phytochemical Test:

Phytochemical tests were carried out on each extract using standard procedures that have been reported in previous studies. These tests include the tests for alkaloids, triterpenoids, steroids, tannins and saponins

Volatile Compound Analysis with GC-MS:

20 mg of betadine leaf part methanol extract was put into a sample bottle, then 4ml of solvent was added in the ratio, methanol (1.7 ml): H₂SO₄ 98% (0.3ml): Chloroform (2ml). The bottle was tightly closed, homogenized using a vortex for 15 minutes, then heated using a heating block at 90ºC for 90 minutes. After heating, the sample bottles were cooled to room temperature for ±5-6 hours. After cooling, 1ml of distilled water was added and then homogenized using a vortex for ± 1 minute. The sample was allowed to stand for a while until 2 layers were formed. Separate the bottom layer into another sample bottle. Then the solution was added Na₂SO₄ until it was insoluble. And filtered using filter paper. Furthermore, the filtrate was transferred into a vial and the sample was ready for GC-MS analysis¹⁰. The condition of the GC-MS tool can be seen in Table 1.

Table 1. GC-MS Analysis Condition

Column	Capillary column type Agilent 19091S-433 HP-5MS
Stationary Phase	Phenyl Methyl Silox
Column Temperature	325 °C
Column Length	29,81 m Diameter 325 °C
Detector	MS (Mass Spectrometer)
Syringe size	10 µL
Injection Volume	1 µL Injection temperature : 250°C
Flow Rate	15 mL/menit,

RESULTS AND DISCUSSION:

Phytochemical Test:

The results of the phytochemical screening test showed that the methanol extract from the leaves of Betadin (Jatropha multifida Linn) contained several active compounds, namely alkaloids, flavonoids, tannins, total phenols and terpenoids. The complete phytochemical test results can be seen in Table 2.

Table 2. Results of Phytochemical Screening of Betadine Leaf Methanol Extract

The Component	Qualitative Result
Alkaloids	++
Flavonoids	+
Phenol	+
Tannins	+
Saponins	-
Terpenoids	-
Unsaturated Steroids	-
Steroid Glycosides	-

Based on Table 2, the methanol extract of betadine leaves were suspected to have a fairly high alkaloid compound, in addition to the presence of flavonoids and phenols and tannins. It was also known that betadine leaves do not contain saponins, steroids and glycosides. These results are quite different from the content contained in different types of Jatropha. As in Jatropha gossypiifolia which has additional content of saponins, steroids and terpenoids^11,12. Phytochemicals are biocompounds contained in plant food, such as fruit, vegetables, grains and other plant foods, that are non-nutrient chemical agents. They can be classified into several groups such as polyphenols, sulfur compounds of organism, carotenoids, alkaloids and chemicals containing nitrogen. Phenolic acids, stilbenes, coumarins and tannins are among the most widely-studied chemicals and can be further broken down into flavonoids (including flavonols, flavones, catechins, flavanones, anthocyanidins and isoflavons)¹³.

Flavonoids are known to be generated by plants to be hydroxylated phenolic compounds and are reported to be antibacterial against a wide variety of in vitro microbes. Recent study has found that flavonoids are strong antioxidants with reported antimutagenic and anticarcinogenic effects In the pharmacology of dietary phytochemicals¹⁴. Alkaloid compounds were reported as chemical compounds that can prevent disease and several derivatives of alkaloids show biological activities such as anti-inflammatory¹⁵, antimalarial¹⁶, antimicrobial¹⁷. In addition, steroid compounds were known to have cardiotonic, antibacterial and insecticidal effects¹⁸and tannins as antibacterial¹⁹. The major part of antioxidants in plants are phenolic chemicals. Several studies have demonstrated that the development of cardiovascular disease, cancer and osteoporosis is restricted by phenolic substances²⁰.

GC-MS Analysis:

Analysis of Gas Chromatography-Mass Spectrophotometry (GC-MS) was carried out to identify volatile compounds in betadine leaf methanol extract. The results of the analysis showed that the compounds detected were: 4H-Pyran-4-one, 2,3-dihydro-3,5-dihydroxy-6-methyl-; 2,4-Di-tert-butylphenol; Dodecanoic acid, ethyl ester; Tetradecanoic acid; 8-Methylnonanoic acid; 9-Hexadecenoic acid, 7-Hexadecenoic acid, methyl ester, Methyl hexadec-9-enoate; cis-9-Hexadecenoic acid methyl ester; Pentadecanoic acid, 14-methyl-; Palmitoleic acid; Ethyl 9-hexadecenoate, E-11-Hexadecenoic acid; 9,12-Octadecadienoic acid (Z,Z)-, Methyl 10-trans,12-cis-octadecadienoate; Phytol; Methyl stearate, Heptadecanoic acid, 16-methyl-; Linoelaidic acid; 9,12,15-Octadecatrienoic acid, (Z,Z,Z)-, 7,10,13-Hexadecatrienoic acid; .alpha.-Tocospiro A, .alpha.-Tocospiro B; .gamma.-Sitosterol, .beta.-Sitosterol; Lup-20(29)-en-3-one, D:C-Friedoolean-8-en-3-one; and Lupeol. The chromatogram can be seen in Figure 1.

Based on the results of the GC-MS test, it showed that there was an abundance of triterpenoids, namely phytol compounds and types of fatty acids that were thought to have antibacterial and fragrance activity, such as 9-Hexadecenoic acid, methyl ester, (Z)-; 7-Hexadecenoic acid, methyl ester, (Z)-; Methyl hexadec-9-enoate; 9,12,15-Octadecatrienoic acid, methyl ester, (Z,Z,Z)-; and Phytol. This was in accordance with the statement of Bhattacharya¹⁶, that phytol was a class of diterpenoid and triterpenoid compounds that were commonly found in seaweed and have microbial activity. In a plant defense mechanism, triterpenoids play a very significant role. It protects plants against insects and environmental stress, whether constitutive or induced²¹.

CONCLUSION:

Based on the results of research on the methanol extract of the betadine leaf, it was suspected that it had a fairly high alkaloid compound, a little flavonoid, a little phenol. The type of chemical compound extracted into methanol solvent showed that the compounds contained in the betadine leaf were polar. The results of GC-MS on betadine leaf extract samples detected 27 peaks of compounds, the 3 most dominant compounds were at retention times of 18,072, 19,950 and 20,058, namely 9-Hexadecenoic acid, methyl ester, (Z)-; 7-Hexadecenoic acid, methyl ester, (Z)-; Methyl hexadec-9-enoate; 9,12,15-Octadecatrienoic acid, methyl ester, (Z,Z,Z)-; and Phytol.

ACKNOWLEDGEMENT:

Authors are thankful to Politeknik AKA Bogor for providing financial support for laboratory and instrumental facilities.

CONFLICT OF INTEREST:

The authors declare that they have no conflict of interest.

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Received on 19.08.2021 Modified on 21.11.2021

Research J. Pharm. and Tech 2022; 15(10):4633-4636.

DOI: 10.52711/0974-360X.2022.00777

Peak	Retention Time	Chemical Compounds	Molecular Weight	Relative Content (%)	Molecular Formula
A	18.072	9-Hexadecenoic acid, methyl ester, (Z)-Methyl hexadec-9-enoate 7-Hexadecenoic acid, methyl ester, (Z)-	268	17,22	C₁₇H₃₂
B	20.058	Phytol	296	13,96	C₂₀H₄₀
C	19.950	9,12,15-Octadecatrienoic acid, methyl ester, (z,z,z)-(Metil Linolenat)	292	12,31	C₁₉H₃₂