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A Qualitative test of Primary and Secondary Metabolites of Bintaro Plant as a Rat (Rattus argentiventer) Pest Repellent

Bintaro is a mangrove plant that grows on the coast and is often used as a shade tree in megacities. The toxic content of the Bintaro plant is found in all parts of the plant. The toxic content of cardiac glycoside compounds contained in the Bintaro plant can be used as a rat repellent. Rat pests are important pests in crops, especially in rice plants which are difficult to control in mechanically and chemically, thus rice production always decreases. Therefore, it is necessary to search for effective, efficient, and environmentalfriendly control technology, namely by using fruit extracts. The plant-based rodenticides made from Bintaro fruit extracts are effective for overcoming rat pests. The purpose of the study was to determine the qualitative levels of primary and secondary metabolites from Bintaro plants which act as antifeedants for rat pests (Ratus argiventer). The method used in this research was qualitative testing using phenol method to test the content of primary and secondary metabolites in the leaves and stems of Bintaro plants. From the results of the research conducted, it was found that the qualitative levels of primary metabolites from Bintaro leaves and stems contained protein content. Fat and carbohydrate content of secondary metabolites found in the stems and leaves of the Bintaro plants were Alkaloids, Saponins, Flavonoids and Polyphenols in Bintaro leaves and on its stems contained Flavonoids, Saponins, Tannins and alkaloids. Keywords— Bintaro, rat pests, primary and secondary metabolites content.


INTRODUCTION
Bintaro is a mangrove plant that grows a lot on the coast and is often used as a shade tree in megacities. This Bintaro plant is known for its high toxic content, where the poison from the Bintaro plant has been used for various uses since the early 15th century. The toxic content in the Bintaro plant is found in all parts of the plant, ,especially its fruit, which has the highest toxic content.
In Asian society, especially Indonesia, Bintaro fruit is widely used as a rat pest repellent. So far, the handling of rat pests has been carried out using commercially-available rodenticides. Considering of many dangers posed by rodents which have anticoagulant-based ingredients, alternative ways of controlling rat pests were developed. One of the alternative ways to control rat pests is to utilize the content of one of the characteristics of Bintaro fruit. The toxic content of cardiac glycoside compounds contained in Bintaro fruit seeds can be used as a rat repellent.
In general, there are two kinds of metabolism, namely primary metabolism and secondary metabolism. Primary metabolism produces primary metabolites, while secondary metabolism produces secondary metabolites. Primary metabolism is present in all organisms with almost the same processes and pathways, whereas secondary metabolism has specific and unique pathways and products for each organism. Primary metabolism is directly involved in growth, whereas secondary metabolism is not involved (Anurag et al., 2015).
Primary metabolites modify and synthesize carbohydrates, fats, proteins and nucleic acids, while In plants, secondary metabolite compounds have several functions, including as attractants (attracting other organisms), defense against pathogens, protection and adaptation to environmental stress, protection against ultraviolet rays, as growth regulators and to compete with other plants (allelopathy). Secondary metabolites are also suspected as waste or plant detoxification products, however, most of the function of secondary metabolites is still unknown This research was conducted to determine the qualitative levels of primary and secondary metabolites from Bintaro which can function as a biorodenticide.

II. RESEARCH METHODOLOGY
The research was conducted in the laboratory of the Faculty of Agriculture, Muhammadiyah University of North Sumatra, Medan.
The materials used in this study were Bintaro plants (leaves and stems), alcohol, methanol, aquades, and others that support the research.
The tools used in this study were beaker glass, test tube, spatula, Erlenmeyer flask, stopwatch, calculator, writing instruments and others that support this research.

Sample Preparation
The samples used in this study were the leaves and stems of Bintaro. A total of 100 grams were cut into small pieces.
Then dried using an oven at 60 o C for 6 hours.

Preparation of standard glucose solution
A total of 0.1 g of glucose powder was weighed and then put into a 100 mL flask. A total of 10 mL of glucose stock solution was taken using a pipette then put into a 100 mL measuring flask and diluted to the limit mark. The glucose standard solution was made with the concentrations of 0, 10, 20, 30, 40, 50 ppm by piping out the glucose standard solution as much as 0, 5, 10, 15, 20 mL then put into a 50 mL measuring flask and diluted to the marking area. The next step was measuring the absorbance with a spectrophotometer at a wavelength of 490 nm, then making its linear equation as standard curve (Bintang, 2010).

Determination of carbohydrates (Phenol Method)
10 grams of Bintaro plant samples (stems and leaves) were weighed, then furnaced for 5 hours. 1 gram of sample ash was taken and dissolved in 10 mL concentrated HNO3, then filtered in a 10 mL measuring flask. Then, the filtrate diluted with distilled water to mark the boundaries. Furthermore, 1 mL was taken and then added 1 mL of phenol 1% and 6 mL of sulfuric acid and 2 mL of distilled water. The mixture was allowed to stand at room temperature and then its absorption was measured at a wavelength of 490 nm. The treatment was repeated twice (duplo)

Determination of Protein (Kjeldhal Method)
A total of 1 gram sample was weighed, then put into a kjeldahl flask, added 1 tablet kjeldhal. Then 10 mL of concentrated sulfuric acid solution was added and all the ingredients were digested (heated) in the Kjeldahl flask until it boiled and dissolved and the liquid turned clear. 75 mL of distilled water was diluted and cooled to room temperature. 10 mL of the filtrate was taken to determine the total nitrogen content and determined using a spectrodirect at a wavelength of 410 nm. The treatment was repeated twice (duplo).

Qualitative test of secondary metabolites
The leaves and stems of fresh Bintaro are cleaned and then cut into small pieces. Furthermore, the cut-offs were dried for 7 days (1 week). After drying, then mashed the sample using a blender until smooth. After that, the leaves, stems and smooth Bintaro fruit were ready to be extracted. The manufacture of Bintaro plant extract began by weighing 10 grams of Bintaro powder (leaves and stems). After that, the sample was put into an Erlenmeyer flask and added with 100 mL of aquades. Then closed the Erlenmeyer flask using aluminum foil and soaked for 3 x 24 hours (48 hours) and shook it using an orbital shaker. After 72 hours the extract was filtered using a filter and the filtrate obtained was used in testing for secondary metabolites. The steps were the same for the extraction treatment with ethanol solvents.

Test of Secondary Metabolite Compounds from Bintaro Plant Extracts Extracted with Water and Ethanol Solvents Alkaloid Test
The test was carried out by taking 2 mL of each sample (leaves and stems) of Bintaro which had been extracted with

Saponin Test
The test was carried out by taking 2 mL of each sample (leaves and stems) of Bintaro which had been extracted with water and ethanol solvents. The sample was put into a test tube, added 10 ml of hot water, cooled and then shook vigorously for 10 minutes). The reaction is positive if foam is formed which is steady for not less than 10 minutes, 1 cm to 10 cm high. With the addition of 1 drop of 2 N hydrochloric acid, the foam does not disappear (Arif et al, 2015).

Polyphenol Test
The test was carried out by taking 2 mL of each sample (leaves and stems) of Bintaro which had been extracted with water and ethanol solvents. Then it was reacted with 1% FeCl3 solution, if it forms green, red, purple, dark blue, blackish blue or greenish black, it indicates the presence of phenolic compounds (Kurratul et al., 2014).

Tannin Test
The test was carried out by taking 2 mL of each sample (leaves and stems) of Bintaro which had been extracted with water and ethanol solvents, then heated for about 5 minutes. After heating, each of them added a few drops of 1% FeCl3. If each solution forms a greenish brown or blue-black color, it is positive for tannins (Marlinda et al, 2012). Alkaloids contain in the stems and leaves of the Bintaro plant are toxic, as food inhibitors and insecticides for insects. According to Cahyadi (2009) alkaloid and flavonoid compounds can act as stomach poisons. Therefore, if the alkaloid and flavonoid compounds enter the insect's body, their digestive organs will be disturbed.

III. RESULTS AND DISCUSSION
Flavonoids found in the stems and leaves of Bintaro are chemical compounds that have insecticidal properties. Flavonoids attack several nerve organs in several vital organs of insects, resulting on nerves weakening, such as breathing and death. Flavonoids work as respiratory inhibitors. Inhibitors are substances that inhibit or decrease the rate of chemical reactions, flavonoids also affect the energy mechanism in the mitochondria by inhibiting the electron transport system (Roqib and Kristanti, 2015).
Saponins found in the stems and leaves of the Bintaro plant can affect the absorption of minerals and vitamins. Southon et al. (1988) stated that saponins reduce iron absorption in tested mice. The decrease in absorption is more due to the influence of the disturbance of Fe transport through mucosal cells compared to the bonds formed between Fe and saponins.