The effect of natural ingredients extracts on the phagocytosis index of the carbon clearance animal models

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INTRODUCTION
The immune system is a mechanism that develops the body's capacity to combat pathogens which would exclude numerous external substances that are taken in order to prevent disease.The body's defense mechanisms against pathogens are all part of the immune system, which can be divided into two types: the non-specific innate immune system and the specific adaptive immune system.Both have cellular and humoral components.The innate immune system will react quickly and nonspecifically if the pathogen breaches the physical barriers.The innate immune system is not able to continuously defend against a given infection (Marshall et al., 2018).
Original Article The effect of natural ingredients extracts on the phagocytosis index of the carbon clearance animal models JAI, Vol 1 No 1, 2023 doi: Immunomodulators are substances that can affect the immune system and have the ability to enhance the immune response or protect against pathogens or tumors.Immunomodulators include immunostimulants, which can boost the immunological response, immuno-suppressants, which can reduce the immune response, and immunorestorants, which can enhance immune system performance (Mahima et al., 2013).
Some plants and natural ingredient have the ability to modify the human immune system, hence enhancing the immune system's ability to combat infections and reducing the occurrence of illness (Swaroop et al., 2022).Immunomodulatory plants typically include secondary metabolites that modulate the immune system by acting as antioxidants during cell metabolism, as enzyme cofactors, promoting the differentiation and proliferation of B and T cells, influencing cytokine synthesis, and decreasing histamine levels (Kumar and Yadav, 2022).

Production of the Simplicia
The simplicia that have been collected are determined at the Pharmacognosy Laboratory of Padjadjaran University.The test material was well cleaned with running water before being dried at 40°C for 24 hours, then the simplicia was ground into a powder using a blender, and last it was sieved.
Extraction 200 grams of the test substance that had been ground into a powder were weighed, placed in a flask with a flat bottom, and then covered with 50% ethanol.Reflux was performed three times for a total of 120 minutes on the filtrated reflux residue.
The filtrate was then dried in an oven at 50°C to produce a thick extract.

RESULTS AND DISCUSSION
The determination results showed that the plats were Ageratum conyzoides, Artocarpus heterophyllus, Nephelium lappaceum, Cucurbita moschata and Citrullus lanatus.
Phytochemical screening was carried out to determine the class of secondary metabolite compounds belonging to the plant.The results of phytochemical screening can be seen in Table 1.
The results showed that all simplicia contain flavonoids, polyphenols, quinones and monoterpenoids/ sesquiterpenoids.The stimulation index indicates the ability of active macrophages to phagocytize colloidal particles of carbon ink based on the optical density value.
The optical density and phagocytosis index values increase with the amount of phagocytosed colloidal carbon particles.The results of the carbon clearance test can be seen in Table 2. activity in modulating the immune response, by activating macrophages, triggering the work of NK cells in producing IFN-γ and suppressing the activity of mTOR mediators in producing T lymphocytes (Hosseinzade et al., 2019).Exocarp of C.lanatus contains the amino acid L-citrulline which is a precursor of L-arginine as a substrate for the synthesis of nitric oxide, which is an important signal molecule in cell regulation in binding radical hydroxyl groups (Aguayo et al., 2021).
It is believed that the Zn concentration in C.moschata contributes to RNA synthesis, lymphoid cell proliferation, T cell maturation, and the creation of regulatory cytokines that are important for the immune system, particularly for the removal of antigens (Hojyo and Fukada, 2016;Purnamasari et al., 2022).C.moschata seeds contain carotenoid compounds which also have antioxidant activity (Kulczyński, Sidor and Gramza-Michałowska, 2020).
This study was conducted to determine the fundamental properties of a substance for nonspecific immunomodulatory activity via the carbon clearance test.Therefore, further study on specific immunomodulatory activity using other animal models or cell lines is required.Specific immunomodulator evaluation may be performed using the humoral antibody response technique, delayed type hypersensitivity response, effect on total leucocyte count, and leucocyte mobilization studies.In vitro testing using a cell line can be performed on the K562 cell line, J 779 macrophage cell, K562 cell line and cutaneous squamous cell carcinoma cell line (Ganeshpurkar and Saluja, 2017).

CONCLUSION
Extracts that have potential as immunostimulants is A.heterophyllus and C. lanatus had potency as immunosupressant.It is important to conduct additional study on the immunostimulatory effect of these natural substances utilizing other, more specialized testing methods.
Blood was obtained from the test animals via the tail vein at the start of the experiment, lysed with 1% glacial acetic acid, and then the transmittance at T0 was determined.The test animals were then separated into eight groups: Control, Zymosan A 10 mg/kg BW, methylprednisolone 4 mg/kg BW, ethanol extract of A.conyzoides leaves 160 mg/kg BW, ethanol extract of A.heterophyllus leaves 500 mg/kg BW, ethanol extract of N.lappaceum peel 650 mg/kg BW, ethanol extract of C.moschata seed 342 mg/kg BW and ethanol extract of C.lanatus rind 55 mg/kg BW.For 15 days, every treatment was administered orally.Every animal fasted on the fifteenth day.On the sixteenth day, a suspension of carbon ink 0.025 mL/10g BW was administered to the animals.Following the injection of the carbon ink suspension, blood samples were obtained at 5, 10, 15, and 20 minutes.Blood samples 25 µL were diluted with 4 mL 1% acetic acid solution, and the transmittance was then determined using a visible spectrophotometer at a wavelength 650 nm.Then the stimulation constant (SC) and stimulation index (SI) were determined(Vikasari et al., 2015).The stimulation constant was calculated from the difference in optical density (OD) at 5 and 20 minutes to the difference in the time of blood sampling.The stimulation index was calculated from the comparison of the KS test to the KS control.Based on the SI value, the classification of the immunomodulatory effect of a substance is no effect (SI = 1.0-1.2),slight (SI=1.3-1.5) and strong

Table 1 .
The results of characteristics and phytochemical screening assay The effect of natural ingredients extracts on the phagocytosis index of the carbon clearance animal models JAI, Vol 1 No 1, 2023 doi: (Utama, Rosidah and Yuandani, 2020)specific immune system in phagocytosing carbon ink colloidal particles(Utama, Rosidah and Yuandani, 2020).

Table 2 .
The results of the immunomodulatory effect using the carbon clearance test Flavonoids have The effect of natural ingredients extracts on the phagocytosis index of the carbon clearance animal models JAI, Vol 1 No 1, 2023 doi: