Kajian Efek Imunomodulator Ekstrak Etanolherba Binara (Artemisia vulgaris L.) secara In Vitro dan In Vivo

Abstract

Herbs from the Binara plant (Artemisia vulgaris L.) are potentially medicinal herbs from Tanah Karo, North Sumatra which have been used traditionally to overcome skin infections and hypertension. Secondary metabolites and contained marker compounds become parameters of pharmacological potential. Infectious diseases require alternatives to synthetic drugs. Research was conducted to analyze the immunomodulatory effects of herbal binara. The test was carried out by qualitative and quantitative determination of secondary metabolites, examining the immunomodulatory effects of extracts both in vitro and in vivo. Pemeriksaan secara kualitatif dan kuantitatif dengan menggunakan High Performance Liquid Chromatography (HPLC) dan Liquid chromatography-mass spectrometry (LCMS). Kadar flavonoid total ekstrak ditetapkan dengan menggunakan Spektrofotometri UV-Visibel. Pengujian efek imunomodulator dilakukan dengan metode pengujian secara in vitro dan in vivo. Pengujian in vitro dilakukan dengan pengujian viabilitas sel untuk melihat konsentrasi sel yang paling aman dan digunakan untuk pengujian proliferasi limposit dengan Microtetrazolium (MTT) assay. In vitro testing is done with cell viability testing to see the safest cell concentration and is used for testing lymphocyte proliferation with Microtetrazolium (MTT) assay. IL-6 gene expression testing using the highest safe concentration was used to analyze gene expression in RAW 264.7 cells by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) method. In vivo testing was carried out by giving extracts for 14 days to nonimmunosuppressant mice and suppressants given cyclophosphamide on days 8 and 13 infected with Staphylococcus aureus on day 4 measuring total leukocytes and differentials, Ig-G levels, and IL-2 with the Enzyme-linked immunosorbent assay (ELISA) method. The results showed that the extract containing quercetin compounds indicated by ethanol extract of binara herb (EEHB) showed a retention time of 5.27 minutes with m/z 300.18-301.18 and quercetin levels in the extract with a concentration of 6.25 ppm of 7.14 μg/ml, 12.50 ppm of 13.42 μg/ml, 25.00 ppm of 23.77 μg/ml, 50.00 ppm of 48.55 μg/ml, and 100 ppm at 100.85 μg/ml. The results of cell viability testing from EEHB showed no toxic effect at a concentration of 12.50; 25 μg/mL, levamisole 1.25; 2.5 μg/mL, quercetin 1.25 μg/mL with percentage of living cells (>90%). EEHB, levamisole, and quercetin increased lymphocyte proliferation with an EC50 value of 4.527; 4,282; 2.436 μg/mL. EEHB is able to increase lymphocyte proliferation. EEHB also increased IL-6 gene expression as seen from absorption band thickness. EEHB also increased the number and differential of leukocytes such as lymphocytes, neutrophils, basophils, and eusinophils, Ig-G levels, and IL-2 levels in nonimmunosuppressant mice and suppressants infected with Staphylococcus aureus. Based on the results of the study, it was concluded that EEHB contains quercetin. EEHB and quercetin did not show toxic effects on RAW 264.7 cells, increased lymphocyte cell proliferation and IL-6 gene expression, also increased total and leukocyte differential, Ig-G, IL-2 cytokine, so EEHB is effective as an immunostimulant.