Abstract
Background:
Peronema canescens Jack is a botanical with immunostimulant and immunosuppressive properties that is capable of modulating the immune system.

Aim:
The immunomodulatory properties of fractions from et leaves, specifically the fraction made from ethyl acetate (PcEA), n-butanol (PcB), and water (PcW) solvents, were evaluated in vitro and then characterized using Fourier transform infrared (FT-IR). Gas chromatography-mass spectrometry (GC-MS) and gene expression were used to analyze the fraction with the best immunomodulatory activity.

Methods:
P. canescens was extracted in 96% ethanol. A liquid-liquid partitioning method yielded PcEA, PcB, and PcW. After testing RAW 264.7 cells for immunomodulation, FTIR characterization was performed. The fractions with the highest immunomodulatory activity metabolites were identified using GC-MS and gene expression profiling. Three replicates were prepared for each parameter assay.

Result:
All fractions at 3.1 to 50 µg/mL concentrations exhibited no toxicity to RAW 264.7 cells in the viability assay. The phagocytic capacity assay showed that all fractions significantly improved phagocytosis compared with the control group (p < 0.05). Three fractions at 1.56 µg/mL significantly enhanced nitric oxide (NO) release relative to the untreated group (p < 0.05). In contrast, PcEA at 3.13 µg/mL tended to lower NO production than the control (p > 0.05). TNF-α production revealed that all fractions at all doses significantly increased in comparison to the control group (p < 0.05). IL-6 production of PcEA at 25 and 50 µg/mL concentrations did not differ substantially from the control (p > 0.05). IL-10 production of PcEA at 25 and 50 µg/mL concentrations, and PcB at 12.5 µg/mL, was significantly elevated compared to the control (p < 0.05). The GC-MS analysis of PcEA revealed eight chemicals, including terpenoids, steroids, fatty acids, and siloxane. TNF-α, IL-6, and IL-10 gene expression demonstrate that PcEA modulates the immune system.

Conclusion:
PcEA has potential as an immunomodulatory agent in vitro. Future research should focus on the isolation and purification of the identified compound and in vivo and mechanistic studies to confirm its therapeutic potential

Key words: Cytokine; FT-IR analysis; GC-MS analysis; Immunomodulatory; Peronema canescens.