Determination of non-cytotoxic dose ranges of selected plant extracts in RBL-2H3 cells

Abstract

Background: Globally, research on identifying bioactive compounds from medicinal plants and characterizing their in vitro and in vivo pharmacological activities is progressing rapidly. Plants native to Mongolia have adapted to unique, extreme continental climatic conditions, accumulating distinct phytochemicals, and have historically been utilized in traditional medicine and dietary practices. Establishing the non-cytotoxic safety threshold of these wild and cultivated plant extracts in RBL-2H3 mast cells a universally accepted model for type I hypersensitivity and inflammation is a vital prerequisite. Determining these baseline non-toxic dose ranges provides the essential scientific foundation for subsequent cellular allergy models, animal studies, and the elucidation of molecular mechanisms underlying their therapeutic efÏcacy. Aim: To determine the cytotoxicity profile and establish the non-cytotoxic dose ranges of extracts from four selected Mongolian plant species (wild and cultivated) in RBL-2H3 cells. Materials and Methods: Plant samples including lingonberry (Vaccinium vitis-idaea), strawberry (Fragaria orientalis), black radish (Raphanus sativus), and blue-berried honeysuckle (Lonicera caerulea L.) were standardly collected, authenticated, and cataloged as herbarium specimens at the Botanic Garden and Research Institute, Mongolian Academy of Sciences. A total of 12 distinct extracts were prepared from plant leaves, stems, and fruits using water and ethanol extraction, followed by concentration via rotary evaporation and freeze-drying. Rat basophilic leukemia (RBL-2H3) cells were maintained in EMEM growth medium and utilized during the exponential growth phase. Cell viability and non-cytotoxic concentration thresholds were evaluated using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. Results: Ethanolic extracts of strawberry leaves, black radish, and honeysuckle leaves (≥6.25 µg/mL), as well as aqueous extracts of honeysuckle fruits (≥6.25 µg/mL) and stems (≥25 µg/mL), exhibited dose-dependent cytotoxicity. Conversely, aqueous extracts of lingonberry leaves (≤200 µg/mL), strawberry leaves (≥100 µg/mL), and black radish (≤200 µg/mL), alongside ethanolic extracts of lingonberry leaves (≥50 µg/mL), demonstrated non-cytotoxic profiles. Furthermore, ethanolic extracts of honeysuckle stems (≤300 µg/mL) and fruits (≥12.5 µg/mL), as well as the aqueous extract of honeysuckle leaves (≤50 µg/mL), significantly enhanced RBL-2H3 cell viability. Conclusion: The tested plant extracts influenced RBL-2H3 cell viability in a solvent polarity and plant organ dependent manner. Ethanolic extracts of strawberry leaves, black radish, and honeysuckle leaves, along with aqueous extracts of honeysuckle fruits and stems, demonstrated relatively high cytotoxicity, reducing cell viability by >40% starting at concentrations of 12.5, 25, and 50 µg/mL. In contrast, aqueous extracts of strawberry leaves and black radish, as well as both aqueous and ethanolic extracts of lingonberry leaves, maintained low cytotoxicity at concentrations below 200 µg/mL. Notably, ethanolic extracts of honeysuckle stems and fruits (>12.5 µg/mL) and its aqueous leaf extract (<100 µg/mL) enhanced cell viability, exhibiting significant cell proliferation promoting activity.