Cytotoxic Evaluation of Alchemilla vulgaris Flavonoids in Normal Blood Lymphocytes

The discovery and identification of a new drugs, which can potentiate the immune function has become an important goal of researches in immunepharmacology.This study demonstrates the favorable effects
of wild type Alchemilla vulgaris active components as immunomodulating agent. A. vulgaris is an important medicinal plant, represented a rich sources of biologically active phytocompounds, which are beneficial for humans. A. vulgaris which is widely known as lady’s mantle, bear’s foot or lion’s foot is traditionally used due to their tannin content for the treatment of inflammation of the upper digestive tract, diarrhea internally and as wound healing and astringent externally. It is also used as gargle against mouth and throat inflammation. Lady’s mantle is reported to be an important remedy in folk medicine in Europe. It is used to heal inflammations in mouth, bleeding of the nose and gynecological diseases. The plant aerial parts dried and extracted in 80% methanol by maceration. Chemical detection of crud plant extracts was performed. The total flavonoids were isolated, subjected to thin layer chromatography (TLC) using different mobile systems. The purified material was read by using high performance liquid chromatography (HPLC).A.vulgaris contain total flavonoids (9.8mg/g dried whole plant) calculated as Rutin. A.vulgaris also contains quareciten and kaempferol, both of them were investigated qualitatively and quantitatively in the present study but their quantities in this herbal plant were less than Rutin.The immunomodulation occur by treated extracted semi-purified flavonoids with Lymphocytes isolated from healthy people to measure the cytotoxic effect of extracted Flavonoids on normal lymphocytes by using MTT dye. MTT is widely chosen as a cell viability-measurement for optimal end point. MTT is a yellow water-soluble tetrazolium dye {3-(4, 5-Dimethylthiazol-2-yl)-2, 5diphenyltetrazolium bromide} reduced by live, but not the dead cells to a purple formazan product that is insoluble in aqueous solution. The
cytotoxic evaluation was read by using sandwich ELISA technique at 450nm. The result of treatment the isolated flavonoids with normal lymphocytes show significant effect by suppressing the lymphocytes
proliferation with the decrease in the extracted flavonoids concentration as compared with control (untreated lymphocytes in suitable media). The immunomodulation effects of A.vulgaris were caused alteration in the IL-2 & IL-10 level estimated by ELISA technique, The extracted flavonoids cause significant increase in both IL-2 and IL-10 level that produced from treated lymphocytes as compared with negative control (normal lymphocytes with media). The reduction in the extracted A.vulgaris concentration accompanied with significant increase in both IL-2 and IL-10 in 2hours and 4hours.  The present study reveal the effects of semi-purified flavonoids on IL-2 and IL-10 production indicated by alteration in gene expression of normal lymphocytes in vitro toward IL-2 and IL-10  production in response to all concentrations.