Estraction conditions of proteins from Echinacea Purpurea L. (Moench) roots and leaves

Abstract

The purple coneflower (Echinacea purpurea L. (Moench)) has been known and used in traditional medicine for decades, but the interest of this plant pharmacological properties isn't decreasing. Previous studies showed hemagglutinating activity of ethanolic extracts from purple coneflower roots and leaves. Nevertheless, information about purple coneflower proteins and their hemagglutinating properties are still not defined. Aim of experiment: To determine the best extraction conditions with highest protein yield and highest hemagglutinating activity. Experiment tasks: 1. Extraction of proteins from coneflower roots and leaves. 2. Determination of extraction conditions for the highest protein yield. 3. Determination of extraction conditions for the highest hemagglutinating activity. Materials and methods: 1. Proteins were extracted out of coneflower leaves (ratio of herb material and extraction buffer - 1:10) and roots (ratio of herb material and extraction buffer - 1:6) in different conditions: A. At 20±2°C for 2 hours in 3 buffers with different pH (Written below); B. At 4±1°C temperature for 2 hours with 5% of protese inhibitor cocktail (Halt™, Thermo Scientific). C: At 4±1°C for 2 hours with 1% of polivynyl polypirolidone, 0,5% of dithiotrethiol and 0,5% of ethylenediaminetetraacetic acid. After extraction proteins were precipitated with 13.3% trichloracetic acid in acetone and 0.2% of β-mercaptoethanol. 2. Protein amount was measured by Bradford assay. 3. Determination of hemagglutinating activity: protein fractions were poured on suspension with 2% trypsin treated rabbit erythrocytes and incubated for 30 min at 20±2°C temperature. Buffers used for extraction: Acetic buffer pH=5.0, Phosphate buffered saline (PBS) pH=7.4, Tris-HCI buffer pH=8.0. [...]Botanikos sodasLietuvos agrarinių ir miškų mokslų centrasLietuvos sveikatos mokslų universitetas. Medicinos akademijaVytauto Didžiojo universiteta