2-Se-nucleosides were further purified at room temperature using a Knauer HPLC system equipped with a Smartline Detector 2600 and an Azura P2.1 L pump. A reversed phase Kinetex® 5 µm Evo C18 column (250 x 21.2 mm) and a flow rate of 21.24 mL min -1 were used. Samples were analyzed at 210 nm. Deionized water and acetonitrile were applied as eluents, while the gradient was modified from the analytical method as summarized in Table . The product purified by silica chromatography was dissolved in deionized water (Table ) and filtered either with a 0.45 µm PES syringe filter or with a 0.45 µm cellulose nitrate filter using a vacuum pump. The collected product fractions were kept on ice to prevent product degradation during the purification process. Collected 2-Se-nucleoside fractions were dried using a Christ Gamma 1-20 freezedryer (Osterode am Harz, Germany).