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Use a stainless steel column (20 cm ¡Á 4.6 mm) packed with end-capped  octadecylsilyl silica gel for chromatography (5 µm) (Hypersil ODS is suitable).
Use a flow rate of 2 mL per minute
48 volumes of methanol  and 52 volumes of a phosphate buffer prepared by  dissolving 4.91 g of sodium dihydrogen orthophosphate and 0.633 g of disodium  hydrogen orthophosphate in sufficient water to produce 1000 mL.
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frwang1983

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