Process for purifying secretin
AssigneeEisai Co., Ltd.
InventorsMikio Uchiyama, Takashi Sato, Hiroshi Yoshino, Yutaka Tsuchiya, Masayuki Konishi, Masahiko Tsujii, Yoshihiko Hisatake, Atsushi Koiwa
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Abstract
Purified secretin is obtained by subjecting crude secretin to reversed-phase high-performance liquid chromatography using, as an eluant, a solvent mixture of an acid or salt thereof and an organic solvent.
Claims10
A process for purifying crude secretin selected from the group consisting of synthetic crude secretin and natural crude secretin, which comprises subjecting the crude secretin to reversed-phase, high-performance, liquid chromatography using, as an eluant, a solvent mixture of (1) an aqueous solution of a substance selected from the group consisting of hydrochloric acid, trifluoroacetic acid, phosphoric acid, perchloric acid, sodium phosphate and sodium perchlorate, and (2) a polar organic solvent which is miscible with water and is selected from the group consisting of acetonitrile, methyl alcohol, ethyl alcohol, isopropyl alcohol and tetrahydrofuran, and using, as a stationary phase, column packing of silica gel particles having alkyl or cyanopropyl groups chemically bonded thereto, and thereby obtaining a purified secretin having a biological activity of higher than 5000 cu/sq;
- (as acetate) and higher than 6000 cu/mg (on dry, free basis).
A process for purifying a natural or synthetic crude secretin which comprises dissolving said crude secretin in a mixture of (1) an aqueous hydrochloric acid solution and (2) acetonitrile, to form a solution of said crude secretin;
- then subjecting said crude secretin solution to reversed-phase high-performance liquid chromatography using a column packing of silica gel having alkyl or cyanopropyl adsorbent groups chemically bound thereto as the stationary phase and using said mixture as eluent;
- then collecting the elute corresponding to a secretin peak fraction, as determined by an elution curve prepared at 210 nm UV;
- and then freeze-drying the collected secretin to obtain a purified secretin having a biological activity higher than 5,000 cu/mg, as acetate, and higher than 6,000 cu/mg on a dry, free basis.
Dependencies are inferred from the claim text.