SummaryA highly purified intrinsic factor preparation (RAS fraction) was prepared in quantity from fresh hog duodenum by saline extraction, pH adjustment and ammonium sulfate precipitation. This fraction was active in pernicious anemia patients in relapse at daily oral dose of 5 mg. Ultracentrifugal separation could not completely separate the components of RAS fraction. RAS fraction was further purified on a DEAE-cellulose ion exchanger to give a fraction which was active in urinary excretion test at a dose of 3 mg. equivalent to a daily oral dose of 1 mg in pernicious anemia patients in relapse. Rechromatography of this active fraction resulted in no further increase in potency. Ultracentrifugal studies of the most active fraction obtained from rechromatography showed essentially 3 peaks with sedimentation constants of 1.0 S, 2.8 S and 11 S.