Separation of putrescine oxidase and spermidine oxidase in foetal bovine serum with the aid of a specific radioactive assay of spermidine oxidase.
1. A sensitive and specific assay for spermidine oxidase is described. The method involves the separation of [14C]spermidine (substrate) from [14C]putrescine (product) and other 14C-labelled products on a Dowex 50 cation-exchange column: 92% of the putrescine applied to the column was eluted by 2.3 M-HCl, but this treatment left 96% of the spermidine bound to the column. Unchanged spermidine could be removed from the column by elution with 6 M-HCl. 2. By means of this assay, foetal and adult bovine serum were each shown to contain spermidine oxidase activity, putrescine being a major product of the oxidation of spermidine by the serum enzymes. 3. In foetal bovine serum, spermidine oxidase activity is separable from putrescine oxidase activity by chromatography on a cadaverine-Sephadex column, by gel filtration and by ion-exchange column chromatography. Putrescine oxidase was purified 1900-fold and spermidine oxidase 130-fold by these procedures. The former oxidized putrescine but not spermidine, and spermidine oxidase exhibited no activity with putrescine as substrate.
D000818:Animals; D002250:Carbon Radioisotopes; D002417:Cattle; D002852:Chromatography, Ion Exchange; D002853:Chromatography, Liquid; D005312:Fetal Blood / Q000201:enzymology*; D006851:Hydrochloric Acid; D008722:Methods; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000097:blood* / Q000302:isolation & purification; D011700:Putrescine / Q000097:blood / Q000302:isolation & purification / Q000378:metabolism; D012588:Scintillation Counting; D013095:Spermidine / Q000378:metabolism; D000097667:Polyamine Oxidase
D002250:Carbon Radioisotopes; D006851:Hydrochloric Acid; D000587:Oxidoreductases Acting on CH-NH Group Donors; D011700:Putrescine; D013095:Spermidine; D000097667:Polyamine Oxidase; C022488:putrescine oxidase