title: A mass spectrometric method to determine activities of enzymes involved in polyamine catabolism.
abstract: An analytical method for the determination of three polyamines (putrescine, spermidine, and spermine) and five acetylpolyamines [N(1)-acetylspermidine (N(1)AcSpd), N(8)-acetylspermidine (N(8)AcSpd), N(1)-acetylspermine, N(1),N(8)-diacetylspermidine, and N(1),N(12)-diacetylspermine] involved in the polyamine catabolic pathway has been developed using a hybrid tandem mass spectrometer. Heptafluorobutyryl (HFB) derivatives of these compounds and respective internal standards labeled with stable isotopes were analyzed simultaneously by TOF MS, based on peak areas appearing at appropriate m/z values. The isomers, N(1)AcSpd and N(8)AcSpd were determined from their fragment ions, the acetylamidopropyl and acetylamidobutyl groups, respectively, using MS/MS with (13)C(2)-N(1)AcSpd and (13)C(2)-N(8)AcSpd which have the (13)C(2)-acetyl group as an internal standard. The TOF MS method was successfully applied to measure the activity of enzymes involved in polyamine catabolic pathways, namely N(1)-acetylpolyamine oxidase (APAO), spermine oxidase (SMO), and spermidine/spermine N(1)-acetyltransferase (SSAT). The following natural substrates and products labeled with stable isotopes considering the application to biological samples were identified; for APAO, [4,9,12-(15)N(3)]-N(1)-acetylspermine and [1,4,8-(15)N(3)]spermidine ((15)N(3)-Spd), respectively; for SMO, [1,4,8,12-(15)N(4)]spermine and (15)N(3)-Spd, respectively; and for SSAT, (15)N(3)-Spd and [1,4,8-(15)N(3)]-N(1)-acetylspermidine, respectively.
authors.affiliation: Department of Molecular Medicine, Tokyo Metropolitan Institute of Medical Science, 2-1-6 Kami-kitazawa, Tokyo 156-8506, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Moriya
authors.forename: Kaori
authors.initials: K
authors.lastname: Iwasaki
authors.forename: Keijiro
authors.initials: K
authors.lastname: Samejima
authors.forename: Koichi
authors.initials: K
authors.lastname: Takao
authors.forename: Kohfuku
authors.initials: K
authors.lastname: Kohda
authors.forename: Kyoko
authors.initials: K
authors.lastname: Hiramatsu
authors.forename: Masao
authors.initials: M
authors.lastname: Kawakita
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry*; D000818:Animals; D057075:Enzyme Assays; D006801:Humans; D013058:Mass Spectrometry; D015394:Molecular Structure; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000737:chemistry*; D011073:Polyamines / Q000737:chemistry* / Q000378:metabolism*; D000097667:Polyamine Oxidase; D000099228:Diamine N-Acetyltransferase
chemical_list: D000123:Acetyltransferases; D000587:Oxidoreductases Acting on CH-NH Group Donors; D011073:Polyamines; D000097667:Polyamine Oxidase; D000099228:Diamine N-Acetyltransferase