Application of oxime formation in a radiometric assay of aminooxy compounds.
A radiometric determination of monoaminooxy analogues of naturally occurring polyamines is described in which [2-14C]acetone is employed as reagent. The reagent is volatile while the oxime product is not allowing unreacted reagent to be removed and the oxime formation to be completed by lyophilization in vacuo. The residual radioactive compound is soluble in water and proportional to the reactive aminooxy content of the reaction mixture and can be quantified by liquid scintillation counting. The assay method is inexpensive and simple and has high specificity and flexibility in sample volume enabling reliable quantification of reactive aminooxy amines in biological extracts at concentrations exceeding 0.25 microM. Optimal pH values for oxime formation of five monoaminooxy analogues of polyamines with acetone were resolved. Reactions via reversible intermediates to irreversible oximes were sped up by removal of water. Complete oxime formation and stability was confirmed by 1H NMR studies. Tested drugs readily formed oximes with pyridoxal 5'-phosphate, too. Diaminooxy analogue of cadaverine formed a volatile oxime with acetone. The method was used to monitor the stability of aminooxy analogues of putrescine and spermidine during storage and under culture conditions and to establish their scant accumulation in, but fast catabolism by, cultured baby hamster kidney cells.
Keinänen
T A
TA
Department of Biochemistry and Biotechnology, University of Kuopio, Finland.
Hyvönen
T
T
Khomutov
A R
AR
Khomutov
R M
RM
Kauppinen
R A
RA
Vepsäläinen
J
J
Eloranta
T O
TO
D000588:Amines / Q000032:analysis* / Q000493:pharmacokinetics; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D004355:Drug Stability; D005069:Evaluation Studies as Topic; D006863:Hydrogen-Ion Concentration; D010091:Oximes / Q000032:analysis*; D011437:Propylamines / Q000032:analysis; D011732:Pyridoxal Phosphate; D012588:Scintillation Counting / Q000379:methods*
D000588:Amines; D010091:Oximes; D011437:Propylamines; D011732:Pyridoxal Phosphate; C046252:1-aminooxy-3-aminopropane