Open AccessDetermination of Protonation Sites in Thermospermine and in Some Other Polyamines by 15 N and 13 C Nuclear Magnetic Resonance Spectroscopy
Author(s) -
TAKEDA Yasuko,
SAMEJIMA Keijiro,
NAGANO Kozo,
WATANABE Mayumi,
SUGETA Hiromu,
KYOGOKU Yoshimasa
Publication year - 1983
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1983.tb07164.x
Subject(s) - protonation , titration , chemistry , titration curve , chemical shift , nmr spectra database , polyamine , nuclear magnetic resonance spectroscopy , proton nmr , spermidine , putrescine , carbon 13 nmr , analytical chemistry (journal) , nuclear magnetic resonance , crystallography , stereochemistry , spectral line , organic chemistry , physics , biochemistry , ion , astronomy , enzyme
The p K values of thermospermine, a novel asymmetric tetraamine (4,8‐diazadodecane‐1.12‐diamine), have been determined for the first time by the conventional titration method with subsequent data analyses. It was found to be the most basic polyamine among naturally occurring tetraamines. The 15 N NMR titration method was applied to determine its protonation site. Natural‐abundance 15 N magnetic resonance spectra of the polyamine have been recorded as a function of pH from its free to the protonated form. Its apparent p K values, as well as the intrinsic chemical shifts for each stage of protonation, have been calculated from the NMR titration curve by using a nonlinear least‐squares method. The four protonation sites of thermospermine have been successfully determined to be N 12 , N 1 , N‐8, and N‐4 in that order from the free to the completely protonated form. In a similar manner the protonation sites of spermidine and two symmetric tetraamines, thermine and spermine, have been established. 13 C NMR experiments for the four polyamines were almost consistent with those of 15 N NMR. Owing to the much higher concentration of the polyamines furnished for NMR titration experiments, the p K values obtained by NMR were larger than those derived by the conventional titration methods.