J. CHEM. SOC., CHEM. COMMUN., 1983 zyxwvutsr 1289 z Isotope-induced Nonequivalence in a Symmetrical Molecule: Measurement of the 31P-31P Geminal Coupling Constant in Pyrophosphate Thomas M . Marschner, Mark A. Reynolds, Norman J. Oppenheimer," and George L. Kenyon" zy Department of Pharmaceutical Chemistry, University of California, San Francisco, zyxw CA 94743, zy U.S.A. zy Asymmetric incorporation of zyxwvut *O into pyrophosphate to perturb the 3l P n.m.r. resonance frequency allows determination of the 31 P-31 P geminal coupling constant in this otherwise symmetrical molecule. We report the use of selective lSO labelling of one phosphorus atom in pyrophosphate to induce chemical shift non- equivalence of the two phosphorus nuclei and thus permit observation of the individual resonances and measurement of an otherwise unobservable mutual coupling constant. lSO induces a small upfield shift in the 31P n.m.r. resonance frequency of phosphates zyxwvutsrq (ca. 0.02 p.p.m./lsO bond), the magnitude of which is related to the P-0 bond ~rder.l-~ Recent work in this laboratory involving the synthesis of 1sO-/3,y-bridge-labelled ATP for use in positional isotope exchange experiments produced an interesting result while assigning resonances in the 31P n.m.r. spectra of lsO-labelled pyrophosphate intermediate^.^ At 202.5 MHz, the 31P n.m.r. spectrum of pyrophosphate with one nonbridging la0 appeared as a closely spaced doublet with a separation of 0.5 Hz. This splitting was presumed to be due to coupling between the nonequivalent phosphorus atoms with the observed doublet corresponding to the central doublet of an AB pattern. By selectively increasing the number of lSO atoms attached to one of the two phosphorus atoms in the pyrophosphate molecule we can increase the nonequivalence of the phosphorus atoms sufficiently to allow observation of Published on 01 January 1983. Downloaded by University of Windsor on 23/10/2014 18:52:24. View Article Online / Journal Homepage / Table of Contents for this issue