Unlike most other NMR experiments for the measurement the sign and magnitude of the three-bond 1 H 1 -13 C II coupling across the glycosidic bond in sucrose at natural isotopic of scalar coupling constants, E.COSY-type NMR experiments offer the additional advantage of containing informa-abundance (Fig. ).
The DQ / ZQ-HC experiment of Fig. is designed for tion on the signs of the coupling constants (1-8). The experiments require at least three spins, A, M, and X. The cou-the generation of heteronuclear mixed DQ / ZQ coherence between the spins H 2 and C II during the evolution time plings J AM and J MX are measured from the A-X cross peak recorded in an experiment where the third spin M is left t 1 , where the DQ and ZQ coherences evolve with the sum and difference of the couplings to the common coupl-undisturbed. The relative signs of the J AM and J MX couplings are encoded in the tilt of the cross-peak multiplet pattern. ing partner H 1 . Using the product-operator notation of Sørensen et al. ( ) and neglecting signs, trigonomet-The absolute signs of the scalar couplings can be determined by relating them, e.g., through a series of E.COSY-type ric factors, and normalization constants, the coherence between H 2 and C II is generated by the following coher-experiments (9), to the sign of a large one-bond 1 H-13 C coupling which is always positive (10, 11). Since hetero-ence transfer pathway:
C)] nuclear three-bond coupling constants can assume positive as well as negative values (10-12), the sign information is -H 1z H 2y C IIy . The term H 1z H 2y C IIy represents the desired mixed DQ / ZQ coherence which evolves during t 1 with necessary for accurate dihedral angle determinations based on Karplus curves.
passive couplings to H 1 . The subsequent 90Њ ( 1 H, 13 C ) pulses convert it back to H 1y H 2z C IIz . This coherence is Sign information is also contained in the recently developed double-quantum/zero-quantum (DQ/ZQ) experiment detected during t 2 at the frequency of H 1 with a multiplet fine structure which is doubly anti-phase with respect to (13). The original work showed that the DQ/ZQ scheme offers improved accuracy in the measurement of scalar cou-J ( H 1 , H 2 ) and J( H 1 , C II ) . If D does not perfectly match 1 / [ 2 J ( H 1 , H 2 ) ] , the mixed DQ / ZQ coherence term pling constants in macromolecules (13). The present Communication shows that, unlike E.COSY experiments, DQ/ H 1x C IIy is present during t 1 , too. This term cannot be removed by phase cycling, but it would generate cross ZQ experiments also yield the relative signs of the couplings J AM and J MX in linear spin systems A-M-X, where J AX Å peaks at F 1 frequencies different from those of the desired cross peaks. 0. The principle is demonstrated with the determination of FIG. 1. Atom numbering in sucrose. The atoms involved in the linear spin system H 2 -H 1 -C II are labeled in bold. Sign and magnitude of the coupling constant J(H 1 , C II ) are measured by the experiment of Fig. 2. 503