TOCSY - total correlation spectroscopy
A fundamentally different method for the transfer of coupling information is utilized in the TOCSY experiment. In contrast to the COSY experiment, in-phase signals are obtained, I.e. the following transfer takes place: I1x I2x . Mixing is achieved while a spin-lock field is applied, and ideally the effect of chemical shift is completely removed. This means, that basically all energy levels are equivalent or “degenerate”, which leads to strongly coupled systems. The spin-lich pulses are termed “isotropic mixing” or Hartmann-Hahn mixing. Isotropic mixing leads to spectra with mixed phases (phase modulation), whereas Hartmann-Hahn mixing pure phases are obtained. The multiplets, even when generated over different mixing times, give rise to combinations of different mirror-symmetric multiplet patterns. Ideally cross peaks are formed between all nuclei in a spin system.
Several pulse sequences are proposed for the TOCSY experiment. The most successful utilizes is so-called MLEV-17 Hartmann-Hahn mixing pulse. In this pulse scheme, a series on 90/180 pulses leads to a constant flip of the magnetization between the Iz and -Iz direction. Another very good sequence is the DIPSI sequence (series of 180). The effect is identical, differences are only the way the spin flip is caused, and how well the 180/90 pulses and their phases are designed to compensate for slight errors in pulse length and phase/direction.
TOCSY spectra are particularly useful in the analysis of bio-macromolecules, as the in-phase signals are insensitive to cancellation (due to small coupling constants). The number of nuclei to which magnetization is distributed is controlled by the mixing time, the length of the spin lock pulse. Short mixing times ( 30-40 ms) produce cross signals practically only between directly coupled nuclei, whereas longer mixing times give rise also to relayed and multiple relayed peaks (65-90 ms).