The 3D NOESY-NOESY (TONO) experiment is specifically designed to obtain homonuclear ¹H-¹H NOE connectivities in a single experiment. The experiment found widely application to study such effects and for the structural refinement of large unlabeled biomolecules in whose 2D NOESY spectra afford severe overlap of signals. The 3D NOESY-NOESY experiment contain more information than the corresponding 2D NOESY counterpart and, in addition, dipolar cross-relaxation effects are more efficient with increased molecular weight.

Implementation on AVANCE spectrometers is feasible.

The 3D NOESY-NOESY pulse sequence ( 91JMR6262-95 , 93JMRB185-101 ) consists of two concatenated NOESY building blocks.

Several structural refinements methodologies have been specifically proposed to quantify cross-peaks intensities:

Two spin approximation ( 91JB83 )

Taylor series expansion of the NOE-rate equation ( 92JMR511-97 and 92JMB156 )

Direct gradient refinement ( 89JMR643-83 and 91JMR626-95 )

Hybrid-Hybrid matrix ( 95JMRB164-106 , 96MRCS125 , and 99JB209-14 )

The 3D NOESY-NOESY experiment could be recorded in automation mode.

The NOESY-NOESY experiment affords a 3D spectrum in which several cross-peaks can be present:

• Diagonal peaks.
• Cross-peak due to single NOESY transfer in the w₂=w₃ and w₁=w₂ planes.
• Cross-peak due to double NOESY transfers.

The basic principles of 3D NMR spectroscopy was published in 89JMR14-84 .