Figure 1 shows an example of a COSY and a NOESY cross peak. We now want to measure the coupling constant in the w2 dimension. This tool can only measure coupling constants along the first dimension in the spectrum. If you want to measure a coupling constant in the other dimension, swap the axes using the Spectrum Zoom window.
The window for setting up the measurement of coupling constants is shown in Figure 2. The method to determine the coupling constant is described in Ludvigsen et al. (Ludvigsen, S., Andersen, K.V. & Poulsen, F.M. (1991) J. Mol. Biol. 217, 731-736). The first two fields select the antiphase and the inphase cross peaks, respectively. The w2 range selects the range in the w2 range, in Hz, valid for both peaks. The two w1 ranges select the ranges for the two peaks in the w1 dimension. The two fields assume that the w2 dimension is dimension 0, and the w1 dimension is dimension 1. The method field selects the method for determining the scale factor, it can be set either to Automatic or Manual. The fields below are the values calculated, when the Go button is hit, except the Scale field: This is used for input if Manual is selected. The Update button transfers the coupling constant calculated into the cross peak catalog. The Separation field is the difference, in Hz, between the two maxima of the antiphase peak, and The Linewidth is the width of either the subtracted or added peak, at half peakheight. The Coupling is measured as the difference between the top position of the sum and difference curve. The Scale refers to the scale used before addition and subtraction of the traces, which both have been normalized to 1.0 relative to their extreme values. A scale factor of 1.0 is valid if the coupling constant is much larger than the line width. Values between 1 and 6 are normal, high values if the actual coupling is small relative to the line width. The method only works for inphase and antiphase doublets, however, small additional passive coupling will not disturb the result. In manual mode (Method: Manual), the routine can be used as a DISCO (Kessler, H. Müller, A. & Oschkinat, H. (1985) Magn. Reson. Chem. 23, 844-852) procedure, if small values for w1 antiphase range are used.
Upper left: antiphase peak.
Upper right: inphase peak.
Lower left: added spectrum.
Lower right: subtracted spectrum.
When the GO button is pressed, a result window, Figure 3, pops up. This window can be used to measure some of the values manually. The value to be measured manually is selected at the bottom of the window. All values measured are ranges, measured in Hz. The left and right of the range to be measured can be set in any of the windows, independently. The left range is set using the left mouse button, and the right range is set using the middle mouse button. When the right range is set, the width of the range is stored as the currently selected value to be modified.
Figure 4 shows the result after performing the calculation of the coupling constant. The result in this case is 5.4 Hz.