4. Acquisition

Sample: 3% strychnine in CDCl₃ Record normal 1D ¹H and ¹³C NMR spectra, optimize the spectral widths for the aliphatic region only, and note the offsets. A narrow spectral window should be used be- cause of the digitization problem in 3D NMR. Switch to the 3D mode of the spec- trometer software and load the gs-HSQC-TOCSY pulse program. You have to set:

td3: 256 data points in F; (¹H) td2: 64 data points in F2 (¹H) td: 128 data points in F; (°c) sw3: 3.3 ppm sw2: 3.3 ppm swl: 56 ppm o1: middle of selected ¹H NMR region o2: middle of selected ¹H NMR region 03: middle of selected ¹³C NMR region p1, p3, p5: 90° ¹H transmitter pulse p2, p4, p6: 180° ¹H transmitter pulse p8, p9: 90° ¹³C decoupler pulse p7, p10: 180° ¹³C decoupler pulse 90° and 180° ¹H pulses within MLEV-16 spin-lock = 40 and 80 μs at transmit- ter attenuation of 16 dB (see Exp. 11.11); loop counter for duration of spin-lock = 40 (must be an even number), giving a spin-lock time of 100 ms dl:2s d2: 1/[4(C,H)] = 1.75 ms, calculated from 'J(C,H) = 145 Hz d3: delay to compensate the duration of field gradient = 1.5 ms g1, g2: sinusoidal shaped field gradients with 5% truncation, 1.5 ms duration and ca. 0.1 T/m strength, with gradient loop counters, ring-down delays (100 is), lock blanking and gradient coil blanking switches according to actual instrumentation used. Gradient strength ratio: 4:1 ¹³C decoupler attenuation and 90° pulse for GARP initial values for f, and f, evolution: 3 μs increment for f; evolution: 1/[2-sw1]; increment for f, evolution: 1/sw2

5. Processing

Apply zero-filling to 128 real data points in F2 and to 256 real data points in F, to ob- ain a matrix of 128x128x256 real data points. This will result in 4 MB of processed data, Use an exponential window with lb = 5 Hz line-broadening in F; and 7/2-shifted squared sinusoidal windows in the other two dimensions. Apply the correct acquisition order parameter (3-1-2) before FT in all three dimensions. Phase correction is not necessary, since a magnitude calculation is performed after the last Fourier transforma-