4. Acquisition

Time requirement: 1.5 h

Sample: 100 mg cholesteryl acetate in CDCl₃.

Load standard ¹³C NMR parameters and the DEPT pulse program. You have to set:

decoupler attenuation for hard decoupler pulses
td	64 k
sw	200 ppm
o1	middle of ¹³C NMR spectrum
o2	middle of ¹H NMR spectrum
p1	90° ¹H decoupler pulse
p2	180° ¹H decoupler pulse
p3	use 45°, 90°, and 135° ¹H decoupler pulses for three successive experiments leading to spectra a, b and c. Spectrum a will give the signals of CH, CH₂, and CH₃ groups all positive, b gives only the signals of CH groups, and the third spectrum c gives the signals of CH and CH₃ groups positive and signals of CH₂ groups negative. The second spectrum b gives a clear indication of whether the decoupler pulse was determined correctly.
p4	90° ¹³C transmitter pulse
p5	180° ¹³C transmitter pulse
d1	2 s
d2	1/[2¹J(C,H)] = 3.5 ms, calculated from ¹J(C,H) = 140 Hz
decoupler attenuation and 90° pulse for CPD
ns	512

5. Processing

Use standard 1D processing as described in Experiment 3.2. For editing purposes the three spectra have to be further manipulated. Subtraction of 0.8·b from a yields spectrum d, where the signals of CH₂ and CH₃ groups both remain positive. Subtraction of 0.6·b from c yields spectrum e, where the signals of CH₂ are negative and those of the CH₃ groups remain positive. Subtraction of 1.2·e from d yields spectrum f with only signals of CH₂ groups, whereas addition of 1.2·e to d yields spectrum g with only signals of the CH₃ groups. The factors of 0.8, 0.6, and 1.2 may be finely adjusted according to the exact duration of pulse p3.

6. Result

The figure shows the three edited DEPT subspectra b (CH groups), f (CH₂ groups), and g (CH₃ groups), obtained from the three different measurements a–c on an AM-400 spectrometer and calculated as described.