71 For a Chn group, we get a signal with the amplitude sin 0 cos(n-0 in the DEPt experiment, compared to sin(TJA,) cosm-( A,) in the INEPT. So the dependence of the DEPT spectrum on the flip angle of the 0 pulse is the same as the dependence of the INEPT on the length of A2 However, the DEPT is much less sensitive to varying JHC values and therefore the preferred experiment for multiplicity editing( usually with a set of three 0 values,θ=45°,90°,135°; corresponding to the INEPT with A2=14J, 12J,314J) Inverse heteronuclear spectroscopy Proton detection Today, most of the heteronuclear experiments are performed in a H detected version, also called inverse detection"(in contrast to the classical X nucleus detection described so far). If the pro equipment is available (re-wired spectrometer console, inverse detection probe!), then inv detection offers such an immense gain in sensitivity that there is(almost) no reason to run any conventional heteronuclear correlation experiments anymore Theoretical relative sensitivities (S/N)for H, X correlation spectra(X=]C, N)*. Method Yexc. Yet 3/2 13C 15 direct detection YXY 2 1.0 10 INEPT/ DEPT YHYx/2 4.0 9.9 reverse INEPT 7.9 31.0 (relative to INEPT=1) 2.0 Invers YHWH 31.6 306.0 (relative to INEPT=1) 31.0 not taking into account other factors, e.g Tl, heteronucl noe, linewidths etc. It has to be remembered that the number of scans (spectrometer time) required goes up with the square of the sensitivity ratio. Thus, a simple id C spectrum might well need almost 1000 times the measuring time of an inverse 2DH. C-correlation
71 For a CHn group, we get a signal with the amplitude sin q cos(n-1) q in the DEPT experiment, compared to sin(pJD2 ) cos (n-1) (pJD2 ) in the INEPT. So the dependence of the DEPT spectrum on the flip angle of the q pulse is the same as the dependence of the INEPT on the length of D2 . However, the DEPT is much less sensitive to varying 1JHC values and therefore the preferred experiment for multiplicity editing (usually with a set of three q values, q = 45°, 90°, 135°; corresponding to the INEPT with D2 = 1/4J , 1/2J , 3/4J ). Inverse heteronucleare spectroscopy Proton detection Today, most of the heteronuclear experiments are performed in a 1H detected version, also called “inverse detection” (in contrast to the classical X nucleus detection described so far). If the proper equipment is available (re-wired spectrometer console; inverse detection probe!), then inverse detection offers such an immense gain in sensitivity that there is (almost) no reason to run any “conventional” heteronuclear correlation experiments anymore. Theoretical relative sensitivities (S/N) for H,X correlation spectra (X=13C, 15N)*. Method gexc. gdet.3/2 13C 15N direct detection gXgX 3/2 1.0 1.0 INEPT / DEPT gHgx 3/2 4.0 9.9 reverse INEPT gXgH 3/2 7.9 31.0 (relative to INEPT=1) 2.0 3.1 invers gHgH 31.6 306.0 (relative to INEPT=1) 7.9 31.0 * not taking into account other factors, e.g., T1, heteronucl. NOE, linewidths etc. It has to be remembered that the number of scans (~spectrometer time) required goes up with the square of the sensitivity ratio. Thus, a simple 1D 13C spectrum might well need almost 1000 times the measuring time of an inverse 2D 1H,13C-correlation!
The first H detected correlation experiment was performed in 1977 by Maudsley Ernst: just the basic 2D H, C three-pulse correlation experiment(antiphase crosspeaks in both dimensions!) reversed"to start onC and end on H ( the H irradiation boosts the C magnetization by the heteronuclear NOE) saturation The term"inverse'"is usually reserved for experiments that start on H and detect H, giving the maximum sensitivity. There are basically two inverse H, X correlation experiments, the hsQC and the hmQc sequence HSQC(Heteronuclear Single Quantum Correlation) Experiment. HILE C The HSQC experiment consits essentially of the elements (INEPT-t1-reverse INEPT-t2); the delay t is set to t=(4JCH)- With product operators, the transfer goes as follows(chemical shift is refocussed during 2t) 90°(1,S) t1 2 IzSy cos(Q2t1) 90°(1,S) IxSz cos(@2t1)-> Iy cos( S2t1) To select only 13C bound protons, a phase cycling scheme has to be used on the 13C 90 pulses.A 180 phase shift on one of these pulses will flip the sign of the detected term, e.g
72 The first 1H detected correlation experiment was performed in 1977 by Maudsley & Ernst: just the basic 2D H,C three-pulse correlation experiment (antiphase crosspeaks in both dimensions!) “reversed” to start on 13C and end on 1H (the 1H irradiation boosts the 13C magnetization by the heteronuclear NOE). The term “inverse” is usually reserved for experiments that start on 1H and detect 1H, giving the maximum sensitivity. There are basically two inverse 1H,X correlation experiments, the HSQC and the HMQC sequence. HSQC (Heteronuclear Single Quantum Correlation) Experiment. t t t t x y x x y 1 H 1 3 C decoupl. The HSQC experiment consits essentially of the elements (INEPT – t1 – reverse INEPT – t2 ); the delay t is set to t = (4JCH) -1 . With product operators, the transfer goes as follows (chemical shift is refocussed during 2t): 90°x D 90°(I, S) t1 Iz ¾¾® I-y ¾® 2 IxSz ¾¾¾® 2 IzSy ¾® 2 IzSy cos(Wt1 ) 90°(I, S) D ¾¾¾® 2 IxSz cos(Wt1 ) ¾® Iy cos(Wt1 ) To select only 13C bound protons, a phase cycling scheme has to be used on the 13C 90° pulses. A 180° phase shift on one of these pulses will flip the sign of the detected term, e.g.: