The COSY experiment

The COSY experiment is the most familiar to 2D NMR spectroscopists. The cross peaks connect protons which are coupled by scalar interactions. Under these 

When the second 90° pulse is applied, the antiphase magnetization of the / and J spins is interchanged (Fig. 8.14B). During t2, the new antiphase coherence, 

To a good approximation, p is the transverse relaxation rate of spin 7 and p is the transverse relaxation rate of spin J. The two cross peaks are distinguished according to which of the two T [ or constants appears in each dimension. From Eq. (8.3), it appears that the maximum information is not contained in the first data points. In fact, considering that Eq. (8.3) represents the evolution of the signal, the derivative of 7 (t) with respect to t (or t2) provides the value of t (or t2) with maximum intensity of the signal  

The coherence transfer provides cross peaks which are antiphase for the various 7//-split components. The antiphase nature of the cross peaks then leads to partial or total cancellation of the cross peaks themselves, especially if they are phased in the absorption mode. This behavior can be simulated (Fig. 8.15) using appropriate treatments of the time evolution of the spin system, for instance using the density matrix formalism [17,18]. It is quite common that signals in paramagnetic systems 

In macromolecular paramagnetic systems, further phenomena may concur which provide COSY cross peaks, when using the sequence of Fig. 8.2C, whose nature is actually dipolar (see Section 8.8). 

Weaker cross peaks are observed between (i) methyl S and protons A, C, and D, (ii) Hq and protons G and R, (iii) Hp and H, and (iv) He and Hr. Since Ha is attached to an sp -hybridized carbon (5), and methyl S, from its chemical shift, might be likewise (Section 3-2a), couplings between Ha and its vicinal partner (Ho) to methyl S (allylic and homoallylic, respectively. Section 4-6) are not unreasonable. Moreover, protons C, M, and R 

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Fiffire 5.38 Pulse sequence for delayed COSY—a modification of the COSY experiment. The fixed delays at the end of the evolution period t and before the acquisition period <2 allow the detection of long-range couplings between protons. 

Fi and F. The off-diagonal peaks (cross-peaks) represent the direct coupling interactions between protons. Working through cross-peaks, one can easily correlate protons that are coupled to each other. Several versions of the COSY experiment have been designed to get optimum performance in a variety of situations (such as DQF COSY, COSY-45°, and COSY-60°). 

SECSY (spin-echo correlated spectroscopy) is a modified form of the COSY experiment. The difference in the pulse sequence of the SECSY experiment is that the acquisition is delayed by time mixing pulse, while the mixing pulse in the SECSY sequence is placed in the middle of the period. The information content of the resulting SECSY spectrum is essentially the same as that in COSY, but the mode... 

The most important difference in the spectrum as compared with RGs released by RG-hydrolase action (Colquhoun et al., 1990) was a doublet at 5 5.81 (J = 3.4 Hz). From the COSY experiments this doublet was found to belong to a four-proton spin-coupling network... 

In this case we pulse at the beginning of the evolution time and then wait before doing our acquisition pulse. If we vary this wait by incrementing it for each successive cycle, we can change what we see in the FID. This is what generates our second dimension. In the case of the COSY experiment, we allow the coupling information to evolve during this period and then read what has happened to it with the acquisition pulse. 

As with the COSY experiment, the sequence starts with a pulse followed by an evolution period, but now the mechanism that couples the two spins (which must be in close proximity, typically <6 A) is the Nuclear Overhauser Effect (NOE). The second pulse converts magnetization into population disturbances, and cross-relaxation is allowed during the mixing time. Finally, the third pulse transfers the spins back to the x-y-plane, where detection takes place. The spectrum will resemble a COSY spectrum, but the off-diagonal peaks now indicate through-space rather than through-bond interactions. 

There are several variations of correlation spectroscopy, all giving rise to different, complementary data. We have already met HH (homo-nuclear) COSY earlier in this chapter (Section 4.5). An obvious extension to the COSY experiment is to use it for heteronuclear correlation, e.g. correlation of all the H and C signals in a molecule... 

An advantage of 2D TOCSY is that the net coherence transfer produced can be arranged to create pure positive absorption spectra, including the diagonal peaks, rather than spectra with equal positive and negative intensities obtained with differential coherence transfer as in the COSY experiment. 

The first term represents inverted z magnetization, the starting point for a transient NOE experiment. All we have to do is add a mixing period (simple delay rm) and a 90° read pulse, and we have a 2D NOE experiment NOESY. For the COSY experiment, we start... 

The first term, which is not observable in the COSY experiment, is now exactly what we need for a transient NOE experiment. We have inverted the Ha magnetization in a way that carries the information of its chemical shift encoded in the cos( 2a t ) term. Depending on the value of t, sometimes Ha will be completely inverted (cosine = 1), leading to a maximum NOE transfer to Hb, and sometimes it will not be inverted at all (cosine = — 1), leading to no NOE transfer to Hb. Thus, the transferred magnetization will also carry the chemical shift information of Ha ... 

A simple variant of the COSY experiment is COSY-35 (sometimes called COSY-45), in which the second 90° pulse is reduced from a 90° pulse to a 35° or 45° pulse (Fig. B.3). The result is that the fine structure of crosspeaks is simplified, with half the number of peaks within the crosspeak. This makes it much easier to sort out the coupling patterns in both dimensions and to measure couplings (active and passive) from the crosspeak fine structure. A more important variant of the COSY experiment is the DQF (double-quantum filtered)— COSY (Fig. B.4), which adds a short delay and a third 90° pulse. The INEPT transfer is divided into two steps antiphase I spin SQC to I,S DQC, and I,S DQC to antiphase S spin SQC. The filter enforces the DQC state during the short delay between the second and third pulses either by phase cycling or with gradients. DQF-COSY spectra have better phase characteristics and weaker diagonal peaks than a simple COSY, so this has become the standard COSY experiment. 

TOCSY (total correlation spectroscopy) is an extension of the COSY experiment, in which the coherence transfer is not limited to a single jump from one proton to another via a J coupling. Instead, coherence is spread out over an entire spin system of coupled protons via multiple /-coupled jumps. For example, in a string of carbons CHa-CHb-CHc-CHd, coherence can be transferred by the TOCSY mixing sequence from Ha to Hc or from Ha to Hj. Thus, crosspeaks will be observed at F% = va and I = i b, vc or (Fig. B.5). 

For example, the COSY experiment illustrated in Fig. 10.2 can be repeated with the first pulse applied in the same direction but with Bj for the second pulse applied along y, rather than x. M is then rotated into the y z plane, where its projection along y, M sin O.,, is detected. The signals for the two experiments are, then,... 

Today, a number of one- and two-dimensional NMR experiments are available for the detection of homonuclear Li, Li and Li, Li couplings. Aside from the COSY experiment, the double quantum filtered COSY (COSY-DQF), the TOCSY, and the ID and 2D INADEQUATE experiments [24] have been successfully employed. An attractive feature of all these experiments is their sensitivity for small scalar interactions which give rise to crosspeaks even if line splittings in the corresponding ID spectra are not resolved. This was first demonstrated with COSY experiments for a paramagnetic nickel complex [82] and for quadrupolar nuclei in the case of boron-11 [83]. 

In the present context, the COSY experiment is best performed with a pulse sequence (i), which enhances small coupling and reduces diagonal signals [84]. 

Figure 6-2 The pulse sequence for the COSY experiment. For (c ), the magnetization M is allowed to evolve a longer time from (b) than for (c) before the final 90 pulse is applied to give (d ).

Figure 6-4 (a) The result of the COSY experiment after double Fourier transformation for a single isolated nucleus such as that in Figure 6-3. (b) The result of the COSY experiment for two uncoupled nuclei. 

Figure 6-6 The stacked representation of the COSY experiment for the two coupled nuclei of p-chloroacetic acid. (Reproduced from A. E. Derome, Modern NMR Techniques for Chemistry Research, Pergamon Press, Oxford, UK, 1987, p. 189.)...

Figure 6-8 The COSY experiment for the illustrated annulene. (Reproduced from R. Benn and H. Gunther, Angew. Chem., Jnt. Ed. Engl, 22, 350(19831).

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