14.19: \(^{13}C\) NMR Spectroscopy

The ¹²C isotope of carbon - which accounts for up about 99% of the carbons in organic molecules - does not have a nuclear magnetic moment, and thus is NMR-inactive. Fortunately for organic chemists, however, the ¹³C isotope, which accounts for most of the remaining 1% of carbon atoms in nature, has a magnetic moment just like protons. Most of what we have learned about ¹H-NMR spectroscopy also applies to ¹³C-NMR, although there are several important differences.

5.6A: The basics of ¹³C-NMR spectroscopy

The magnetic moment of a ¹³C nucleus is much weaker than that of a proton, meaning that NMR signals from ¹³C nuclei are inherently much weaker than proton signals. This, combined with the low natural abundance of ¹³C, means that it is much more difficult to observe carbon signals: more sample is required, and often the data from hundreds of scans must be averaged in order to bring the signal-to-noise ratio down to acceptable levels. Unlike ¹H-NMR signals, the area under a ¹³C-NMR signal cannot be used to determine the number of carbons to which it corresponds. This is because the signals for some types of carbons are inherently weaker than for other types – peaks corresponding to carbonyl carbons, for example, are much smaller than those for methyl or methylene (CH₂) peaks. Peak integration is generally not useful in ¹³C-NMR spectroscopy, except when investigating molecules that have been enriched with ¹³C isotope (see section 5.6B).

The resonance frequencies of ¹³C nuclei are lower than those of protons in the same applied field - in a 7.05 Tesla instrument, protons resonate at about 300 MHz, while carbons resonate at about 75 MHz. This is fortunate, as it allows us to look at ¹³C signals using a completely separate 'window' of radio frequencies. Just like in ¹H-NMR, the standard used in ¹³C-NMR experiments to define the 0 ppm point is tetramethylsilane (TMS), although of course in ¹³C-NMR it is the signal from the four equivalent carbons in TMS that serves as the standard. Chemical shifts for ¹³C nuclei in organic molecules are spread out over a much wider range than for protons – up to 200 ppm for ¹³C compared to 12 ppm for protons (see Table 3 for a list of typical ¹³C-NMR chemical shifts). This is also fortunate, because it means that the signal from each carbon in a compound can almost always be seen as a distinct peak, without the overlapping that often plagues ¹H-NMR spectra. The chemical shift of a ¹³C nucleus is influenced by essentially the same factors that influence a proton's chemical shift: bonds to electronegative atoms and diamagnetic anisotropy effects tend to shift signals downfield (higher resonance frequency). In addition, sp² hybridization results in a large downfield shift. The ¹³C-NMR signals for carbonyl carbons are generally the furthest downfield (170-220 ppm), due to both sp² hybridization and to the double bond to oxygen.

Example 5.11

How many sets of non-equivalent carbons are there in each of the molecules shown in exercise 5.1?

Solution

Example 5.12

How many sets of non-equivalent carbons are there in:

1. toluene
2. 2-pentanone
3. para-xylene
4. triclosan

(all structures are shown earlier in this chapter)

Solution

Because of the low natural abundance of ¹³C nuclei, it is very unlikely to find two ¹³C atoms near each other in the same molecule, and thus we do not see spin-spin coupling between neighboring carbons in a ¹³C-NMR spectrum. There is, however, heteronuclear coupling between ¹³C carbons and the hydrogens to which they are bound. Carbon-proton coupling constants are very large, on the order of 100 – 250 Hz. For clarity, chemists generally use a technique called broadband decoupling, which essentially 'turns off' C-H coupling, resulting in a spectrum in which all carbon signals are singlets. Below is the proton-decoupled¹³C-NMR spectrum of ethyl acetate, showing the expected four signals, one for each of the carbons.

While broadband decoupling results in a much simpler spectrum, useful information about the presence of neighboring protons is lost. However, another modern NMR technique called DEPT (Distortionless Enhancement by Polarization Transfer) allows us to determine how many hydrogens are bound to each carbon. For example, a DEPT experiment tells us that the signal at 171 ppm in the ethyl acetate spectrum is a quaternary carbon (no hydrogens bound, in this case a carbonyl carbon), that the 61 ppm signal is from a methylene (CH₂) carbon, and that the 21 ppm and 14 ppm signals are both methyl (CH₃) carbons. The details of the DEPT experiment are beyond the scope of this text, but DEPT information will often be provided along with ¹³C spectral data in examples and problems.

Below are two more examples of ¹³C NMR spectra of simple organic molecules, along with DEPT information.

Example 5.13

Give peak assignments for the ¹³C-NMR spectrum of methyl methacrylate, shown above.

Solution

One of the greatest advantages of ¹³C-NMR compared to ¹H-NMR is the breadth of the spectrum - recall that carbons resonate from 0-220 ppm relative to the TMS standard, as opposed to only 0-12 ppm for protons. Because of this, ¹³C signals rarely overlap, and we can almost always distinguish separate peaks for each carbon, even in a relatively large compound containing carbons in very similar environments. In the proton spectrum of 1-heptanol, for example, only the signals for the alcohol proton (H_a) and the two protons on the adjacent carbon (H_b) are easily analyzed. The other proton signals overlap, making analysis difficult.

In the ¹³C spectrum of the same molecule, however, we can easily distinguish each carbon signal, and we know from this data that our sample has seven non-equivalent carbons. (Notice also that, as we would expect, the chemical shifts of the carbons get progressively smaller as they get farther away from the deshielding oxygen.)

This property of ¹³C-NMR makes it very helpful in the elucidation of larger, more complex structures.

Example 5.14

¹³C-NMR (and DEPT) data for some common biomolecules are shown below (data is from the Aldrich Library of ¹H and ¹³C NMR). Match the NMR data to the correct structure, and make complete peak assignments.

• spectrum a: 168.10 ppm (C), 159.91 ppm (C), 144.05 ppm (CH), 95.79 ppm (CH)
• spectrum b: 207.85 ppm (C), 172.69 ppm (C), 29.29 ppm (CH₃)
• spectrum c: 178.54 ppm (C), 53.25 ppm (CH), 18.95 ppm (CH₃)
• spectrum d: 183.81 ppm (C), 182. 63 ppm (C), 73.06 ppm (CH), 45.35 ppm (CH₂)

Solution

5.6B: \(^{13}C\)-NMR in isotopic labeling studies

Although only about 1 out of 100 carbon atoms in a naturally occurring organic molecule is the ¹³C isotope, chemists are often able to synthesize molecules that are artificially enriched in ¹³C at specific carbon positions, sometimes to the point where the ¹³C isotope is incorporated at a given position in over half of the molecules in the sample. This can be very useful, especially in biochemical studies, because it allows us to 'label' one or more carbons in a small precursor molecule and then trace the presence of the ¹³C label through a metabolic pathway all the way to a larger biomolecule product. For example, scientists were able to grow bacteria in a medium in which the only source of carbon was acetate enriched in ¹³C at the C₁ (carbonyl) position. When they isolated a large molecule called amino-bacterio-hopanetriol (very similar in structure to cholesterol) from these bacteria and looked at its ¹³C-NMR spectrum, they observed that the ¹³C label from acetate had been incorporated at eight specific positions. They knew this because the ¹³C-NMR signals for these carbons were much stronger compared to the same signals in a control (unlabeled) molecule.

This result was very surprising - the scientists had expected a completely different pattern of ¹³C incorporation based on what they believed to be the metabolic pathway involved. This unexpected result led eventually to the discovery that bacteria synthesize these types of molecules in a very different way than yeasts, plants, and animals (Eur. J. Biochem. 1988, 175, 405). The newly discovered bacterial metabolic pathway is currently a key target for the development of new antibiotic and antimalaria drugs.

Organic Chemistry With a Biological Emphasis by Tim Soderberg (University of Minnesota, Morris)