# Nitrogen-15 nuclear magnetic resonance spectroscopy

Nitrogen-15 nuclear magnetic resonance spectroscopy (nitrogen-15 NMR spectroscopy, or just simply $^{15}$N NMR) is a version of nuclear magnetic resonance spectroscopy that examines samples containing the $^{15}$N nucleus. $^{15}$N NMR differs in several ways from the more common $^{13}$C and $^1$H NMR. To circumvent the difficulties associated with measurement of the quadrupolar, spin-1 $^{14}$N nuclide, $^{15}$N NMR is employed in samples for detection since it has a ground-state spin of ½. Since$^{14}$N is 99.64% abundant, incorporation of $^{15}$N into samples often requires novel synthetic techniques.
Nitrogen-15 is frequently used in nuclear magnetic resonance spectroscopy (NMR), because unlike the more abundant nitrogen-14, that has an integer nuclear spin and thus a quadrupole moment, $^{15}$N has a fractional nuclear spin of one-half, which offers advantages for NMR like narrower line width. Proteins can be isotopically labeled by cultivating them in a medium containing nitrogen-15 as the only source of nitrogen. In addition, nitrogen-15 is used to label proteins in quantitative proteomics (e.g. SILAC).

## Implementation

$^{15}$N NMR has complications not encountered in $^1$H and $^{13}$C NMR spectroscopy. The 0.36% natural abundance of $^{15}$N results in a major sensitivity penalty. Sensitivity is made worse by its low gyromagnetic ratio (γ = −27.126 × 10$^6$ T$^{−1}$s$^{−1}$), which is 10.14% that of $^1$H. The signal-to-noise ratio for $^1$H is about 300-fold greater than $^{15}$N at the same magnetic field strength.

### Physical properties

The physical properties of $^{15}$N are quite different from other nuclei. Its properties along with several common nuclei are summarized in the below table.
| Isotope | Magnetic dipole moment (μ$_N$) | Nuclear spin number | Natural abundance (%) | Gyromagnetic ratio (10$^6$ rad s$^{−1}$ T$^{−1}$) | NMR frequency at 11.7T (MHz) |
| $^1$H | 2.79284734(3) | 1/2 | ~100 | 267.522 | -500 |
| $^2$H | 0.857438228(9) | 1 | 0.015 | 41.066 | -76.753 |
| $^3$H | 2.97896244(4) | 1/2 | 0 | 285.349 | -533.32 |
| $^{10}$B | 1.80064478(6) | 3 | 19.9 | 28.747 | -53.718 |
| $^{11}$B | 2.6886489 | 3/2 | 80.1 | 85.847 | -160.42 |
| $^{13}$C | 0.7024118(14) | 1/2 | 1.1 | 67.238 | -125.725 |
| $^{14}$N | 0.40376100(6) | 1 | 99.6 | 19.338 | -36.132 |
| $^{15}$N | -0.28318884(5) | 1/2 | 0.37 | -27.126 | 50.782 |
| $^{17}$O | -1.89379(9) | 5/2 | 0.04 | -36.281 | 67.782 |
| $^{19}$F | 2.628868(8) | 1/2 | ~100 | 251.815 | -470.47 |
| $^{31}$P | 1.13160(3) | 1/2 | ~100 | 108.394 | -202.606 |
From these data, one can see that at full enrichment, $^{15}$N is about one tenth (-27.126/267.522) as sensitive as $^1$H.

### Chemical shift trends

The International Union of Pure and Applied Chemistry (IUPAC) recommends using CH$_3$NO$_2$ as the experimental standard; however in practice many spectroscopists utilize pressurized NH$_3$(l) instead. For $^{15}$N, chemical shifts referenced with NH$_3$(l) are 380.5 ppm upfield from CH$_3$NO$_2$ (δ$_{NH3}$ = δ$_{CH3NO2}$ + 380.5 ppm). Chemical shifts for $^{15}$N are somewhat erratic but typically they span a range of -400 ppm to 1100 ppm with respect to CH$_3$NO$_2$. Below is a summary of $^{15}$N chemical shifts for common organic groups referenced with respect to NH$_3$, whose chemical shift is assigned 0 ppm.

### Gyromagnetic ratio

Unlike most nuclei, the gyromagnetic ratio for $^{15}$N is negative. With the spin precession phenomenon, the sign of γ determines the sense (clockwise vs counterclockwise) of precession. Most common nuclei have positive gyromagnetic ratios such as $^1$H and $^{13}$C.

## Applications
### Tautomerization

$^{15}$N NMR is used in a wide array of areas from biological to inorganic techniques. A famous application in organic synthesis is to utilize $^{15}$N to monitor tautomerization equilibria in heteroaromatics because of the dramatic change in $^{15}$N shifts between tautomers.

### Protein NMR

$^{15}$N NMR is also extremely valuable in protein NMR investigations. Most notably, the introduction of three-dimensional experiments with $^{15}$N lifts the ambiguity in $^{13}$C–$^{13}$C two-dimensional experiments. In solid-state nuclear magnetic resonance (ssNMR), for example, $^{15}$N is most commonly utilized in NCACX, NCOCX, and CANcoCX pulse sequences.

### Investigation of nitrogen-containing heterocycles

$^{15}$N NMR is the most effective method for investigation of structure of heterocycles with a high content of nitrogen atoms (tetrazoles, triazines and their annelated analogs). $^{15}$N labeling followed by analysis of $^{13}$C–$^{15}$N and $^1$H–$^{15}$N couplings may be used for establishing structures and chemical transformations of nitrogen heterocycles.

## INEPT

Insensitive nuclei enhanced by polarization transfer (INEPT) is a signal resolution enhancement method. Because $^{15}$N has a gyromagnetic ratio that is small in magnitude, the resolution is quite poor. A common pulse sequence which dramatically improves the resolution for $^{15}$N is INEPT. The INEPT is an elegant solution in most cases because it increases the Boltzmann polarization and lowers T$_1$ values (thus scans are shorter). Additionally, INEPT can accommodate negative gyromagnetic ratios, whereas the common nuclear Overhauser effect (NOE) cannot.