*3.2. Ammonium Ions*

Ammonium ions and alkyl ammonium ions play an important role in biology [19,20]. Attempts to mimic enzymes were carried out by Lehn et al. [21] An example is SC-24 (Figure 6). Other confinements have been investigated recently [22].

**Figure 6.** Structure of SC-24.

Arginine showed deuterium isotope effects at the η nitrogen, 0.307 ppm measured in a H2O/D2O mixture 1:1. Due to the technique used it is an average for the two Nη nitrogen obtained, which means that the measured isotope effect is close to one half of <sup>1</sup>ΔN(D). The values served as a reference for non-interacting residues in lysozyme [9].

Platzer et al. [23] measured deuterium isotope effects at the side-chain nitrogen of protonated and non-protonated lysines and argines being part of a tripeptide, acetyl-Gly-X-Glyamide. For the side-chain nitrogen of lysine, the values were 1.05 ppm for ND3 <sup>+</sup> and ~1.9 ppm for ND2. For arginines, the value for the ε nitrogen in the protonated state was 1.0 ppm and 1.4 ppm for the η nitrogen. In this case, it is not a pure one-bond isotope effect as long-range effects are present. In the case of lysines, the very different values between the protonated and non-protonated cases may be used to estimate the protonation state of lysines.

In Figure 7 is shown a plot of <sup>1</sup>ΔN(D)4 vs. the heavy atom distances of halide ammonium ion salts, I−, Br−, Cl− and F is shown−. Marked with squares are data points for SC-24 and water. It is seen, that as the distance decreases, the one-bond deuterium isotope effect decreases. [24] A similar trend is found from theoretical calculations. [25] Furthermore, it is found that water is more effective than the halide ions. For SC-24, the one-bond deuterium isotope effects were found to be independent of the counter ions [26].

**Figure 7.** A plot of <sup>1</sup>ΔN(D)4 vs. the heavy atom distances of halide ammonium ion salts. The second square is data for water. Data from Ref. [24].

The two-bond deuterium isotope effects for complexes with 18-crown-6, 18-crown-6(COOH)4, dicylohexano 18-crown-6, kryptofix's 2.2.2, 2.2.1 and 5, SC-24 and the ionophore and nonactin, are negative. The two-bond deuterium isotope effects are roughly proportional to the NH chemical shifts. The higher the NH chemical shifts, the more negative the two-bond deuterium isotope effects. In other words, the more negative the stronger the hydrogen bond. For SC-24, 18-crown-6(COOH)4 and nonactin, the two-bond deuterium isotope effects on 1H chemical shifts are independent of the counter ion. For the others, the more negative values are usually found for Cl− rather than for NO3 − or I− counter ions [27].

A plot of <sup>2</sup>ΔH(D) vs. <sup>1</sup>ΔN(D) shows very little correlation (Figure 8). It has been suggested that the 15N chemical shifts and with that the one-bond isotope effects depend on orbital overlap with the counter ion, whereas the two-bond deuterium isotope effects on 1H depend on electric field effects. In both cases, the isotope effect monitors ion pair formation.

**Figure 8.** A plot of <sup>2</sup>ΔH(D) vs. <sup>1</sup>ΔN(D). Data from Ref. [26].
