NMR investigation of magnesium chelation and cation-induced signal shift effect of testosterone.

We have previously reported that testosterone (Tes) is able to interact with magnesium chloride dissolved in methanol. In this study, we have applied 1H and 13C NMR spectroscopies to a series of Tes solutions containing Mg2+ at various concentrations. High-resolution 13C NMR spectra of Tes/Mg2+ revealed well-resolved 13C signals, and the intensities of those arising from C3, C5, C16, and C17 decreased linearly with increasing Mg2+ concentration. The magnitude of the chelation affinity could be deduced from the slopes of the 13C intensity variations; typically, the greater the slope the higher the chelation affinity. The results revealed Tes/Mg2+ chelation to be mediated by the oxygen atom attached to C3 in ring A, and the hydroxyl group attached to C17 in ring D. With regard to the chelation specificity, we showed that Tes chelates Mg2+, but not Ca2+ or Zn2+. We also explored the cation-induced signal shift effects of Tes in the presence of Mg2+, Ca2+, or Zn2+. We demonstrate that high-resolution 13C NMR spectroscopy provides a better probe than 1H NMR for the detection of cation chelation and cation-induced signal shift effects for steroid compounds such as Tes.