Figure 6: Pharmacophore of 2-APB and ¹H-NMR (¹H-nuclear magnetic resonance) spectra reveal structural changes of 2-APB, DPBA and DPB by acidic pH.

(A) Structures of 2-APB, DPBA, diphenyl methanol (DPM), and diphenyl borate (DPB). (B) Whole-cell TRPV3 currents at −60 mV showing that the response to 10 μM DPB was potentiated by pH 5.5 after sensitization by 100 μM 2-APB. (C) Whole-cell TRPV3 currents at −60 mV showing the lack of response to DPM. TRPV3 channels were first sensitized by repeated applications of 100 μM 2-APB and the responses to 10 μM 2-APB were notably enhanced by pH 5.5. However, no detectable activity was evoked by 30 μM or 500 µM DPM, or the combination of 30 μM DPM and pH 5.5. At the end, 2-APB (100 μM) was applied to ensure that channels stilled functioned normally. (D) Summary of relative currents elicited by various combinations of compounds and pH conditions as indicated. The pH was 7.4 if not mentioned. Error bars represent SEM. (E) ¹H-NMR spectra of 2-APB and its analogs under various pH environments. For 2-APB, DPBA, and DPB, the signals in the aromatic area were obviously shifted downfield at pH 5.5 as compared to pH 7.4 and 8.5. The intensities for the signals in aliphatic areas also differ between pH 5.5 and pH 7.4 or 8.5. These imply that the compounds exist in different configurations at different pH conditions. However, for DPM, there was only one configuration in solutions of various pH because of the near identical NMR signals. NMR experiments were performed at least twice with similar results. Upper panels show full spectra; lower panels show expanded spectra for the aromatic areas.