Molecular mechanism of high pressure action on lupanine

High hydrostatic pressure (HHP) is an emerging tool for studying conformational changes in lipids, proteins and nucleic acids. Although many thermodynamic parameters describing those processes have been determined, a molecular mechanism of HHP action is poorly recognized. To get insight into that, we have studied quinolizidine alkaloid, lupanine. It consists of two quinolizidine moieties, one of which contains lactam group. Using Fourier-transform infrared (FT-IR) spectroscopy, we have showed that at 6 kbar, the intensity of amide band at 1589 cm−1 decreased and a new band at 1556 cm−1 appeared. These changes are due to the hydrolysis of lupanine to lupanic acid. That reaction was confirmed with 13C NMR spectra of lupanine exposed to HHP. The NMR signals at 176.6 and 184.3 ppm were assigned to lactam group of lupanine and carboxylic group of lupanic acid, respectively. The ring opening reaction of lupanine under HHP is reversible at ambient pressure, as evidenced by CD measurements. A slightly acidic condition induced by HHP causes protonation of lactam group and carbocation is formed, while on the other hand, water molecule as nucleophile attacks electrophilic carbon of lactam and electrons move towards oxonium ion. Finally, CN bond breaks down and carboxyl group is formed.