The thermodynamic stability of a cytosine(C)‐rich i‐motif tract of DNA, which features pH‐sensitive [C..H..C]+ moieties, has been studied as function of both pressure (0.1–200 MPa) and pH (3.7–6.2). Careful attention was paid to correcting citrate buffer pH for known variations that stem from changes in pressure. Once pH‐corrected, (i) at pH >4.6 the i‐motif becomes less stable as pressure is increased (KD decreases), giving a small negative volume change for dissociation (ΔDV°) of the i‐motif – a conclusion opposite to that which would be drawn if the buffer pH was not corrected for the effects of pressure; (ii) the i‐motif's melting temperature increases by more than 30 K between pH 6.5 and 4.5, the consequence of an enthalpy for dissociation (ΔDH°) of 77(3) and 90(3) kJ (mol H+)−1 at 0.1 and 200 MPa, respectively; (iii) below pH 4.6 at 0.1 MPa (pH 4.3 at 200 MPa) the melting temperature decreases as a result of double protonation of cytosine pairs, and ΔDH° and ΔDV° change signs; and (iv) the combination of ΔDH° and ΔDV° lead to the melting temperature at pH 4.3 being 3 K higher at 200 MPa than at 0.1 MPa.

Monitoring the spin states of species in solution is a crucial aspect of understanding magnetic properties as well as spin-labile sensing, supramolecular, catalytic and biochemical processes. Herein, we describe the first quantitative variable-pressure and variable-temperature method of determining spin states in solution, demonstrate that it is accurate, and identify a simultaneous T and P sensor system.

The relatively low chemical stability of cytosine compared with other nucleobases is a key concern in origin-of-life scenarios, but the effect of pressure on the rate of hydrolysis of cytosine to uracil remains unknown. Through in situ NMR spectroscopy measurements, it has been determined that the half-life of cytosine at 373.15 K decreases from (18.0±0.7) days at ambient pressure (0.1 MPa) to (8.64±0.18) days at high pressure (200 MPa). This yields an activation volume for hydrolysis of (-11.8±0.5) cm mol ; a decrease that is similar to the molar volume of water (18.0 cm mol ) and consistent with a tetrahedral 3,3-hydroxyamine transition-state/intermediate species. Similar behaviour was also observed for cytidine. At both ambient and high pressures, the half-life of cytosine decreases significantly as the pH decreases from 7.0 to 6.0. These results provide scant support for the notion that RNA-based life forms originated in high-temperature, high-pressure, acidic environments.