*3.3. Hirshfeld Surface Analysis*

Hirshfeld surface analysis was performed [9,35,36] in order to gain a better understanding of the differences in hydrogen bond networks and the contribution of weaker contacts within these conformational polymorphs. Since the Hirshfeld surface depends on the spherical atomic electron densities of a particular molecule within its crystal structure, surface differ even between polymorphs [29]. Examination of the Hirshfeld surfaces in Figure 6 shows the distinguishable features of each form, particularly the position and intensity of some of the red areas, which represent close contacts. The Hirshfeld surface of form I is characterized by more red spots relative to that of form II, which as remarked earlier through the full interaction maps, has a folding feature that hinders its accessibility to hydrogen bonds.

The nature of all non-bonding contacts in both polymorphs is presented through the fingerprint plots in Figure 7, by plotting *di* against *de* and hence translating the information provided by the Hirshfeld surface into a 2D format. The plots are overall strongly related, probably due to the similarities between these conformational polymorphs. The greatest proportion of interactions H ... H are with a higher percentage contribution (Figure 8) and smaller minimum distance *di* ≈ *de* ≈ 1.1 Å associated with form II. Although such contacts are believed to be repulsive in nature, Matta et al. demonstrated how the net result of their presence has a stabilizing effect of up to a 10 kcal/mol decrease in the total energy of that particular structure [37].

**Figure 6.** Hirshfeld surfaces with *dnorm* as mapped function, constructed for (**a**) form I and (**b**) form II, with neighboring molecules to illustrate the network of non-bonding contacts, represented by dotted lines (not only hydrogen bonds).

**Figure 7.** Fingerprint plots of (**a**) form I and (**b**) form II. The arrows represent interactions between different atoms, as indicated in the legend below. Given that the plots are approximately symmetrical, the arrows can be mirrored through the x–y diagonal.

**Figure 8.** Bar-chart representing the percentage contribution of the main contacts in the Hirshfeld surfaces of forms I and II. The category of "other" includes C . . . N and C . . . O interactions.

The spikes at the bottom left of both plots are very prominent, exhibiting the dominance of N ... H and O ... H contacts, due to the highly polar functional groups in ganciclovir. The presence of these contacts is in agreemen<sup>t</sup> with the full interaction maps and the hydrogen bonds predicted by *Mercury.* The greener shades on the spikes of the form I surface plot indicate a higher frequency of contacts having relatively shorter distances. The percentage contribution of O ... H interactions is equivalent for both forms, but the global minimum distance *di* + *de* of around 1.7 Å pertains to form I.

The C–H ... π contribution is evident in both polymorphs, as shown in the Hirshfeld surface maps; however, it is more significant in form II (Figure 8). Although these interactions are weaker and less directional than the conventional hydrogen bonds, they still contribute to self-assembly and molecular recognition processes, as they reinforce the stability within the supramolecular structure [38]. On the other hand, the absence of large maxima around the area at *di* + *de* ≈ 3.8 Å and of the typical bold red and blue pairs of triangles on the shape index surfaces, indicate the less prominent π ... π contacts [39]. Examination of the network of interactions in the two forms, reveals how the side chain

prevents close direct stacking of the aromatic systems, hence minimizing the effect of π ... π contacts. Any C ... C contact seems to be distant and concentrated on the periphery of the guanine ring system, which hints that the aromatic systems are either far apart and/or shifted relative to one another.
