Co-crystallization of N′-benzylidenepyridine-4-carbohydrazide and benzoic acid via autoxidation of benzaldehyde

The 1:1 co-crystal N′-[(2-methylphenyl)methylidene]pyridine-4-carbohydrazide–benzoic acid (1/1) formed unexpectedly after autoxidation of benzaldehyde during the slow evaporation process of a solution of isoniazid in benzaldehyde. The original intent of the synthesis was to modify isoniazid with benzaldehyde and crystallize the product in order to improve efficacy against Mycobacteria species, but benzoic acid formed spontaneously and co-crystallized with the intended product, N′-benzylidenepyridine-4-carbohydrazide.


Chemical context
Mycobacterial infections are a historic tribulation to mankind, and are managed with an array of drugs ranging from natural to synthetic derivatives that possess antimicrobial properties. However, these strategies have failed over time due to the emergence of resistant mycobacteria (Cully, 2014). A number of constituents such as isoniazid (INH) have been modified to try and curb the scourge of tuberculosis (Cully, 2014). Some of the resulting modified INH derivatives have been shown to render the active pharmaceutical ingredients (API) a lot more active to the circulating resistant strains of TB (Hearn & Cynamon, 2004;Suarez et al., 2009). It was for this reason that the covalent modification of API's was adopted to synthesize new analogues by modifying the NH 2 group of the hydrazide moiety of INH (Smith et al., 2015), believed to assist in the evasion of the N-arylaminoacetyl transferases, an enzyme capable of reducing the efficacy of INH in particular, by acetylating the NH 2 position, thus ultimately preventing its reaction with nicotinamide adenine dinucleotide (NADH) (Vishweshwar et al., 2006;Smith et al., 2015).
Benzaldehyde is known to undergo autoxidation resulting in the formation of benzoic acid. The formation of benzoic acid occurs when benzaldehyde is exposed to air at room temperature (293 K) where the rate of the reaction is increased by the presence of a catalyst. However, this phenomenon can occur spontaneously without a catalyst over a prolonged period (Sankar et al., 2014). The synthesis of this co-crystal was interesting as there were three separate processes that took place within the reaction mixture to create the final product. Firstly, benzaldehyde reacted with isoniazid to form N 0 -benzylidenepyridine-4-carbohydrazide. Secondly, excess benzaldehyde spontaneously autoxidized to form benzoic acid as described above (no benzoic acid was added to the reaction mixture). Lastly, the carbohydrazide moiety cocrystallized with the benzoic acid (as shown in Fig. 1) to form the product, N 0 -[(2-methylphenyl)methylidene]pyridine-4carbohydrazide-benzoic acid (1/1).

Structural commentary
The asymmetric unit contains one molecule of N 0 -benzylidenepyridine-4-carbohydrazide (C 13 H 11 N 3 O 1 ÁC 7 H 6 O 2 ) and one molecule of benzoic acid (as shown in Fig. 2). This cocrystal crystallizes in the Pbca space group. The benzoic acid molecule lies in the plane of the pyridine ring of the benzylidene derivative. All bond lengths and angles are normal.

Supramolecular features
Each carbohydrazide moiety is hydrogen bonded by a strong O2-H2Á Á ÁN2 hydrogen bond (Table 1) to a benzoic acid molecule to form a co-crystal. This interaction is supported by a weaker C-HÁ Á ÁO hydrogen bond that stabilizes the coplanar arrangement of the carboxylic acid moiety and the pyridine ring. The graph-set notation for this would be R 2 2 (7) (Bernstein et al., 1995), and is observed in other isoniazid cocrystals (Lemmerer et al., 2010) (Fig. 2). This co-crystal is another example of the robust carboxylic acidÁ Á Ápyridine heterosynthon (Shattock et al., 2008;Aakerö y et al., 2007). Each carbohydrazide moiety is also hydrogen bonded via its N1-H1 donor to the carbonyl oxygen (O1) acceptor of an adjacent carbohydrazide moiety. This results in a mono-periodic hydrogen-bonded chain along the b-axis direction, with graph-set notation C(4). Overall, the combined carbohydrazide moiety with the benzoic acid forms a ribbon motif (as shown in Fig. 3a). Viewed along the b-axis, the ribbons forms a X-shaped motif seen in other carbohydrazide moieties (Hean et al. 2018) (Fig. 3b).

Synthesis and crystallization
All reagents were commercially sourced and used without further purification. 1.00 g of isonicotinic acid hydrazide (isoniazid) (7.29 mmol) were dissolved in 15 ml of benzaldehyde in a 50 ml amber Schott bottle. The mixture was placed on a stirring heating block and heated to 333 K while stirring with a magnetic stirrer bar. Once the isoniazid had completely dissolved, the lid was tightly sealed. The solution was then allowed to react for 24 h. To maintain the temperature throughout the duration of the experiment, the amber Schott bottle was covered with an inverted round glass evaporation dish. After 24 h, the solution was allowed to cool to ambient temperature. The stirrer bar was retrieved and the sample was left to evaporate slowly for 6 weeks at ambient temperature without a lid. Over the 6 weeks, the temperature in the laboratory fluctuated between 298 and 300 K. Due to the fact that benzaldehyde evaporates extremely slowly, the Schott bottle was placed in the laminar flow biohazard safety level 2 cabinet to facilitate evaporation. Crystals (colourless blocks) started forming on the rim on the outside of the bottle as the benzaldehyde evaporated. One of these crystals was sampled for XRD analysis.

Refinement
Crystal data, data collection and structure refinement details are summarized in Table 2. C-bound H atoms were first located in the difference map, then positioned geometrically and allowed to ride on their respective parent atoms, with thermal displacement parameters 1.2 times of the parent C atom. The coordinates and isotropic displacement parameters of the O and N-bound H atoms involved in hydrogen-bonding interactions (H1 and H2) were allowed to refine freely.

Special details
Experimental. Absorption corrections were made using the program SADABS (Sheldrick, 1996) Geometry. All esds (except the esd in the dihedral angle between two l.s. planes) are estimated using the full covariance matrix. The cell esds are taken into account individually in the estimation of esds in distances, angles and torsion angles; correlations between esds in cell parameters are only used when they are defined by crystal symmetry. An approximate (isotropic) treatment of cell esds is used for estimating esds involving l.s. planes. Refinement. The crystal structure was solved through direct methods using SHELXT. Non-hydrogen atoms were first refined isotropically followed by anisotropic refinement by full matrix least-squares calculations based on F 2 using SHELXL.