Template for crystallisation

Glass surfaces functionalised with alkylsilanes were selected as templates for crystallisation experiments. Silanisation is a well established technique for functionalisation of surface with hydrolysable functional groups (Cras et al., 1999). Glass or silica surfaces when cleaned and hydroxylated exhibit hydroxyl functional groups on the surface. The alkylsilanes with ethoxy or methoxy functional groups react with the hydroxyl groups on the surface and form siloxane bonds, thereby forming chemisorbed layer on such surfaces. This occurs through a two stage reaction. The alkoxy groups of silanes are first hydrolysed by surface water on hydrated glass, followed by the condensation of silanes on to glass surface. Thermal curing after silanisation ensures cross linking of silanes. The schematic representation of the functionalisation reaction and the final structure of cured silane layer are provided in Figure 3.3. Different methods for silanisation procedure on glass/ silica surface involving elevated or room temperature from organic, aqueous or vapour phase deposition has been reported (Halliwell & Cass, 2001). In ideal situation, each trialkoxysilane molecule can result in one or two attachments with the glass surface based on availability of surface hydroxyl groups and their spacing. Upon thermal treatment after surface attachment, the

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0.22 μm filter to avoid particulate impurities. The vials were hermetically sealed to avoid evaporation of solvent. Temperature was maintained at 15, 20 or 25 °C by regulating the temperature of the incubator in which the glass vials were placed for crystallisation.

The inner surface of glass vials form an interface with the crystallising solution and hence act as the template surface for crystallisation. Therefore, the whole inner surface of each vial was functionalised to avoid the effect of multiple surfaces or interfaces in contact with crystallising solution. Ideally, the solution would be in contact with only two interfaces – one with the functionalised vial surface and with the solvent saturated air above the solution in the sealed vial. Functionalised glass cover slips were also used as template surfaces in certain instances to facilitate removal of crystals while attached to surface. In the case of glass cover slips, the templates were placed at the bottom of vials or crystallisation dishes into which the supersaturated solution was poured. The functionalisation method used for both vials and cover slips are the same and is explained in next section.

3.4.2. Template functionalisation

Prior to silanisation, the glass surfaces were cleaned and pretreated to form hydroxyl surface groups to enhance silanisation efficiency. Cleaning involves repeated washing of vials or cover slips with de-ionised water (DI water) and ethanol to remove dust and other organic contaminants from the surface. Surfaces are further cleaned overnight with 5 N nitric acid. This removes any covalently bonded surface impurities and readies the template for hydration process. Acid content from the surface is removed by washing with copious amount of DI water, followed by hydration in boiling water for 4 – 6

hours. During hydration, the surface groups of glass are hydroxylated resulting in silanol (Si – OH) groups. Expression of hydroxyl content on template surface is essential for successful silanisation process (Cras et al., 1999). However, excess water content on surface can inhibit silanisation process and hence vials and cover slips are dried at room temperature after hydration process. All templates including control surfaces (clean glass cover slips) undergo these pretreatment steps while other templates are further functionalised through silanisation process. Liquid phase silanisation is carried out in toluene with 1% silane content. The silanisation solution is poured into the vials and left overnight at room temperature. For cover slips, they are dipped into a silanising solution inside Corning® silanisation jar. Individual slots for each cover slip ensure full surface exposure to solution. After silanisation, the solution is poured out and the vials/ cover slips are washed with toluene and ethanol to remove any excess silane from surface. Templates are then subjected to heat treatment at 80 – 100 °C for 20 – 24 hours to complete surface attachment and cross linking of silanes.

3.4.3. Solubility measurement

Solubilities of form II and form III CBZ crystals in ethanol were measured using the Crystalline system (Avantium, The Netherlands). The apparatus works on the principle of ‘clear point’ measurement, the temperature at which a suspension with a known concentration of solid turns into a clear solution upon heating. In the case of stable form III polymorph, the clear point temperatures of a slurry suspension of CBZ were determined at various heating rates. By extrapolating the data to zero heating rate, the actual dissolution temperature that is independent of the heating rate was determined.

Solubility of form II CBZ was determined using an adapted bracketing method. In this method, multiple vials with solution samples saturated with a known quantity of the stable crystal form were first prepared and held at 0.25 °C (±0.1 °C) temperature intervals. 5 mg of form II crystals was added to each of the equilibrated solutions. If the initial solution concentration was below the solubility of the metastable form at that particular temperature, the added crystals fully dissolved (typically within 2 – 3 minutes). On the other hand, if the solution concentration was above the solubility limit, the added crystals did not dissolve fully and the excess solid underwent solution mediated transformation to the stable form over a period of time. Among the multiple solution samples, the lowest temperature at which the added form II crystals dissolved completely corresponded to the saturation temperature with respect to the metastable form.