Virtual screening of co-formers for ketoprofen co-crystallization and the molecular properties of the co-crystal

S. Siswandi, Taofik Rusdiana, Jutti Levita Department of Pharmaceutics and Formulation Technology Faculty of Pharmacy, Universitas Padjadjaran Jl. Raya Bandung-Sumedang km.21 Jatinangor Sumedang, Indonesia. Indonesian Airforce Pharmaceutical Institution Jl.Abdurachman Saleh, Lanud Husein Sastranegara, Bandung, Indonesia. Department of Pharmaceutical Analysis and Medicinal Chemistry Faculty of Pharmacy, Universitas Padjadjaran Jl. Raya Bandung-Sumedang km.21 Jatinangor Sumedang, Indonesia.


INTRODUCTION
Ketoprofen (Fig. 1) or [2-(3-benzoylphenyl)propionic acid] is a nonsteroidal antiinflammatory and analgesic agent.The positive qualities of ketoprofen are based on optimal physicochemical and structural characteristics, its ability to penetrate into and accumulate in the inflammation centers and compatibility with other classes of drugs (Khormosh, 2009).Ketoprofen is soluble in ethanol, chloroform, and ether.It is practically insoluble in water, therefore as most of NSAID drugs, this compound is categorized as Biopharmaceutics Classification System (BCS) class II, i.e. drugs with low solubility and high permeability character (Hussain, 2002).Various techniques have been widely used to enhance the solubility of poorly water soluble drugs; co-crystallization is one amongst them.A co-crystal is a multi-component crystal which all components are solid at room temperature in a stoichiometric ratio and it involves non-covalent interactions such as hydrogen bonds, van-der Waals bonds, ionic bonds in a crystal lattice (Shan, 2008;Patel, 2012).The selection of co-formers is based on their ability to form reversible or non-covalent interaction with the API.Both API and co-former should contain hydrogen bond donors (HBDs) or acceptors (HBAs), e.g.ether, thioether, alcohol, ketone, thioketone, ester, thioester, carboxylic acid, amide, primary and secondary amines, etc (Vinesha, 2013).Other interactions involved in the formation of co-crystals are Van der Waals force, pi-pi (π-π) interaction, aromatic stacking, and halogen bonding (He, 2008).

Molecular modelling
2D structures of ketoprofen (ChemSpider ID: 3693) and its co-formers in .molformat were downloaded from www.chemspider.com.All molecules were geometry optimized using molecular mechanics method in Portable_Hyperchem_8.0.and calculated their QSAR properties using the same program.

Molecular docking
All .molfiles of the molecules were converted into .pdbfiles by employing OpenBabelGUI 2.2.3.The files then were opened in AutoDock 4.2.3 and converted into .pdbqfiles by adding polar hydrogen and Kollman charges.The .pdbq files were converted into .pdbqtby calculating their torsion angles and were ready to be used for docking.The dimension of grid box was set to 40x40x40 Å 3 and other parameters were set to default.Docking was repeated 10 times for each co-former.Parameters observed were type and energy (Ei) of interactions.

Solubility assay
Instruments used were mechanical agitator (Kotterman 4010), Shimadzu UV 1700 spectrophotometer, and chemical glasswares.Solubility assay was carried out according to Higuchi and Connor method on ketoprofen and all co-crystals formed.An accurately weighed 100 mg of ketoprofen was dissolved to saturation in water and was shaken vigorously using mechanical agitator for 24h 120 rpm.The mixture was filtered and passed through milipore membrane ø 25 µm.Filtrate was measured at 250-260 nm.

Dissolution test
The best co-former according to virtual screening and solubility assay is equimol mixed and ground with ketoprofen, and 5 mL of methanol is added drop wise to the mixture.
The crystal formed is kept at room temperature (27 o C) for 24 h.Dissolution test is performed to both ketoprofen and ketoprofen co-crystal at 10 minutes interval for 1 h.

RESULTS AND DISCUSSION
The molecule of ketoprofen contains two aromatic rings as well as 1 HBD (hydrogen atom of the carboxylate) and 3 HBAs (oxygen atoms of the carbonyl and carboxylate), hence it is possible to form co-crystals with certain co-formers.Co-formers chosen in this work were benzoic acid, fumaric acid, malonic acid, citric acid, stearic acid, succinic acid, saccharin, and methyl paraben.The result of virtual screening of co-formers using molecular docking is showed in Table 1.
Mixtures of ketoprofen with the co-formers were tested for their solubilities and compared with ketoprofen itself.Results could be seen in Table 2.
Based on solubility assay carried out using Higuchi and Connor method, ketoprofen is slightly soluble in water (117.18μg/mL).By mixing it with citric acid, the solubility increases 142.67% (167.18μg/mL), whereas with benzoic acid and saccharin the increasing of solubility are 162.00%(189.84 μg/mL) and 256.54% (300.62 μg/mL), respectively.The interactions of ketoprofen with citric acid, benzoic acid, and saccharin could be proposed and seen in Fig. 2.

Table 1 .
Virtual screening of co-formers using molecular docking Co