Modulating The Solubility of Mesalamine by Means of Cocrystals

One of the major problems in drug discovery and development is the poor solubility of new chemical entities (NCEs). Compounds with low solubility not only complicate the drug development process but also hinder the effectiveness of in vitro and in vivo assays during drug discovery. Successful absorption of drug requires the drug to be dissolved in gastrointestinal fluids, and dissolution depends on the drug's solubility in the surrounding medium. Advancements in pharmaceutical sciences have led to the development of various strategies to address the problem of low aqueous solubility. Methods such as micronization, salt formation, cosolvency, micellar solubilization, and cyclodextrin complexation have been employed to enhance solubility and dissolution. However, the effectiveness of these techniques largely depends on the specific physicochemical properties of the molecules involved. Solubility is defined as the quantity of solute that can be dissolved in a saturated solution at a specific temperature. Crystal engineering  designs drug delivery systems by improving dissolution properties without compromising stability. It focuses on understanding and applying non-covalent interactions, like hydrogen bonding and π-π interactions, to form solid structures with desired characteristics, helping to address challenges such as poor aqueous solubility and enhance bioavailability. Desiraju expanded this concept to define “supramolecular synthons,” which are structural units within supermolecules formed by intermolecular interactions, also referred to as “cocrystals”. These supramolecular synthons can be formed by various groups, including acids (e.g., carboxylic, sulfonic, phosphonic, and boronic), primary and secondary amides, alcohols, amino-pyridines, ketones, aldehydes, ethers, esters, and amines. Supramolecular synthons are categorized into two types:

(a) Supramolecular homosynthons: Composed of identical self-complementary functional groups.

(b) Supramolecular heterosynthons: Composed of different but complementary functional groups.

Crystal engineering, in general, involves designing molecular solids to customize their physical and chemical properties, which can be used to control the solubility and dissolution rates of crystalline substances. Cocrystallization, provides a more flexible approach for engineering drug molecules. A cocrystallization can be applied to any active pharmaceutical ingredient (API), regardless of whether it contains ionizable groups. Pharmaceutical cocrystals offer advantages such as enhanced solubility, dissolution rate, physical and chemical stability, and improved bioavailability of the API. Additionally, the availability of many coformers listed in the GRAS (Generally Recognized as Safe) and EAFUS (Everything Added to Food in the US) databases provides opportunities for patenting and extending the life cycle of existing APIs. “Cocrystals are typically defined as crystalline substances consisting of at least two components: an API (neutral or ionic) and a neutral coformer, bound together through non-covalent, reversible interactions”. A pharmaceutical cocrystal requires that at least one of these components is a drug, and the other is a pharmaceutically acceptable solid coformer. Rapid onset and improved bioavailability are desirable for analgesics. Hence, there is strong scientific and clinical need to prepare novel forms of mesalamine possessing modified solubility and dissolution rate which can be formulated for oral administration.  Hence, it was selected for solubility enhancement via cocrystallization.

4. Materials and Methods

Table 1: Drug and Coformers

4.2 Equipment

Table 2: List of Equipment used

4.3 Liquid Assisted Grinding Method for preparation of cocrystal

Mesalamine and coformers at 1:1 stoichiometric ratio was grounded in acetonitrile for 10 minutes using a mortar and pestle and left it for solvent evaporation to obtain free flowing cocrystal.

Table 3: drug and cocrystal quantity in formulation of cocrystals

4.4 Formulation of pure mesalamine tablet

Composition for the formulation of pure mesalamine tablets were presented in table. The accurately weighed ingredients were mixed and were directly compressed to form tablet.

Table 4: Formulation of pure mesalamine tablet

4.5 Formulation of mesalamine cocrystal tablet

Composition for the formulation of pure mesalamine: Saccharin sodium (cocrystal with highest solubility) cocrystal tablets were presented in table.

Table 5: Formulation of mesalamine cocrystal tablet

4.6 Analysis of solubility and evaluation of cocrystals:

4.6.1 Analytical Method Development

The UV method was developed in water and phosphate buffer pH 6.8. The mesalamine solution was scanned in the 200-400 nm range in order to obtain the absorption spectrum.

Calibration Curve of Mesalamine in Distilled Water

 The standard stock solution of mesalamine was prepared in distilled water and phosphate buffer pH6.8 to produce the concentration of 100 μg/ml by dissolving accurately weighed quantity of drug.

Preparation of Working Solutions

Series of working solutions were prepared in the range 10-100 μg/ml by appropriate dilutions of the standard stock solution.

Construction of Calibration Curve

The calibration curve was created by recording the absorbance of the working solutions and plotting the absorbance against concentration.

4.6.2 Saturated solubility study

The solubility was determined by dissolving excess quantity of pure drug and cocrystal in the 10 ml vials containing water. The vials were subjected to agitation on rotary shaker for 6 hrs and allowed to stand for equilibration. The samples were filtered after 1 hr using Whatman filter paper, diluted with distilled water and analysed by UV Spectrophotometer at 330 nm.

4.6.3 Fourier Transform Infrared Spectroscopy (FTIR)

IR spectroscopy was employed for the characterization of cocrystal. The samples were scanned using Bruker Alpha II IR Spectrophotometer.

4.6.4 Differential Scanning Calorimetry

DSC analysis of mesalamine and cocrystal was done using METTLER TOLEDO DSC1 STARe System. About 3 mg of sample was taken in aluminium pans and sealed using lids. The event was run within the temperature range of 100-350°C at a ramp of 10°C/min, under a nitrogen atmosphere with a flow rate of 50 mL/min.

4.6.5 Crystal Structure Determination From PXRD

The crystal structure was determined from its PXRD pattern using Ultima IV, Rigaku Corporation, Japan. The silicon sample holders were used to obtain diffraction patterns of pure mesalamine. The instrument was equipped with a fine focus X-ray tube (X-ray Source: Cu K-α) and each sample was analysed at X-ray wavelength of 1.5406 Å.

4.6.6 Scanning Electron Microscopy SEM

Mesalamine and M+SacchSod cocrystal was subjected to evaluation by SEM to study the surface morphology and result is depicted. Due to greater solubility and considering analytical evaluation only M+SacchSod cocrystal was evaluated by SEM. However remarkable change was observed in the M+SacchSod 1:1 cocrystal as structure was observed in the SEM image.

4.6.7 Pre-Compression studies

1. Bulk Density

Bulk Density of compound varies substantially with the method of crystallization, milling or formulation. Bulk density was determined by placing powder mix into a graduated cylinder via a large funnel and measuring its volume and weight.

Bulk Density=weight of powder bulk volume of powder mixed

2. Tapped Density

Tapped density was determined by placing powder mix into a graduated cylinder via large funnel and tap the cylinder 50 times and measuring its volume and weight.

Tapped density=weight of powderTapped volume of powder mixed

3. Angle of Repose

The method used to find angle of repose is to pour the powder through funnel on flat surface and measure the inclined angle with the horizontal.

Tan θ=Height of the heap Radius of the heap

4. Hausner’s Ratio

It indicates the flow properties of the powder and ratio of tapped density to the bulk density of powder.

Hausner’s Ratio =Tapped density of powder mixedbulk density of powder

5. Compressibility index

Compressibility index was measured using the values of bulk density (BD) and tapped density (TD). The following equation was used to find the compressibility index.

4.6.8 Post compression tests of tablets

1. Hardness

Hardness is force required to break a tablet in diametric compression. Hardness of tablet was determined by Monsanto hardness tester. The tablet hardness of 5 kg is considered as suitable for handling the tablets.

2. Tablet Size and Thickness

The thickness of the tablet was measured by vernier calipers scale. The thickness of tablet is related to the tablet hardness and can be used as initial control parameter. Tablet thickness should be controlled within range ±5%

3. Friability

The test was performed to evaluate the ability of tablet to withstand abrasion in packing, handling and transporting. Initial weight of 20 tablets was taken and these were placed in friabilator, rotating at 25 rpm for 4 min. the difference in weight is noted and expressed as percentage. It should be preferably between 0.5% to 1.0%.

% friability= (weigth of tablet before test-weigth of tablet after test )×100weight of tablet  before test

4. Weight Variation Test

It is desirable that all the tablets of a particular batch should be uniform in weight. If any weight variation is there, that should fall within the prescribed limits. 20 tablets were taken randomly and weighed accurately. The average weight was calculated by using formula,

Average weight = weight of 20 tablets20

5. Disintegration test

To test disintegration, one tablet was placed in each of the six disintegration, one tablet was placed in each of six disintegration tube of basket rack assembly which was kept in one liter beaker of phosphate buffer pH6.8, simulated intestinal fluid maintained at 37^oc.

4.7 Dissolution Studies

Drug dissolution study was carried out in 900 ml of phosphate buffer pH6.8 at 37^oc, using USP paddle method at stirring speed of 50 rpm. At regular time interval of 10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, 110 min, 5 ml of samples were withdrawn and immediately replaced with an equal volume of fresh dissolution medium. Dissolution profile was compared with that of mesalamine tablet.

6. Results and Discussion

6.1 Analytical Method Development

The analytical method for mesalamine was developed and validated for the quantification of drug in the bulk and pharmaceutical formulation according to ICH Q2R1 guidelines. The method was developed in the distilled water and phosphate buffer pH 6.8. The results are given in table. The calibration curve for the piroxicam in distilled water and phosphate buffer pH 6.8 is represented in fig. 1. The absorption spectrum of mesalamine in water and phosphate buffer pH 6.8 is given in figure.

Fig 1: Absorption spectrum of mesalamine in distilled water

Table 6: Concentration and absorbance of mesalamine in distilled water

Fig 2: Calibration curve of mesalamine in water

Fig 3: Absorption spectrum mesalamine in phosphate buffer pH6.8

Table 7: Concentration and absorbance of mesalamine in phosphate buffer pH6.8

Fig 4: Calibration curve of mesalamine in phosphate buffer pH6.8

6.2 Saturation Solubility study of pure drug and cocrystal

Cocrystal of mesalamine with Ascorbic Acid, Sodium Acetate, urea, Saccharine sodium was observed with more solubility enhancement almost more than 40%. so are selected for further analysis DSC, XRD, SEM, etc. which will confirm the formation of cocrystals.,

Fig 5: mesalamine cocrystal solubility study in distilled water

Analytical characterization of mesalamine and mesalamine Cocrystal

6.3 Infrared spectroscopy

Comparison of the IR spectrum of drug and cocrystal, clearly indicates that there is hydrogen bond formation between drug and coformer and new supramolecular synthon is formed.

1. IR Spectrum of mesalamine:

Fig 6: IR Spectra of Mesalamine

6.4 IR Spectrum of M+SacchSod cocrystal:

Fig 7: IR Spectrum of M+SacchSod cocrystal

Table 9: Major shifts in absorption peaks of M+SacchSod cocrystal

6.5 Differential scanning calorimetry

Mesalamine and cocrystals are characterized by DSC. Pure drug and cocrystal shows characteristic endothermic peak at different temperature corresponding to melting point. This indicates possibility of formation of cocrystal. The change in thermal properties were reported as evidence for the formation of cocrystal.

1. DSC thermogram of Mesalamine

Fig 8: DSC thermogram of Mesalamine

DCS thermogram of M+SacchSod cocrystal:

Fig 9: DSC thermogram of M+SacchSod Cocrystal

The cocrystal showed substantial difference in melting point 265.52°Cin comparison to pure drug 284.06°C. moreover, the peak onset for pure drug was obtained at 288.50°C whereas 263.16°C for cocrystal which indicates possibility of formation of cocrystal.

6.6 Powder X-ray diffraction (PXRD)

1. Powder X-ray diffraction of Mesalamine

The PXRD Patterns of cocrystals are shown in figure . Material in powder state give distinctive peak of varying intensity at certain position. The PXRD patterns of cocrystals was distinguishable from components and additional peaks were appeared which did not exist in pure drug. The appearance of new diffraction peak in diffractogram of cocrystals shows formation of new crystalline face.

Fig 10: PXRD pattern for Mesalamine

2. Powder X-ray diffraction of M+SacchSod cocrystal:

Fig 11: Powder X-ray diffraction of M+SacchSod cocrystal

6.7 Scanning Electron Microscopy SEM

The surface morphology of the Mesalamine was studied by SEM. The scanning electron microscopic images of Mesalamine at 500X, 1000X, 2000X and 5000X magnification shows its crystalline structure.

The Mesalamine, M+SacchSod cocrystal was subjected to evaluation by SEM to study the surface morphology and result is depicted in figure. Due to greater solubility and considering analytical evaluation only M+SacchSod cocrystal was evaluated by SEM. Remarkable change was observed in the M+SacchSod cocrystal. The substantial enhancement in crystallinity supports the formation of cocrystal

6.8 Preformulation Study of Pure Drug and Cocrystal

Table 10: Preformulation study of pure drug and cocrystal

The pure mesalamine and M+SacchSod cocrystal were subjected to Preformulation study and results are given in table. The results indicate that bulk and flow properties of cocrystal were improved in comparison to pure drug. This may be due to formation of new crystalline phase with altered particle properties like size, shape etc. Hence cocrystal may provide opportunity to enhance the bulk and flow properties of the API with improved processing properties.

6.9 post-compression parameters of tablet

Table 11: Post-compression parameters of tablet

6.10 Dissolution Studies

The dissolution rate plays crucial role in the bioavailability of drugs with poor solubility. The dissolution experiment was conducted on the tablet of pure drug and cocrystals. The dissolution profile of the pure drug and the prepared cocrystal is shown in Figure 32. The dissolution profile of pure drug tablet indicates slow dissolution rate with only 11.25% of the drug dissolved in the 60 min. The total amount of drug dissolved in 120 min was 49.35%. However, cocrystals of the M+SacchSod cocrystal tablet resulted in significant increase in the dissolution rate. The amount of drug dissolved in first 60 min was 16.70% and. The total amount of drug dissolved in 120 min was 65.24%. Greater dissolution of mesalamine from cocrystal can be attributed to enhanced solubility of cocrystal in the dissolution media.

Fig 14: drug release profile of tablet of pure mesalamine and M+SacchSod cocrystal tablet