Evaluating the Role of Celosia argentea Powder and Fenugreek Seed Mucilage as Natural Super-Disintegrating Agents in Gliclazide Fast Disintegrating Tablets

The bioavailability of the drug is dependent on in vivo disintegration, dissolution, and various physiological factors1. Tablet disintegration has received considerable attention as an essential step in obtaining fast drug release2. The emphasis on the availability of drugs highlights the importance of the relatively rapid disintegration of a tablet as a criterion for ensuring uninhibited drug dissolution behavior3. “Disintegrants are agents added to tablet and some encapsulated formulations to promote the breakup of the tablet and [capsule “slugs”] into smaller fragments in an aqueous environment thereby increasing the available surface area and promoting a more rapid release of the drug substance”4. Superdisintegrants are generally used at a low level in the solid dosage form, typically 110 % by weight relative to the total weight of the dosage unit5. Diverse categories of superdisintegrants such as synthetic, semi-synthetic, natural, and co-processed blends, etc. have been employed to develop effective immediate-release tablets and to overcome the limitations of conventional tablet dosage form6.


INTRODUCTION
The bioavailability of the drug is dependent on in vivo disintegration, dissolution, and various physiological factors 1 . Tablet disintegration has received considerable attention as an essential step in obtaining fast drug release 2 . The emphasis on the availability of drugs highlights the importance of the relatively rapid disintegration of a tablet as a criterion for ensuring uninhibited drug dissolution behavior 3 . "Disintegrants are agents added to tablet and some encapsulated formulations to promote the breakup of the tablet and [capsule "slugs"] into smaller fragments in an aqueous environment thereby increasing the available surface area and promoting a more rapid release of the drug substance" 4 . Superdisintegrants are generally used at a low level in the solid dosage form, typically 1-10 % by weight relative to the total weight of the dosage unit 5 . Diverse categories of superdisintegrants such as synthetic, semi-synthetic, natural, and co-processed blends, etc. have been employed to develop effective immediate-release tablets and to overcome the limitations of conventional tablet dosage form 6 .
Superdisintegrants is used as excipients in the tablet formulation as it has to meet certain criteria other than its swelling properties. The requirement placed on the tablet disintegrant should be clearly defined 7 . A group of superdisintegrants including croscarmellose sodium (ac-di-sol) sodium starch glycolate (Primojel and Explotab) and crospovidone (Polyplasdone XL) alleviate most of these problems 8 . The use of the superdisintegrants in a fast dispersible tablet is possible as a tablet shows optimum physical properties. Some superdisintegrating agents are natural in origin and are preferred . ABSTRACT over synthetic substances because they are comparatively cheaper, abundantly available, non-irritating, and nontoxic in nature 9 . The natural materials like gums and mucilage's have been extensively used in the field of drug delivery for their easy availability, cost-effectiveness, eco-friendliness, emollient, non-irritant nature, non-toxicity, capable of a multitude of chemical modifications, potentially degradable, and compatible due to natural origin 10 . There are several gums and mucilages are available which have superdisintegrating activity such as Plantago ovata seed mucilage (isphagula), Lepidium sativum mucilage, gum karaya, fenugreek seed mucilage, locust bean gum, Cassia fistula gum, and guar gum 11 .
The major objective was to develop a fast disintegrating tablet (FDTs) is accomplished by using suitable excipients and superdisintegrants to achieve fast disintegration within a short period of time. FDTs tablet was developed that is safe, efficacious and to get the comparable dissolution profile. The task of developing the FDTs tablet of gliclazide (GLZ) by using superdisintegrants was needed because of its low aqueous solubility and low bioavailability. The research work was aimed to design and characterization of fast disintegrating GLZ-based anti-diabetic tablet using the natural disintegrating agent; fenugreek seed mucilage (Batch: FS1-FS3) and Celosia argentea (Batch: FM4-FM6) as well as a synthetic disintegrating agent; sodium starch glycolate (Batch: FC7-FC9), which results in a stable dosage form.

Materials
GLZ was obtained as a generous gift from Zim Laboratories Ltd., Nagpur. The fenugreek seeds were purchased from the local market and C. argentea flower was obtained from a local farm. Sodium starch glycolate and microcrystalline cellulose were procured from Loba Chem Ltd., Mumbai. All other reagents and solvents employed in this study were of analytical grade and purchased from HiMedia Ltd., Mumbai.

Extraction of fenugreek seed mucilage
The seeds were powdered using a grinder and 50 g of the powder was extracted with n-hexane to remove lipophilic compounds using a soxhlet apparatus. To remove pigments and to deactivate enzyme, the defatted powder was boiled in ethanol for 20 min. This treated powder was then soaked in 5 litres water and the pH was adjusted to 3.5 using 0.5 M hydrochloric acid. The mixture was stirred by a mechanical stirrer for 12 h and then filtered through filtration paper. The filtrate was centrifuged (5000 g) and the supernatant was concentrated in vacuum to 50% of its initial volume. The resulting solution was mixed with the same volume of 96% ethanol and stored in a refrigerator for 4 hr. The precipitated mucilage was separated by centrifugation (5000 g). The collected mucilage was re-suspended in distilled water, agitated for 20 min, and re-precipitated one more time to eliminate chloride ions and other impurities. Finally, the residue was washed with diethyl ether and acetone and dried overnight at 45°C, resulting in an off-white powder.

Preparation of C. argentea powder
The dried C. argentea flowers were collected and dried in sunlight. The size reduction was done in the grinder and sieved through mesh #80 and stored in a desiccator.

Phytochemical screening
The phytochemical investigation was done for the presence of glycoside, alkaloid, cellulose, mucilage, steroid, protein, reducing sugar, and tannin, according to Kamble et al., 2017 12 .

Swelling ratio
The swelling ratio is the volume in ml occupied by 1 g of the drug; including any adhering mucilage after it has been swollen in an aqueous liquid for 4 hr. One gram of powder was taken in a 25 mL ground glass stoppered cylinder graduated over a height of 120 to 130 mm in 0.5 divisions. To this 25 mL of water was added and this was shaken vigorously every 10 m for 1 hr and then allowed to stand for 24 hr 13 . The volume occupied by the disintegrating agent including adhering mucilage was measured. The swelling ratio was calculated from the mean of three determinations.
Swelling ratio = V 2 -V 1 / V 1 where, V 1 = initial volume of material before hydration; V 2 = volume of hydrated material.

Loss on drying
Loss on drying was carried out by using a hot air oven at 105˚C for 2 hr 14 .

Characterization of Drug and Excipients
Pre-formulation study on GLZ The GLZ was subjected to various pre-formulation studies like solubility, description, and pH.

Fourier Transform Infrared (FTIR) Spectroscopy
This study has been done to check whether there is any compatibility related problems are associated with drug and the excipients used for the formulation of fast disintegrating tab-lets. The drug and excipients must be compatible with one another to produce a product that is stable, efficacious, attractive, and easy to administer and safe. If the excipients are new and not been used in formulations containing the active substance, the compatibility studies are of paramount importance. FTIR can be used to investigate and predict any physicochemical interactions between components in a formulation and can therefore be applied to the selection of suitable chemically compatible excipients. The study was conducted with an intention to check the compatibility of disintegrants fenugreek seed mucilage, sodium starch glycolate, and C. argentea powder with GLZ. Also, it helps to check the suitability of disintegrants for the preparation of fast disintegrating tablets. FTIR spectra were studied using a Shimadzu FTIR (Shimadzu IR Affinity-1, Japan) spectrophotometer by KBr disks in the spectral range of 4000 cm −1 to 500 cm −1 15 .

Differential Scanning Calorimetry (DSC)
The thermal behavior of pure drug and polymer were studied using a differential scanning calorimeter (Mettler Toledo ® ) at a heating rate of 10°C/min. The measurements were performed at a heating range of 30-400°C under nitrogen atmospheres 16 .

Formulation of fast disintegrating Tablets (FDTs)
The drug and all excipients were weighed accurately in separate polybags. The active pharmaceutical ingredient (API), fenugreek seed mucilage, C. argentea, sodium starch glycolate, microcrystalline cellulose, mannitol were sifted through sieve no. #40. The above-sifted material was transferred and mixed carefully. Magnesium stearate, talc, and aerosil were sifted through sieve no. #80 and the lubricating material i.e. magnesium stearate was transferred into the above blend materials and mixed for the duration of 5 minutes ( Table 1). The lubricating material of the above blend was compressed in a 10 station compression machine using 8 mm concave punch.

Bulk Density
It refers to the packing of particles. Bulk density was used to determine the amount of drug that occupies the volume in g/ ml. The bulk density of the ingredients was evaluated using a graduated cylinder. It is the ratio of the total mass of powder to the bulk volume of powder. It was measured by pouring the weighed quantity of powder into a graduated measuring cylinder and the volume was noted. It was expressed in g/ml and is calculated by using the following formula.

Tapped density
It is the ratio of the total mass of powder to the tapped volume of powder. The tapped volume was measured by tapping the powder 10, 500, and 1250 taps in tap density apparatus (Electro Lab USP II) according to USP. The blend was subjected for 500 taps; % volume variation was calculated and subjected for additional 1250 taps, and the % variation was calculated.

Compressibility index (Carr's index)
The compressibility index is an important measure that can be obtained from the bulk and tapped densities. In theory, the less the compressible material, the more flowable it is. A material having values of less than 20% is defined as the freeflowing material. The relationship between % compressibility indexes with flowability can be given by:

Weight of tablet
Twenty tablets randomly selected from each batch and individually weighed. The average weight and standard deviation of 20 tablets were calculated. The batch passed the test for weight variation if not more than two of the individual tablet weight deviates from the average weight by more than the percentage shown in officials and none deviate by more than twice the percentage shown in the tablet.

Tablet dimensions
The thickness of the tablets was measured using a Vernier caliper. It was determined by checking ten tablets from each formulation. It is expressed in mm.

Hardness
Hardness indicates the ability of a tablet to withstand mechanical shock while handling. For each formulation, the hardness of the tablet was determined by using Monsanto hardness tester. It is expressed in N. Ten tablets were selected and hardness was measured.

Friability
For tablets with an average weight of 650 mg or less take a sample of whole tablets corresponding to about 6.5 mg and for the tablet with an average weight of more than 650 mg take a sample of 10 whole tablets. The tablets carefully dedusted and the required number of tablets was weighed accurately. Place the tablet in the Roche friabilator. The friabilator was operated at 25 rpm for 4 min then removes any loose dust from them and weighs them accurately. A maximum loss of weight not greater than 1.0 % is acceptable for the tablet. The percent friability was calculated by using the following equation.

Disintegration time
The in vitro disintegration time of each tablet formulation was determined by using a beaker at 37°C±2°C in 900 mL disintegration media. 6 tablets of each formulation were taken and placed in a glass beaker. The time taken for complete disintegration was noted.

Wetting time and water absorption ratio
Double folded tissue paper was placed in a petridish and 6 mL of water was added to the petridish. A tablet was carefully placed on the surface of the tissue paper in the petridish at a temperature of 25ºC. The time required for water to reach the upper surface of the tablets and to completely wet them was noted as the wetting time. These measurements were carried out in replicates of six. Wetting time was recorded using a stopwatch. The wetted tablet was then weighted and the water absorption ratio (R) was determined by using the equation:

R= (W b -W a /W b ) × 100
where, W a = weight of tablet after water absorption; W b = weight of tablet before absorption.

Drug content
Standard solution 100 mg of pure GLZ was dissolved in little quantity of pH 6.8 phosphate buffer solution in a volumetric flask and then the volume was made to 100 mL mark with buffer solution and sonicated for 10 min. The above solution produced 1 mg/mL solution of GLZ which was further diluted with methanol to produce a series of concentrations ranging between 5-25 μg/ mL. The absorbance was measured using spectrophotometer at 226 nm against buffer solution as blank 19 . The standard calibration curve was plotted.
Sample solution 20 tablets from each batch were randomly selected, weighed accurately, and then finely powdered in a mortar. To a powder equivalent to 80 mg of GLZ, about 50 mL of 6.8 pH solution was added and dissolved with the aid of the shaker for 24 hrs. A sufficient quantity of buffer solution was added to produce 100 mL in a volumetric flask, mixed well, and filtered using a membrane filter. 1 mL of the above filtrate was further diluted to 10 mL using buffer solution and mixed well. The absorbance of the resulting solution (10 μg/mL) was measured at the 226 nm using a blank in the reference cell 20 . The total content of the drug in the solution was calculated with the help of a standard graph.

Dissolution study
In vitro dissolution study was carried out for optimized formulation of GLZ in 0.1 N HCl and pH 6.8 phosphate buffer was analyzed spectrophotometrically at 226 nm. The API has pH-dependent solubility in nature; its solubility is inversely proportional to the pH. It has good solubility in strong acidic conditions. During the multimedia dissolution of the given discriminating dissolution media, we had performed dissolution with various dissolution media. The dissolution was performed in pH 6.8 phosphate buffer and 0.1 N HCl 21 .

Stability study
The optimized batches were packed in aluminum foil and performed the stability study at 40°C temperature/ 75% Relative Humidity (RH). The stability samples were evaluated initially, after one month, and finally after three months22.

RESULT AND DISCUSSION
The drug powders were subjected to preformulation studies. The preformulation characteristics are within the pharmacopoeia specification. The preformulation studies were carried out and the results were found to be satisfactory.

Preliminary Phytochemical test
The phytochemical studies of fenugreek seed mucilage indicated the presence of carbohydrate, protein, and mucilage whereas other components such as cellulose, flavonoids, alkaloids, tannins, and reducing sugar. C. argentea powder showed the appearance of carbohydrate, protein, and cellulose while mucilage, flavonoids, alkaloids, tannins, and reducing sugar were absent ( Table 2).

Physicochemical parameters of natural disintegrants
Both the natural disintegrants share similar characteristics in terms of % yield (~30), pH (~5.8), swelling ratio (~7), ash value (~0.88), acid-soluble ash (~0.3), water-soluble ash (~0.37), % LOD (~0.46), and taste (tasteless) which indicated their imperative properties suitable for pharmaceutical applications ( Table 3). Compatibility studies FTIR spectra were recorded for GLZ, C. argentea powder, fenugreek seed mucilage, sodium starch glycolate, and physical mixture. All the above characteristics peaks of the drug appeared in the spectra of the physical mixture at the same wave number indicating no modification or interaction between the drug and the excipients (Figure 1). The drug and excipients compatibility was carried out by the FTIR method and physical observation showed there was no interaction between them.

Differential Scanning Calorimetry
The DSC thermograms were recorded for GLZ, C. argentea powder, fenugreek seed mucilage, sodium starch glycolate, and their physical mixtures. The DSC heating and cooling curves were recorded as a plot of enthalpy vs. temperature in (ºC). The thermal analysis of the drug and disintegrants was confirmed by differential scanning calorimetry at a scanning rate of 10°C/min. GLZ showed sharp melting endothermic peak at 187.52°C, C. argentea powder at 151.08°C, fenugreek seed mucilage at 217.12°C, sodium starch glycolate at 130.05°C whereas the thermogram of physical mixture shows a respective endothermic peak at 167.30 °C for C. argentea powder, 166.69-209.70°C for fenugreek seed mucilage, 171.91°C for sodium starch glycolate (Figure 2), respectively. DSC studies had proven the interaction between the drug and disintegrants where the melting point was found to be reduced because of physicochemical interaction and embedding of the drug into disintegrants network. The study indicated that there was no significant interaction between the drug and the excipients.

Pre-Compression parameters
In the above table characteristic of the powder blend of FS3, FM6, and FC9 is given. The blend ready for compression was evaluated for bulk density, tapped density, compressibility index, and Hausner's ratio. It was found that the blend had a compressibility index from 5.63% to 5.90% and Hausner's ratio from 1.05 to 1.06, which indicate that blend ready for compression. From the values of the Compressibility index and Hausner's ratio, it was observed that a blend of the above formulation has excellent flow properties and compressibility index ( Table 4).

Post-Compression parameters
Total of nine FDT formulations were prepared using different concentrations of fenugreek seed mucilage, C. argentea powder, and sodium starch glycolate. The various formulation of GLZ was prepared by using a direct compression method because of the good flow property of API. The tablets were formulated by using excipients such as microcrystalline cellulose, mannitol, talc, aerosol, and magnesium stearate. The optimized three batches out of all nine batches on the basis of hardness, friability, disintegration time, weight variation, thickness, and drug content were screened. The thickness of the tablets was found to be in the range of 3±0.04 mm. The results showed that the thickness of all formulates tablets are found to be uniform. The hardness of the all tablet formulation was found to be in range (2-3 N) which indicated that all the tablets have adequate mechanical strength. The accepted percentage deviation of ±7.5% for less than 324 mg weight tablets was taken in to account for all the produced formulations. The weight of tablets found within limits. In the friability test, the maximum weight loss should be no more than 1%. The result revealed that the tablets passed the friability test.  Table 5). The formulation of a fast disintegrating tablet was done by direct compression method by using natural disintegrating agents (fenugreek seed mucilage, C. argentea powder) and synthetic disintegrants (sodium starch glycolate) which shows less disintegration time and higher drug release profile. Multimedia Dissolution Data of Fast Disintegrating Tablet: The multimedia dissolution was carried out by using different dissolution media over a period of 60 min using the USP type II (paddle) dissolution apparatus at 50 rpm under 0.1 N HCl and pH 6.8 phosphate buffer. From the result, it was observed that FDTs in 0.1 N HCl media showed improved dissolution rate with a formulation containing C. argentea powder ( Table 6). The % drug release of fast disintegrating tablets in 0.1 N HCl was found to be greater than pH 6.8 phosphate buffer dissolution media (Figure 3).

Stability studies
The optimized batches were packed in aluminum foil and performed stability study at 40ºC / 75% RH for three months. The stability samples were evaluated initially and after one and three months which showed no change in physical appearance, assay, and drug release which indicate the tablet was stable.

CONCLUSION
Fast disintegrating tablet prepared by direct compression. The natural disintegrants (C. argentea powder) showed bet-ter results than synthetic disintegrants (sodium starch glycolate). Form the above result, it can be concluded that batch FC9 showed better results than batch FS3 and FM6, and also shows less disintegration time improvement in dissolution rate by using superdisintegrants that means improving dissolution ultimately improvement in bioavailability. Hence, the anti-diabetic drug GLZ can be successfully formulated as a fast disintegrating tablet with natural disintegrants like C. argentea and fenugreek seed mucilage.