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Year : 2020  |  Volume : 31  |  Issue : 5  |  Page : 685-693
Evaluation of mucoadhesive dexamethasone sodium phosphate gel in the treatment of arecoline-induced oral submucous fibrosis in wister albino rats: A cross-sectional study

1 Department of Pharmaceutical Techonology, HKES's Matoshree Taradevi Rampure Institute of Pharmaceutical Sciences, Kalburgi, Karnataka, India
2 Department of Pharmacology, HKES's Matoshree Taradevi Rampure Institute of Pharmaceutical Sciences, Kalburgi, Karnataka, India
3 Department of Pharmacognosy, HKES's Matoshree Taradevi Rampure Institute of Pharmaceutical Sciences, Kalburgi, Karnataka, India

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Date of Submission02-Sep-2019
Date of Decision22-Dec-2019
Date of Acceptance31-Mar-2020
Date of Web Publication08-Jan-2021


Aim: The present work aimed to prepare an oral mucoadhesive gel of dexamethasone sodium phosphate to serve the purpose of treating oral submucous fibrosis by incorporating the drug in a polymeric matrix to facilitate the localisation of the drug at the absorption site, to prolong drug delivery and to provide patient convenience. Materials and Methods: The formulations F1, F2 and F3 were prepared using 2, 2.5 and 3% of carboxymethyl cellulose sodium, formulations F4, F5 and F6 were prepared using 2, 2.5 and 3% of hydroxypropyl methylcellulose, respectively, and formulations F7, F8 and F9 were prepared using equal mixtures of carboxymethyl cellulose sodium and hydroxypropyl methylcellulose in the concentrations of 1, 1.25 and 1.50%, respectively. The prepared formulations were subjected for screening of physicochemical parameters, viz, homogeneity, grittiness, viscosity studies, spreadability, extrudability, mucoadhesive strength, pH, drug content uniformity, in vitro drug diffusion, Fourier transform infrared spectroscopy spectral analysis and stability studies. Results: Among the nine formulations prepared, the formulation F8 containing 1.25% carboxymethyl cellulose sodium, 1.25% hydroxypropyl methylcellulose having a mucoadhesive strength of 12.600 ± 0.01 g and drug release of 88.473 ± 0.457% was considered as the promising one and was further used for in vivo study. Conclusion: Oral application of the gel for 4 months in arecoline-induced oral submucous fibrosis rats showed more than 80% reduction in fibrosis. The histopathological results supported these findings.

Keywords: Carboxymethyl cellulose sodium (Na CMC), dexamethasone sodium phosphate, hydroxypropyl methylcellulose, mucoadhesive, oral submucous fibrosis

How to cite this article:
Desai VC, Shirsand SB, Malpani A, Hiremath S. Evaluation of mucoadhesive dexamethasone sodium phosphate gel in the treatment of arecoline-induced oral submucous fibrosis in wister albino rats: A cross-sectional study. Indian J Dent Res 2020;31:685-93

How to cite this URL:
Desai VC, Shirsand SB, Malpani A, Hiremath S. Evaluation of mucoadhesive dexamethasone sodium phosphate gel in the treatment of arecoline-induced oral submucous fibrosis in wister albino rats: A cross-sectional study. Indian J Dent Res [serial online] 2020 [cited 2021 Nov 27];31:685-93. Available from:

   Introduction Top

Oral submucous fibrosis (OSMF) is a generalized pathological state of oral mucosa wherein the connective tissue of lamina propria, which undergo a fibroelastic change causing mucosal rigidity of varying intensities leading to increased risk of oral cancer.[1],[2] The symptoms and signs of OSMF include inflammation, hypovascularity, depapillation, blanching, dryness, burning sensation, ulceration in the mouth, hypomobility of tongue and soft palate associated with difficulty in maintaining oral hygiene, proper speech, normal mastication and swallowing.[3],[4] Increase in fibrinogen and its degradation products in plasma are seen in OSMF which cause excessive deposition of fibrin in connective tissue. This brings about the progressive restriction of mouth opening and also affects pharynx and oesophagus.[5],[6]

OSMF is commonly caused by frequent oral consumption of tobacco, catechu, slaked lime, betel quid and areca nut in its various forms like gutkha, pan masala for longer duration as habit.[7] The contents of the areca nut like alkaloids (arecoline) and flavonoids (catechin, tannin) enhance collagen production, strengthen cross-linking and reduce degradation of collagen.[8] The friction caused by coarse fibres of areca nut cause micro trauma facilitating alkaloid diffusion into subepithelial connective tissues resulting in juxtaepithelial inflammation of oral mucosa.[9]

OSMF can be effectively treated with submucosal injection of dexamethasone alone and also in combination with hyaluronidase and placental extract.[10],[11],[12] Dexamethasone, an immunosuppressive agent acts antagonistically on the soluble factors generated by sensitised lymphocytes following the activation by nonspecific antigens. Fibrosis is prevented by reducing the fibroblastic proliferation and deposition of collagen.[13]

Intralesional injection of dexamethasone suppresses inflammatory reaction, reduces trismus and burning sensation.[14] Treatment with submucosal injection causes mechanical injury by repeated use of needles at multiple sites of inflamed tissue. The study was planned to design a non-invasive method of drug delivery system.[15] Hence oral mucoadhesive gels can be effectively used to target the local disorders of mucosal surfaces to increase patient convenience and compliance. These gels are easily dispersed throughout the mucosa for drug delivery. The rich vascularisation of oral mucosa facilitates the drug absorption at a constant rate for prolonged periods by increasing its bioavailability.[16]

The objective of the present work is to design and formulate oral mucoadhesive gels of dexamethasone sodium phosphate using hydrophilic synthetic polymers, viz. sodium CMC and HPMC K100 at different concentrations and combinations. The formulated gels are subjected to physicochemical evaluations to determine their viscosity, mucoadhesive strength, drug release and to select the best formulation among them for in vivo study.

   Materials and Methods Top

Dexamethasone sodium phosphate, hydroxypropyl methylcellulose K100 (HPMC K100), carboxymethyl cellulose sodium (Sodium CMC), sodium meta bisulphite and glycerol.

Formulation of oral mucoadhesive gels of dexamethasone sodium phosphate

The polymers sodium CMC and HPMC were used alone and in combination of equal mixtures of both for preparing the gels.[17],[18] Dexamethasone sodium phosphate was dissolved in glycerine, mixed with polymer and preservatives like sodium meta bisulphite and further subjected for hydration for 24 h. The prepared gels were filled in empty aluminum tubes and labeled accordingly.[19] The formulation details are given in [Table 1].
Table 1: Formulation of oral mucoadhesive gels

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Characterisation of prepared mucoadhesive gels

The prepared formulations were subjected for the evaluation of the following physicochemical parameters:


The prepared gels were allowed to set in a clean glass beaker and inspected for proper appearance and presence of aggregates.[20]


All the formulations were evaluated microscopically for the presence of particulate matter.[20]


One gram of the prepared gel was placed between two horizontal plates (20 cm × 20 cm) to which 125 g weight was applied on the upper plate. The spreading diameter of the gel was measured after 1 min.[20]


The gel under study was filled in one-ounce aluminum collapsible tube having a nasal opening of 5 mm. The extrudability of the gel was determined by measuring the amount of gel extruded out through the tip when a constant load of 1 kg was applied. The extruded gel was collected on the pan and weighed.[20]

Determination of pH

About 5 ± 0.1 g gel was dispersed in 45 ml of water and the pH of this suspension was determined at 27° using the pH meter.[20]

Determination of drug content uniformity

About 1 g of gel (containing 1000 μg of drug) was dissolved in 100 ml of 6.4 pH phosphate buffer solution to give 10 μg/ml. The absorbance was measured at 240 nm by UV spectrophotometer against blank. The blank solution was prepared in the same manner as above using gel containing the respective polymers and other additives without drug.[20]

Determination of viscosity

A Brookfield Capcalc V3.0 Build 20.0 viscometer was used to carry out the viscosity studies of the gels using spindle-01. The measurements were recorded at various speeds ranging from 10, 15, 20, 25 and 30 rpm for 30 seconds between two successive speeds as equilibration time and then in a descending order, with shear rate ranging from 133 s-1 to 400 s-1.[21]

The viscosity data was plotted for rheograms

  1. Shear rate versus shear stress
  2. Log of shear rate versus log of shear stress
  3. Viscosity versus speed.

Mucoadhesive studies

The bioadhesive force of the prepared gels was determined by using assembled device fabricated in a laboratory. The sections of fresh goat buccal tissue were fixed using cyanoacrylate adhesive allowing the mucosal surface outside upon two glass vials separately which were maintained at 36.5 ° for 10 min. One vial being connected to the balance, the other vial was placed on a height-adjustable pan. About 1 g of gel was applied onto the buccal tissue of one vial. Subsequently, the height of the other vial was adjusted so that the gel applied on the mucosal surface of one vial would coincide and adhere to the mucosal tissue surface of the other vial vertically. The weights were gradually added in ascending order until the two vials detached. Bioadhesive force was determined from the minimal weights required to separate the two vials. The buccal tissue pieces were changed for each measurement.[22]

In vitro drug diffusion studies

A glass cylinder of 10 cm height, 3.7 cm outer diameter, 3.1 cm inner diameter having both ends open was taken. Cellophane membrane soaked in distilled water (24 h before use) was fixed to one end of this cylinder with an adhesive to result the permeation cell. One gram of the gel under study was kept in it. A beaker containing 100 ml of 6.4 pH buffer solution acts as receptor compartment. The sample was immersed to a depth such that it should be below the surface of medium in the receptor compartment. The medium in the compartment was agitated using a magnetic stirrer at the temperature 37° ± 1°. 10 ml of samples were withdrawn every 10 min interval and assayed at 240 nm upto 1 hour. The volume withdrawn each time was replaced by equal amount of medium. All the studies were conducted in triplicates and standard deviation (S.D) was calculated.[23]

The results of in vitro release were fitted into four models of data treatment as follows:

  1. Percent cumulative drug release versus time
  2. Log percent cumulative drug remaining versus time
  3. Percent cumulative drug release versus square root of time
  4. Log percent cumulative drug released versus log of time.

Drug polymer interaction studies

The studies were carried out using FTIR method with the help of a SHIMADZU IR PRESTIGE spectrophotometer. The FTIR spectrum of the pure drug dexamethasone sodium phosphate and dexamethasone sodium phosphate with other excipients in gel formulations were studied for their interactions.

Stability studies

For the period of 6 mo the prepared gels were stored at room temperature to confirm the changes in physical appearance, pH, drug content uniformity, viscosity and mucoadhesive strength.

In vivo studies for evaluation of reduction in fibrosis in arecoline induced OSMF rats

Male Wister albino rats weighing 150 to 250 g were used for in vivo study. The animal experimental study was carried out as per CPSCSEA (HKES/MTRIPS/IAEC/93/2017-18).

The experimental animals were divided into three groups of nine rats each.

Group I- Normal control (G1)

Group II- OSMF-induced group (G2)

Group III- Drug-treated OSMF-induced group (G3).

The in vivo study was carried in two phases: Induction of OSMF in rats (4 mo) followed by treatment of OSMF-induced rats (4 mo).[24],[25]

Induction of OSMF

To induce OSMF, 1% arecoline hydrobromide mucoadhesive gel [Table 2] was applied with the help of cotton bud unilaterally on to the buccal mucosa of the experimental animals daily for 4 months. After application of the gel, the animals were fasted for a period of six hours and then were fed with regular diet. At the end of first, second and fourth month respectively, one rat from each group was sacrificed and the biopsy sample of oral mucosa was taken using skin biopsy punch No. 3.5. The biopsy samples were preserved in normal saline vials and were subjected for histopathological studies. The biopsy samples of oral mucosa of control group were also collected and set aside for comparison.
Table 2: Formulation of oral mucoadhesive arecoline gel

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After four months of successive OSMF induction, the animals were treated by the formulation F8 (best formulation). F8 was applied daily on to the affected part of oral mucosa of the rats for the next fourth month. At the end of first, second and fourth month respectively, two rats from each group were sacrificed and the biopsy samples of oral mucosa were taken and subjected for histopathological studies to evaluate the effect of the treatment.

Statistical studies

Statistical studies were carried out by One-way Analysis of Variance (ANOVA) using Dunnett's multiple comparisons test.

   Results and Discussion Top

The characterization of prepared mucoadhesive gels exhibited the following results


All the gel formulations prepared were free from lumps and grittiness. No particulate matter was present in the gels when observed under light microscope [Table 3].
Table 3: Evaluation parameters for oral mucoadhesive gel formulations

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The spreadabilities of the gels were in the range of 26 ± 2.0 to 71 ± 1.5 mm after 1 min [Table 3]. The viscosities of the gel formulations F7, F8 and F9 prepared by using equal proportions of sodium CMC and HPMC were low, hence these formulations showed high spreadability and good extrudability.

Mucoadhesive strength

The mucoadhesive strengths of the gels were between 7.800 ± 0.005 to 13.900 ± 0.05 g [Table 3]. The gel formulations F7, F8 and F9 containing equal proportions of sodium CMC and HPMC exhibited good mucoadhesive strength even though less viscous than other formulations. The mucoadhesive strength depends upon the type, concentration, and combination of the polymers used.


The pH of the gels were between 6.4 ± 0.06 to 6.7 ± 0.21 [Table 3]. Since the pH of the gels was same as the oral pH of saliva, they were free from irritation.

Drug content uniformity

The percentage drug contents of all the formulations were in the range of 97.00 ± 0.599 to 99.70 ± 0.173 [Table 3]. All the gels showed uniformity in drug content.


The viscosities of the gels were in the range of 6872 to 12991 cps at low shear rate and 3799 to 7087 cps at high rate of shear, respectively [Table 3].

The viscosity measurements were made at varying speed and shear rates. All the prepared gel formulations showed shear thinning/pseudoplastic behaviour at room temperature wherein the viscosity of the gels decreases as the shear rate is increased [Table 4]. Upon plotting the log of shear stress versus log of shear rate, a straight line was obtained with slope N [Figure 1]. The slope value N for F1, F2, F3, F4, F5, F6, F7 and F8 were 1.621, 3.270, 2.194, 1.650, 3.335, 2.794, 1.865, 4.563 and 3.355, respectively. For the pseudoplastic property, the N value should be greater than 1 and a higher value of N signifies better pseudoplastic behavior. The N value was highest for F8.
Table 4: Viscosity data of oral mucoadhesive gel formulation F8 at room temperature

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Figure 1: Log of shear stress vs. log of shear rate (F8)

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For all shear thinning systems, rheogram was obtained by plotting the shear rates at various shear stresses, here the 'up curve' and 'down curve' do not superimpose [Figure 2].
Figure 2: Shear rate vs. shear stress (F8)

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The graph was plotted for viscosity versus speed. The viscosity of the prepared gels is inversely proportional to speed. At high speed, the curve appears straight indicating minimum viscosity [Figure 3].
Figure 3: Speed vs. viscosity (F8)

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Increase in the viscosity of the gels decreases the drug release. The order of decreasing percentage drug release in 2 hour were F7 > F4 > F1 > F8 > F9 > F5 > F6 > F2 > F3.

In vitro drug diffusion studies

The formulations exhibited a cumulative percent drug release in the range of 73.353 ± 0.753 to 98.802 ± 0.792 [Table 5] and [Figure 4]. The regression coefficient values of the kinetic equations like zero-order plots, first-order plots, Higuchi diffusion plots, and Korsmeyer Peppas plots were nearer to one, suggesting that the plots are fairly linear. The drug release is by non-Fickian release mechanism as the slope values obtained from Korsmeyer Peppas plots were in the range of 0.941 to 0.985 [Table 6].
Table 5: Percent cumulative drug release from gel formulations at various time intervals

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Figure 4: Percent cumulative drug release from gel formulations at various time intervals

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Table 6: Kinetic data of oral mucoadhesive gels

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Drug polymer interaction studies

The FTIR of dexamethasone sodium phosphate was adopted to characterize the potential interactions with the excipients used [Figure 5]a and [Figure 5]b.
Figure 5: FTIR Spectra of the Dexamethasone sodium phosphate alone (a) and with excipients and polymers (b) in the optimized formulation F8

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In the spectrum of dexamethasone sodium phosphate, the broad band at 3272 cm-1 corresponded to the free -OH groups. The peak at 1651 cm-1 was caused by -C = C- structure. The peak at 1031 cm-1 is due to the -C-F structure. The peak at 1209 cm-1 is due to -C-O-C- structure.

The FTIR spectrum of dexamethasone sodium phosphate mucoadhesive gel formulations showed the characteristic absorption bands, which resemble that of the pure drug dexamethasone sodium phosphate, which indicates formation of compact drug-polymeric network with no drug-excipients interactions.

Stability studies

After 6 months of stability studies, physical changes like color fading or separation of liquid exudates of the gels were not observed. The pH of all the gels was not affected and was in the range of 6.4 to 6.8. The strength of all the prepared gels remained unchanged after 6 months and the drug content was in the range of 98.20 to 99.85%.

All the prepared gels passed the stability studies with marginal significant changes in physical appearances, viscosity, pH, drug content and mucoadhesive strength.

In vivo studies

Among all the formulations, F1, F4 and F7 containing 2% polymer showed the highest release and spreadability but exhibited very low mucoadhesive strength. The F3, F6 and F9 formulations containing 3% polymer had high mucoadhesive strength but exhibited slow release rate and poor spreadability. However F2, F5 and F8 containing 2.5% polymer showed significance in release rate, mucoadhesive strength, spreadibility, and extrudability. The F8 formulation exhibited prominent mucoadhesive strength of 12.600 ± 0.01 g and drug release of 88.473 ± 0.457% compared to F2 and F5. Hence the formulation F8 was considered to be suitable for carrying out in vivo studies.

The in vivo study was focused on the array of histomorphological changes and to observe the percentage of healing in the oral mucosa of untreated and mucoadhesive dexamethasone sodium phosphate gel-treated OSMF-induced rats.

The histopathological study of the oral mucosa of the OSMF-induced group under phase I study showed redness of mucosa, no alterations in the thickness of epithelial lining and the collagen content of submucosa at the end of 1st months [Figure 6]b. Biopsy taken at the end of 2nd months showed mild pink colour of mucosa, rare thinning of epithelial lining and moderate increase in collagen content of submucosa [Figure 6]c. A white mucosal patch, minute epithelial lining and dense collagen at submucosa was observed with biopsy taken at the end of 4th months [Figure 6]d. However, no change was observed in the oral mucosa of the rats under control group [Figure 6]a.
Figure 6: Histopathological changes in the oral mucosa of rats at different observation periods during induction of OSMF: 6a - Normal oral mucosa; 6b - 1 month induction: no change in the thickness of the epithelial lining (a), no significant alteration of collagen content in submucosa (b); 6c - 2 months induction: slight thinning of epithelial lining (a), moderately increased collagen content in submucosa (b); 6d - 4 months induction: very thin epithelial lining (a), white patch in the mucosa and thick collagen deposition in submucosa (b)

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The experimental animals showed significant (P < 0.0001) reduction in body weight from 146 ± 4 to 91 ± 4 g at the end of 1st and 4th months of induction, respectively [Table 7].
Table 7: Body weights of rats during induction and treatment period of OSMF

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The histopathological study of the oral mucosa of OSMF-treated group under phase II study showed atrophic epithelium, abundant collagen at submucosa and chronic inflammation at the end of 1st months [Figure 7]b. The biopsy of 2nd months showed restoration of epithelium to normal thickness and moderate decrease in collagen of submucosa [Figure 7]c and that of the 4th months showed almost normal epithelium with 80% dissolution of collagen of submucosa [Figure 7]d. The animals showed significant (P < 0.0001) weight gain from 116 ± 4 g to 185 ± 3 g from the end of 5th months till the end of 8th months of treatment, respectively, [Figure 8] due to improvement in mouth opening.
Figure 7: Histopathological changes in the oral mucosa of rats at different observation periods during treatment of OSMF: 7a - 4 months induction: very thin epithelial lining (a), thick collagen deposition in submucosa (b), white patch in the mucosa (c); 7b - 1 month treatment: atrophic epithelium (a), dense collagen deposition in submucosa (b), pale pink coloured mucosa (c); 7c - 2 months treatment: epithelium regaining its normal thickness (a), moderate decrease in collagen in submucosa (b), appearance of blood capillaries in mucosal layer (c); 7d – 4 months treatment: epithelium almost regaining its normal thickness (a), more than 80 % dissolution of collagen in submucosa (b)

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Figure 8: Changes in the body weights of rats at different observation periods during OSMF induction and OSMF treatment

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In the first phase of histopathological studies of OSMF induction, there was progressive increase in fibrosis in submucosa from 1st months to 2nd months and by the end of the 4th months, definitive evidence of OSMF were visible in the rats. In comparison with the control group, the OSMF-induced rats exhibited difficulty in mouth opening, a dim coat color, activity decline, lack of appetite, anemia and significant (P < 0.0001) weight loss during the later stage of the experiment. In the second phase of histopathological studies of OSMF treatment, 80% collagen reduction in submucosa, improvement in mouth opening and weight gain was noticed by the end of 8th months.

   Conclusion Top

The physicochemical evaluation results of the prepared dexamethasone sodium phosphate oral mucoadhesive gels indicate that formulaton F8 had enhanced mucoadhesive property, drug delivery and bioavailability to the oral mucosa. In vivo pharmacological evaluation of the same in the treatment of arecoline-induced OSMF rats showed a noticeable decline in collagen content in submucosa towards reversing the symptoms of OSMF. Therefore, our future studies will be planned to warrant the existing study findings in healthy human volunteers suffering from OSMF.

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Conflicts of interest

There are no conflicts of interest.

   References Top

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Correspondence Address:
Vijaybhaskar C Desai
Department of Pharmaceutical Techonology, HKES's Matoshree Taradevi Rampure Institute of Pharmaceutical Sciences, Kalburgi - 585 105, Karnataka
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ijdr.IJDR_685_19

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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8]

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6], [Table 7]


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