Cellulose buccoadhesive film bearing glimepiride: physicomechanical characterization and biophysics of buccoadhesion
Jaya Gopal Meher1, 2, Magdaline Tarai1, Ansuman Patnaik1, Paresh Mishra1, Narayan Prasad Yadav*2
1. Indira Gandhi Institute of Pharmaceutical Sciences, IRC Village, Bhubaneswar-751015, India
2. Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Lucknow-226016, India
Supplementary Material
Present investigation was directed to the formulation and development of buccoadhesive films with the unique combination of various functional and reactive polymers as well as to investigate the effect(s) of the polymeric combination (developed matrices) on various physico-mechanical parameters of films. Apart from these parameters, the drug release/permeation characteristics and mechanistic insight to the mucoadhesion were also scrutinized. New instrumental techniques were employed and methods of in vitro film assessment were developed. The supplementary materialsare provided in support to the experiments performed in the investigation. These materials will be describing the following issues.
- Test specifications of Contemporary characterization (Film strength, Bucoadhesive strength, Tensile strength) performed with CT3 texture analyzer.
- Zeta potential and particle size of mucin-film complex.
- In vitrobuccocompatibility with pro-biotic microorganisms and film formulations.
- Stability studies data.
- Test specifications of Contemporary characterization (Film strength, Buccoadhesive strength, Tensile strength) performed with CT3 texture analyzer
Any muco/buccoadhesive formulations are characterized for mucoadhesive-/tensile-strength and also for film strength characteristics. But till date the conventional, age-old methods of modified or locally fabricated methods are adopted for these purpose. Since last few years (2-3 years) new instrumental techniques are emerging, which give precise and reproducible results. But these are yet to get popularity/recognition and reasons for this might be the inadequate availability of literature, awareness among the researchers as well as unavailability of respective instruments/infrastructure. Present investigation focuses on the highly precise, reproducible instrumental techniques and for the evaluation of film strength, buccoadhesive strength, tensile strength new methods are developed. The detail method and specification for the same are presented below (Table 1).
Table S-1: Test specification of film strength, buccoadhesive strength and tensile strength characterization by CT3 texture analyzer.
Evaluation parameters / Test Mode / Pre-test speed (mm/sec) / Test speed(mm/sec) / Post-test speed
(mm/sec) / Probe/
Base / Target value
(g/mm) / Trigger force
(g)
Buccoadhesive Strength / Tension (Hold Time) / 1.00 / 0.05 / 0.05 / TA-DEC
TA-BT KIT / 20.0 / 3.00
Film strength / Compression / 1.00 / 2.00 / 2.00 / TA 42
TA-FSF / 5.00 / 5.00
Tensile strength / Compression / 1.00 / 1.00 / 1.00 / TA-DGA / 5.00 / 5.00
- Buccoadhesive Strength
The test procedure is described concisely in the manuscript (section 3.3.2.2). Figure S-1 a representative photograph of buccoadhesive strength test. The experiment was commenced with this programmed test protocol by CT3 texture analyzer. The moment atrigger force of 3.0 g was detected on the film surface, the probe started to get detached from the film at a test-speed of 0.05 mm/sec and the resistance exerted by the sample on the probe is indicated by a peak force i.e.buccoadhesive strength. Area under the peak represented the work done to detach the film from the buccal mucosal membrane. Each film formulation was evaluated by the same protocol and peak force and work done was noted for interpretation. In order to investigate the mechanistic insight to the buccoadhesion, the same test was performed at different time intervals (1, 2, 4, 6 and 8 h) with each film formulation. Results were computed by Texture Pro CT V1.3 Build 15 software for buccoadhesive strength evaluation.
Figure S-1: Representative photograph of buccoadhesive strength test
- Film strength
The test procedure is mentioned briefly in the manuscript (section 3.3.2.1). As and when a trigger force of 5.0 g was detected, the test started. The probe approached to make puncture on the sample and travelled to a deformation distance of 5 mm. The maximum peak represented the force required to cause the initial breaking of the film material. The area under the curve represented the work done in the procedure. Figure S-2 shows the photographic representation of film strength evaluation. Results were computed by Texture Pro CT V1.3 Build 15 software for buccoadhesive strength evaluation.
Figure S-2: Representative photograph of film strength test
C. Tensile strength
The test procedure is mentioned briefly in the manuscript (section 3.3.2.3). After getting a trigger force of 5.0 g the test was initiated. The probe was pulled up as per the tension mode set in the texture analyzer, and the maximum peak force required to cause breaking of the film material was measured. The area under the curve represented the work done in the procedure. Each film was subjected to the same test for tensile strength evaluation. Results were computed by Texture Pro CT V1.3 Build 15 software for buccoadhesive strength evaluation.
- Zeta potential and particle size of mucin-film complex
Zeta potential and the particle size of the polymer matrix-mucin mixture was determined to investigate the mucoadhesive phenomena. The polymeric film was homogenized with small volume of distilled water and allowed to stand for 6 h. Meanwhile mucin was dispersed in distilled water to make stock solutions of 1% w/w. Equal volume of both dispersions were mixed and diluted with distilled water. The zeta potential of the mixture was measured using ZetasizerNanoZS (Malvern, UK) at 22±1°C. The mean particle size and particle distribution (polydispersity index; PDI) were measured by the same instrument.
Figure S-3 (A): Particle size of mucin
Figure S-3 (B): Particle size of buccoadhesive film-mucin mixture
Figure S-3 (C): Zeta potential of mucin
Figure S-3 (D): Zeta potential of buccoadhesive film-mucinmixture
The above figures i.e. Figure S-3 (A), (B), (C) and (D) exhibit the particle size and zeta potential of mucin as well as buccoadhesive film-mucin mixture.Mucin showed zeta potential -22.46 ± 7.34 mV and size 140 ± 25 nm. With the addition of Carbopol (Ultrez-21) and poloxamer (pluronic F68) from film to mucin, the zeta potential increased which was because of increase in total negative charge. Particle size of polymeric film-mucin mixture was found to be 540-710 nm.
- In vitrobuccocompatibility
The in vitrobuccocompatibility was investigated against three pro-biotic microorganisms of the buccal micro-flora. Agar diffusion method was employed for this purpose and the results are given in Table S-2. Films without Poloxamer exhibited bacteriostatic action whereas films with Poloxamer showed very weak bactericidal action. The weak bactericidal action might be due to the presence of Poloxamer which is reported to have its own antimicrobial action. A very few literature is available in this regard and we deem it should be done as an essential evaluation criteria before clinical investigation.
In our current investigation we have taken these three micro-organisms as these were found in the buccal cavity. The results are in agreement with the previously reported literature that Poloxamer is having antimicrobial action, whereas other ingredients as well as drug did not exhibited any antimicrobial action.
Table S-2: Buccocompatibility investigation of developed films against pro-biotic micro-organisms
Buccoadhesive Films / Net zone of growth inhibition (mm)Lactobacillus acidophilus / Bifidobacterium
infantis / Lactobacillus
rhamnnosus
GM1 / BS / BS / BS
P-GM1 / BS / BS / BS
GM2 / 3 ± 1 / 2 ± 1 / 3 ± 1
P-GM2 / 2 ± 1 / 1 / 2 ± 1
GM3 / BS / BS / BS
P-GM3 / BS / BS / BS
GM4 / 2 ± 1 / 4 ± 1 / 2 ± 1
P-GM4 / 1 / 1 / 2 ± 1
GM5 / 3 ± 1 / 3 ± 1 / 4 ± 1
P-GM5 / 2 ± 1 / 2 ± 1 / 2 ± 1
GM6 / 4 ± 1 / 4 ± 1 / 2 ± 1
P-GM6 / 2 ± 1 / 2 ± 1 / 1
P-GM; placebo buccoadhesive films, BS; bacteriostatic action
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- Stability studies
The film formulation GM4 was found to be the best among all formulations and hence subjected to the stability evaluation. As discussed in the method section of the manuscript, the films were properly sealed and stored in two different storage conditions i.e. 4 ± 0.5°C and 22 ± 1°C. At regular time intervals of 1, 3 and 6 months the films were investigated for the physical appearance and other physico-mechanical evaluation parameters. Based on the results the film formulations stored at 22 ± 1°C were found to be comparatively stable than the films stored at 4 ± 0.5°C. Table S3 exhibits the data of all tested physico-mechanical investigation at different storage conditions. GM4 films were also subjected to in vitro release and ex vivo permeation studies. No significant difference (P< 0.05) was found in the drug release and permeation of the films after 6 months of storage at 22 ± 1°C and the order as well as mechanism were also found to be the same. The in vitro and ex vivo drug release and permeation zero order plot is shown below (Figure S-4).
Figure S-4: Comparative in vitro drug release and ex vivo drug permeation zero order plot of GM4 film formulation
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Table S-3: Stability evaluation of buccoadhesive films (GM4) at different storage conditions.
Evaluation parameters/ Time intervals (months) / Storage condition (4 ± 0.5°C) / Storage condition (22 ± 1°C)0 / 1 / 3 / 6 / 1 / 3 / 6
Constriction indexa (%) / 0 / 0 / 2±0.58 / 3±0.45 / 0 / 0 / 1.02±0.28
Surface pH / 6.45±0.48 / 6.5±0.28 / 6.25±0.16 / 6.0±0.46 / 6.40±0.28 / 6.35±0.65 / 6.45±0.18
Film thickness (µm) / 740±30 / 740±20 / 735±28 / 738±25 / 742±10 / 738±20 / 736±25
Swelling index b (%) / 26.28±2.62 / 25.18±1.48 / 27.22±1.68 / 23.22±1.18 / 26.00±3.68 / 27.82±2.64 / 27.00±4.18
Drug content (%) / 97.30±2.08 / 98.10±1.06 / 96.15±1.52 / 95.12±3.26 / 98.00±2.68 / 99.00±0.68 / 97.00±1.45
Folding endurance (times) / 100±18 / 98±10 / 90±8 / 85±12 / 100±20 / 100±15 / 95±10
In vitro residence time (min) / 90±5.00 / 94±3.00 / 84±8.00 / 80±10.00 / 92±8.00 / 94±6.00 / 96±7.50
Buccoadhesive strengthc / 28.8±1.37 / 26.18±2.37 / 24.34±3.72 / 20.34±6.20 / 28.0±2.44 / 28.1±2.40 / 26.8±1.52
Film strengthd / 320±8.5 / 310±9.5 / 280±6.5 / 270±4.55 / 325±10.5 / 316±9.15 / 322±6.8
Tensile strengthe / 260±6.88 / 270±5.18 / 250±4.32 / 220±6.12 / 255±8.45 / 265±5.78 / 260.8±5.24
Mean ± SEM, n = 3
aZero percent constriction implies 100% flatness.
bSwelling index was determined after 4 h of exposure of the film in phosphate buffer medium.
cDual extrusion cell (probe-TA-DEC); fixture base table (TA-BT KIT); compression mode.
dCylindrical probe-3 mm (probe-TA 42); film support fixture (TA-FSF); compression mode.
eDual grip assembly (probe-TA-DGA); tension mode.
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