Sodium Alginate in Mucoadhesive Drug Delivery Systems: A Comprehensive Review of Properties, Dosage Forms, And Characterization Methods

Mucoadhesion has evolved as a revolutionary advanced drug delivery approach with excellent advantages over all other conventional delivery methods owing to its capability of holding to defined sites for both local and systematic therapeutic action(1). The above property plays a vital role particularly concerning local retention of formulations in muco-ciliated regions that can lead to augmenting their bioavailability and therapeutically effective delivery(2). Mucoadhesive drug delivery systems (MDDS) are especially advantageous for drugs that require localized delivery or for therapeutic agents with low solubility and stability(3). Recent advancements in mucoadhesive systems have led to the development of a variety of dosage forms, including tablets(4–8), gels(7,9–11), films(12–15), patches(14,16–18), and microspheres(19–21), which can be applied to various mucosal sites such as the gastrointestinal (GI) tract, nasal passages, and oral cavity(22). These systems not only improve patient compliance but also provide triggering and sustained release features, prolonging therapeutic effects while minimizing the risk of adverse effects compared to traditional dosage forms(2). Sodium alginate, extracted from brown seaweed, this bio-sourced, biocompatible, and non-toxic polysaccharide is capable of forming gels(10). Sodium alginate can be classified based on viscosity (approximately 200 mPa·s) and a molecular weight range of 10,000 to 400,000 g/mole, allowing for modifications tailored to specific drug delivery applications. Its ability to form hydrogels, beads, and films makes it versatile for various mucoadhesive drug delivery systems(4,6–8,11,14,15,23–26). In particular, the formation of gels in the presence of divalent cations such as calcium enhances the stability and efficacy of drug formulations(27). Novel drug delivery methods utilizing sodium alginate have opened new avenues for preparing diverse dosage forms, including buccal tablets, nasal sprays, ocular inserts, and patches. For example, buccal tablets is for painless administration directly into the mouth, bypassing hepatic metabolism, while nasal sprays enable rapid absorption through the mucous membrane. Ocular inserts, designed for slow-release applications, are particularly effective for treating eye-related disorders. The versatility of sodium alginate in forming various dosage forms significantly enhances the efficacy of mucoadhesive drug delivery system(4,6–8,11,14,15,23–26). This review aims to explore the mucoadhesive properties of all-natural polymers, with a specific focus on sodium alginate as a standard reference. It will examine various dosage forms, including conventional ones such as tablets, films, beads, and gels, while considering different mucosal areas(22). The evaluation section will classify characterization methods into four categories: in vivo, in vitro, ex vivo, and advanced techniques, each explained with accompanying diagrams. Notably, this review will detail both static and dynamic mucoadhesive properties, which are crucial for determining the strength and sustained effect of dosage forms. By synthesizing current knowledge and highlighting the unique aspects of sodium alginate-based mucoadhesive systems, this review aims to provide a comprehensive understanding of their potential in enhancing drug delivery efficacy compared to existing literature(28).

Mucoadhesion process

Mucin is a glycoprotein which covers mammals different body part as a protector and lubricator, basically made up with glucose and protein subunit. Mucin acts as a barrier to the surface of epithelium and also acts to remove the extruder. Mucus gland and goblet cells are responsible for mucin secretion, which contains water (95% w/w), mucin (0. 2 to 5. 0% w/v), globular proteins (0. 5% w/v), salts (0. 5 to 1. 0% w/w), lipids (1–2% w/w), DNA, cells and cellular debris. Mucins (10–40 MDa) are polymeric gel-forming glycoproteins secreted by epithelial goblet cells and submucosal glands . Mucin fibers are filamentous O-linked glycoproteins with ‘PTS’ (proline, threonine, and serine) repeated domain which is highly glycosylated and found to have a carbohydrate density of more than 70 percent. It is mainly with N-acetyl galactosamine (GalNac), N-acetyl glucosamine (GlcNac), fucose, galactose (Gal) and sialic acid and only slightly with mannose and sulfate Mucins because they are rich products of glycosylation are situated in the form of bristles(29). Within the secretory glands, high levels of calcium ions assist in muco-thinning by neutralizing the negatively charged sulfate and sialic acid groups. The solubilization of mucins also causes some spectacular changes in volume, increasing by more than 500 times. Further, the steric hindrance by the O-linked GalNac residues with protein core is also seen to account for the broader mucin conformation. In the secretion cavities of the secretory glands, calcium ions help in the neutralization of the negative charges of the sulfate and sialic acid groups which promote the condensation of mucin. One of the key features of mucins is that, after secretion, they increase in volume many folds, to be more precise, 500 times and more. Furthermore, it is the steric impact involving the O-linked GalNac residues pared with the protein core in explaining mucin’s large and expanded structure in figure 1. In addition, the PTS-domains are flanked by hydrophobic globular regions with a rich content of cysteine that seem to form intramolecular disulphide bonds as part of the longer linear oligomers that gives the mucus its adhesive characteristics as well as its swellable nature. At acidic pH, the mucins shed their random coil structure and; gain an extended conformation as well as a gel phase in the mucus. These conformational changes were intended to allow cross-links between mucin macromolecules owing to hydrophobic interactions at a low pH in transition sol to gel state. Also, fluidity can be regulated by the change in concentration of ions, for instance it has been showed that calcium ions can enhance aggregation of mucins into large linear or branched macromolecular structures in figure 1. Therefore, attributes such as composition, pH, ionic strength, and conformation are essential in the assembly as well as the role and physical characteristics of mucus . Mucus controls the passage of molecules and particles by several mechanisms suggested as size restriction, hydrogen bonding, charge and hydrophobicity interactions and other types of molecular binding interactions(30).

       
           
       
    Figure 2: Theories of mucoadhesion

Mainly six theories are there that explains Mucoadhesion phenomenon figure 2.(53) They are

Where:

• L: Depth of interdiffusion, t: Contact time, Db: Diffusion coefficient, SAB: Spreading coefficient ?A, ?B, ?AB: Surface energies of mucoadhesive, biological surface, and their interface, WA: Work of adhesion , F: Friction force, ?: Coefficient of friction, N: Normal force, ?: Fracture strength, Fm: Maximum detachment force, A0: Initial overlap contact area, Wa: Work of adhesion, ?: Elongation at point of detachment(22,40,44,46,54–59)

3.Mucoadhesive properties of Sodium Alginate

Sodium alginate is a water-soluble, biodegradable natural polysaccharide composed mainly of mannuronic and guluronic acids. It's widely used in the pharmaceutical industry due to its ability to gel through interaction with calcium ions, undergoing ionotropic gelation. This property allows sodium alginate to be used in various dosage forms, including mucoadhesive systems, microspheres, microcapsules, tablets, and sutures, with controlled release capabilities(60). As an anionic linear polysaccharide, sodium alginate consists of ?-D-mannuronic acid (M) and ?-L-guluronic acid (G) connected by 1,4-glycosidic bonds. These can form homopolymer or heteropolymer blocks, with acetyl groups distributed throughout the chain. Mucoadhesion occurs through physical and chemical interactions between sodium alginate's COO- groups and mucin's NH3+ groups, primarily via electrostatic interaction or hydrogen bonding. Sodium alginate's molecular mass ranges from 12,000 to 180,000 Da, influencing its hypoglycemic and hypocholesterolemic effects. Alginates with molecular weights of 50 kDa and above have shown potential in preventing obesity and diabetes-like conditions. At a pH of approximately 4, carboxyl groups are neutralized to COO- sodium anions, enabling cross-linking with calcium ions for drug encapsulation. This process is fundamental in developing new dosage forms. Sodium alginate's ability to chelate metal cations, particularly calcium ions, is valuable in producing microcapsules for drugs, food, and biotechnological products. Its non-toxic and non-antibody forming properties make it an ideal material for pharmaceutical formulations(60). Now, to determine the mucoadhesivity property, all the evaluation methods we can major classify as static or dynamic. Based on the name we can easily described that if the evaluation test procedure is fixed means if the system (dosage form attached with mucoadhesive media that means beads attached with excised intestinal goat tissue) is not moving and remain in the tissue surface then it can be termed as static method and if the test procedure is not fixed means if the tissue is moving with then the method is termed as dynamic method. Depending on the instrument used it characterized accordingly explained in table 2 and figure 3. So, we are dividing this into static and dynamic because in our body their physiological difference is there. Means in GIT we have peristalsis movement so we take it dynamic method and in different part of the body causing different mucociliary clearance level or different mucin turnover  rate 5ml/ minute for stomach(peristalsis propagation rate 8cm/s)(61). And if we are applying rectal mucoadhesive tablet mucus turn over rate 1hr and no additional movement then there was taken as static method. (62).

         
           
       
Figure 3: Static and Dynamic method of mucoadhesion

2. Comparison of Different Dosages forms

Tablets have shown promising results in various studies. Pawar et al. (2018) developed bilayer mucoadhesive tablets of Pantoprazole sodium using Carbopol® 974P, HPMC K4M CR, and sodium alginate, demonstrating mucoadhesive properties lasting up to 8 hours. Similarly, Bartoníková et al. (2024) prepared vaginal tablets using sodium alginate as the main mucoadhesive agent, achieving drug release retention for up to 10 hours. Bakr et al. (2022) formulated buccal tablets of Labetalol Hydrochloride, finding that sodium alginate-containing formulations showed better in-vitro drug release and ex-vivo drug permeation rates compared to Carbopol-934, with an ex-vivo residence time of around 8 hours. Ghadge et al. (2023) developed mucoadhesive buccal tablets of eletriptan hydrobromide using HPMC K4M and sodium alginate, achieving 97.43% drug release over 8 hours with good mucoadhesive properties. Comparing these tablet formulations, we can observe that sodium alginate consistently performs well as a mucoadhesive agent, with residence times ranging from 8 to 10 hours across different studies. This suggests that sodium alginate-based tablets may offer superior mucoadhesion compared to other polymers like Carbopol. Gels offer unique advantages in terms of ease of application and intimate contact with mucosal surfaces. Samani et al. prepared Nystatin-loaded alginate microparticles incorporated into a Carbopol 934 gel, demonstrating that the presence of alginate increased the mucoadhesive force from 1.88 ± 0.04 g/cm2 to 1.94 ± 0.15 g/cm2. Dias et al. and Mali et al. formulated thermoreversible mucoadhesive nasal gels of Metoclopramide Hydrochloride using Poloxamer 407 (PF127) and mucoadhesive polymers, with mucoadhesive forces ranging from 466 to 781 dyne/cm2. Yuan et al. prepared thermosensitive and mucoadhesive rectal gels of Nimesulide, demonstrating that mucoadhesive strength increased with polymer concentration. Comparing these gel formulations, we can see a wide range of mucoadhesive forces, from 1.94 g/cm2 to approximately 7.81 g/cm2 (converting 781 dyne/cm2). This variation highlights the importance of polymer selection and concentration in gel formulations. Thermoreversible gels, in particular, show promising results with higher mucoadhesive forces, suggesting they may be more suitable for certain applications requiring stronger adhesion.

Beads offer the advantage of being easily dispersed and providing a large surface area for adhesion. Nayak et al. synthesized glimepiride-loaded alginate-ispaghula beads, which showed 66-70?herence to goat intestinal mucosal tissue in acidic conditions and 38-43% in phosphate buffer. Another study on Piroxicam-loaded alginate-pectin beads demonstrated mucoadhesive strengths between 3.29-6.56 g/cm2 and mucoadhesive times of 180-480 minutes. Nizatidine-loaded alginate-chitosan beads showed a maximum mucoadhesive strength of about 91% in an in vitro wash-off test. Pal et al. prepared metformin HCl-loaded tamarind seed polysaccharide-alginate beads, which exhibited stronger mucoadhesion in gastric pH compared to intestinal pH. Comparing these bead formulations, we observe that mucoadhesive strength varies significantly depending on the polymer composition and pH conditions. The alginate-chitosan beads showed the highest mucoadhesive strength (91%), suggesting that this combination may be particularly effective. The pH-dependent behavior of beads is a notable feature, with stronger adhesion in acidic conditions, making them potentially suitable for gastric-targeted delivery systems. Films offer the advantage of a large surface area for adhesion and easy application. Pamlényi et al. developed buccal films containing cetirizine dihydrochloride with mucoadhesion forces ranging from 8.41 to 9.1 N. Wang et al. created chitosan-sodium alginate-ethyl cellulose films with a bioadhesion time of approximately 120 seconds. Another study on buccal films showed that films with 2% sodium alginate concentration had 7.44 ± 0.23 N adhesion force, while 3% sodium alginate films approached 18 N. Silvestre et al. prepared sodium alginate films with a maximum mucoadhesion force of 3N. Comparing these film formulations, we can see that mucoadhesive forces range from 3 N to 18 N, with sodium alginate concentration playing a crucial role in adhesion strength. The bioadhesion time of 120 seconds reported by Wang et al. is notably shorter than the residence times observed for tablets and beads, suggesting that while films may offer strong initial adhesion, their duration of adhesion might be shorter. This could make films more suitable for rapid-onset, short-duration drug delivery applications. When comparing these different mucoadhesive formulations, several trends emerge. Sodium alginate, HPMC, and Carbopol are commonly used across various formulations due to their excellent mucoadhesive properties. The combination of multiple polymers often leads to synergistic effects in mucoadhesion. Each formulation type has its advantages: tablets and beads offer precise dosing and ease of administration, with tablets showing the longest mucoadhesion times (8-10 hours); gels provide intimate contact with mucosal surfaces and are easily applied, with mucoadhesive forces ranging from 1.94 to 7.81 g/cm2; films offer a large surface area for adhesion and can be suitable for local drug delivery in the oral cavity, demonstrating the highest mucoadhesive forces (up to 18 N) but potentially shorter adhesion durations(4–8). The mucoadhesive strength varies widely depending on the formulation and testing method. Generally, higher polymer concentrations lead to increased mucoadhesive strength. Films and gels tend to show higher mucoadhesive strengths compared to beads and tablets, likely due to their larger contact surface area. Most formulations demonstrated adhesion times ranging from 8 to 12 hours, which is sufficient for once or twice-daily dosing regimens, with tablets showing the longest durations. Several studies noted that mucoadhesion was stronger in acidic conditions compared to neutral or alkaline environments, which can be advantageous for gastric-targeted delivery systems, particularly evident in bead formulations. Mucoadhesive formulations generally showed sustained drug release profiles, with many achieving controlled release over 8-12 hours, making them suitable for various therapeutic applications requiring prolonged drug delivery(4–8).

When comparing these different mucoadhesive formulations, several trends emerge. Sodium alginate, HPMC, and Carbopol are commonly used across various formulations due to their excellent mucoadhesive properties. The combination of multiple polymers often leads to synergistic effects in mucoadhesion. Each formulation type has its advantages: tablets and beads offer precise dosing and ease of administration; gels provide intimate contact with mucosal surfaces and are easily applied; films offer a large surface area for adhesion and can be suitable for local drug delivery in the oral cavity(4,9,10,14,18,22,63,64). Mucoadhesive formulations offer significant quantitative advantages over non-mucoadhesive formulations. They can adhere to mucosal surfaces for 8-12 hours, a 4-6-fold increase compared to non-mucoadhesive formulations, allowing for sustained drug release and improved absorption. Studies have shown that mucoadhesive formulations can increase drug bioavailability by 1.5-3 times, with a buccal mucoadhesive tablet of propranolol demonstrating a 2.4-fold increase compared to an oral tablet. The prolonged release profile can reduce dosing frequency by 50-75%, from 3-4 times daily to once or twice daily, significantly improving patient compliance. Some mucoadhesive polymers have shown to increase drug permeation by 2-5 times, with chitosan-based formulations demonstrating a 3.7-fold increase in insulin permeation across intestinal epithelium. Mucoadhesive formulations can achieve 5-10 times higher local drug concentrations and modulate drug release over 8-12 hours, compared to non-mucoadhesive immediate-release formulations that typically release the drug within 30 minutes to 2 hours. Buccal and sublingual mucoadhesive formulations can bypass hepatic first-pass metabolism, potentially improving bioavailability by 20-80% for drugs that undergo extensive hepatic metabolism. Additionally, the controlled release properties can reduce inter- and intra-patient variability in drug absorption by 30-50%, leading to more predictable therapeutic outcomes. These quantitative advantages demonstrate the significant potential of mucoadhesive formulations to improve drug delivery efficiency, patient compliance, and overall therapeutic outcomes compared to traditional non-mucoadhesive formulations(4–10,11,13–16,48,62,64).

5. Evaluation methods of mucoadhesion property

5.1 In vitro method

1.Method based on tensile strength: As earlier highlighted in both of these methods the force used which is utilized in peeling a model membrane off a test polymer is measured figure 4 (14).

1.1 Tensiometer: This instrument comprises of two jaws that are made from flat glass. The upper one is non-moving while the lower one is in contact with a screw-facilitated elevating surface. The upper fixed glass is placed on a high precision weighing machine. The adhesive tablets are then applied on the lower glass surface then the surface is brought upwards to make contact with the upper glass surface. The lower glass is subsequently lowered so that the tablet is clearly seen as pulled off from the upper glass. The maximum tensile force is computed and noted down in dyne/cm² typical range: 0.1 - 100 dyne/cm?2; (0.01 - 10 N/m?2;) and Ideal range for mucoadhesion  40 - 100 dyne/cm?2; (4 - 10 N/m?2;)(25).

1.2 Modified balance method: The apparatus that used as the Bioadhesion test is the modified double beam physical balance. The mucus membrane is sutured with mucosal side up, using the thread over Teflon block. The fixed weight which is kept on the right side of the pan was removed to raise the balance beam. From this, the Teflon cylinder together with the tablet came down over the mucosa. The extra weight on the right-hand side indicated the Bioadhesive strength of the tablet in grams Typical range: 1 - 200 grams and ideal range for mucoadhesion is between 50 - 150 grams (66).

1.3 Microbalance Methods: The microforce balance technique is employed to the quantitative measurement of adhesion force in particulate systems.  This necessarily involves a microtensiometer and a microforce balance, with the result giving both the contact angle and the surface tension.  Tissues of mucous membrane are sitting in small mobile place where even pH and physiological temperature is regulated.  The size of microsphere is one of the most critical features of the analysed material, while microsphere size is a property individually connected through a specific amount of the thread to the microbalance.  Another apparatus is consisting of a suitable chamber mounted to hold a mucous membrane which is lifted up to come in contact with the microsphere and after a definite period, it is again lowered back to its position, it measured in micro gram Typical range: 1 - 1000 ?g and ideal range for mucoadhesion is between 500 - 1000 ?g.(53). 

2.Method for determining shear stress: The shear stress method of mucoadhesive systems is the force necessary to remove a mucoadhesive from a mucosal surface or mu?in coated substrate by sliding or shearing it. The process involves applying the mucoadhesive material on the surface and preserving the process for a certain number of hours/minutes. A texture analyzer or shear testing apparatus is then used to apply a lateral force on the material until it peels off. Shear stress is determined from the equation; ? = F / A where F is the total shear force, A is the contact area. This technique also enables on determination of the mucoadhesive strength with dyne/cm² or N/m² of the material. Higher shear stress value being associated with better adhesion Typical range: 100 - 40,000 dyne/cm?2; (10 - 4000 N/m?2;) and ideal range for mucoadhesion is between 10,000 - 30,000 dyne/cm?2; (1000 - 3000 N/m?2;).(67)

3. Detachment force method: In the detachment force method of mucoadhesion test, mucoadhesive strength of the material is determined by the force needed to delaminate the material from the mucosal surface or a mucin-coated substrate. In this method a small quantity of the mucoadhesive material is placed directly on the mucosa or a layer of mucoadhesive and allowed to adhere for a predetermined time. They place the material under a force that is applied by a testing equipment like the texture analyzer or a locally-developed tester until the two interfaces debond. Depending on the mucoadhesive strength, the force which is being measured at this point is the force at which the film separated from the mucosa. This method is commonly applied in the assessment of mucoadhesive drug delivery systems measured by N. Typical range: 0.1 - 5 N Ideal range for mucoadhesion: 0.5 - 3 N (68)

4.Falling film method: It was established that there are two modes of particle adhesion: Flow over the surface and ratio of the adhering part of the particles to the overall number of particles on the tissue surface.  Where quantification can be done by the aid of coultercurrent such as incubation flow of coultercurrent sucrose gel.  This is a quantitative technique that is mainly done in the research laboratory. Typical range: 20 - 90?hesion Ideal range for mucoadhesion: >80?hesion (53). 

5.Wash off method: Wash-off method for evaluating mucoadhesion of a mucoadhesive material determines the degree of the material made compact with a mucosal surface after subjecting the system for a subsequent washing process. In this method, the mucoadhesive material, beads is firstly placed on the mucosal tissue or surface with the mucin coating and is allowed to attach for a specific time. next, the surface is then allowed to go through a controlled washing using a simulated physiological media such as PBS. The number of beads still attached to the surface after washing is then determined. This counting can be done by gravimetrically. Basically this measurement designed for beads type dosage form, whose ability to resist itself from being washed off is further calculated by counting the number. Typical range: 50 - 95% retention after washing Ideal range for mucoadhesion: >80% retention after washing (67)

6. Colloidal gold staining: Interaction with the adhesive particles is to be seen based on the red colour formed on the mucin covered surface.  Red-colloidal gold particles are used in this experiment and the mucin molecule forms complexes with the gold particles.  Then, bioadhesive hydrogels get a red colour on the Surface and it did not aggregate with mucin-gold conjugates(69). 

7. Adhesion no. determination method: This is the proportion of the particles that forms a solid layer on the substrate to the overall particles that were used in a particular experiment.  It is normally expressed in terms of a percentage (%)Typical range: 50 - 95% retention after washing Ideal range for mucoadhesion: >80% retention after washing (69). 

8.Viscosity determination methods: Hassan and Gallo developed this experiment by using viscometer, in this test bioadhesive bond strength of mucin-polymer, i e resistance to flow, was determined.  Their viscosity was further determined using a Brookefield viscometer both in its unadulterated form and after incorporating some selected non-ionic, anionic and cationic polymers measured in centipoise or pascal/s. Typical range: 10 - 100,000 cP (0.01 - 100 Pa·s) Ideal range for mucoadhesion: 5,000 - 50,000 cP (5 - 50 Pa·s) (70). 

9.Drug permeation method: The drug permeation method is used generally to assess the extent of mucoadhesion in a mucoadhesive material as far as the enhancement of or sustained drug delivery through a mucosal membrane is concerned. In this approach, a mucoadhesive formulation is administered to a mucosal membrane or tissue including artificial mucosal layer or biological mucosal layer. This configuration generally involves the use of the diffusion cell or any other related system where the mucoadhesive material is placed at one side of the membrane and the drug solution is placed at the other side. At different time points samples are taken from the receptor compartment so as to determine how much of the active drug gets to cross the mucosal barrier. This method determines the degree of improvement the mucoadhesive material has on the transport of the drug across the mucosa which helps to evaluate the material as a drug delivery system. Design was similar to Franz diffusion cell, and drug concentration determined by spectrophotometrically. Then plot amount of drug permeate with time was measured. Typical range: 1 - 100 ?g/cm?2;/h Ideal range depends on the specific drug and application(65)

10.Fluroscence probe method: In this method the membrane lipid bilayer was labelled by pyrene and membrane proteins by adhesion wet). Occasionally the cells were combined with the mucoadhesive agents and changes in fluorescence spectra were observed. This provided a direct feedback of polymer binding and how it affects polymer adhesion. (69).

11.Adhesion determination method / Mucoretentibility study: The adhesion determination method or mucoretentibility study investigates the extent to which a mucoadhesive material will be attached to a mucosal surface at a given time in precise known periods of time. This technique is carried out through applying the mucoadhesive material on a mucosal tissue or a surface that is coated with mucin, and the efficiency of the material is then tested under conditions that simulate physiological conditions. Usually, the mechanical interaction consists of rinsing or rubbing of the material to expose it to physiological movements and forces. The degree of material hold back is then determined, usually through observation method, determination of weight if the size of the particle is larger, or colorimeter test if the size of the particle is very small. In this method the ability of the material to make and sustain contact with mucosal surfaces which is very vital in drug delivery systems. Typical range: 40 - 95% retention over time Ideal range for mucoadhesion: >80% retention over the desired time period (71)

12.Surface pH study methods: The surface pH study method involves determination of pH on the surface of the mucoadhesive material to check its compatibility. In this method, a small amount of the mucoadhesive material is brought into contact with a pH – sensitive electrode or probe that is help to measure the pH at the surface of the material. This is usually done either by positioning the electrode onto the sample or by dipping the material into a small volume of the buffer solution and letting the pH electrode stabilize. The actual reading of the pH of the surface is important to avoid having the mucoadhesive material elicit some adverse effects on the mucosal tissues. Typical range: pH 4 - 8 Ideal range for mucoadhesion: pH 5.5 - 7.0(57)

13.Scanning electron microscopy: Scanning electron microscopy (SEM) is a valuable technique for mucoadhesion studies as it provides high-resolution images of the surface morphology of mucoadhesive materials and their interaction with mucosal tissues. In this method, the mucoadhesive material is applied to a mucosal surface, and after a specified adhesion period, the sample is prepared and coated with a conductive layer if necessary. SEM is then used to examine the surface of both the material and the mucosal tissue at a microscopic level, revealing details such as surface texture, adhesion points, and the extent of interaction between the mucoadhesive and mucosal surface. This technique helps in understanding the physical mechanisms underlying mucoadhesion and optimizing material formulations.(22)

14.Texture analyzer method: The evaluation of the value of the rupture tensile strength is made by equipment that is called: Texture analyst or universal testing machinery. Therefore, in addition to rupture tensile strength this method helps to define the texture of the formulations and other mechanical characteristic values of the system. Here, it will be described by the force necessary to remove the formulation from the model membrane which might be a disc obtained from mucin or piece of animal mucous membrane commonly porcine nasal turbinate or rat intestinal mucus. Typical range: 0.1 - 10 N Ideal range for mucoadhesion: 0.5 - 3 N(69).

15.Adhesion wet method: The adhesion wet method for mucoadhesion testing involves applying a mucoadhesive material to a mucosal surface or a mucin-coated substrate and then measuring the force required to detach it. The procedure includes preparing the mucosal surface (either actual tissue or a mucin layer), applying the mucoadhesive material, and using an apparatus to measure the detachment force. This force indicates the strength of the mucoadhesive bond. Typical range: 0.1 - 5 N Ideal range for mucoadhesion: 0.5 - 3 N(56)

16.Flow channel method: The flow channel method is used to characterize the static and dynamic behaviours of a bioadhesive polymer particle placed on the mucin gel at different times using a camera for the determination of its adhesive property Typical range: 20 - 90?hesion over time Ideal range for mucoadhesion: >80?hesion over the desired time period(70).

17.Mechanical spectroscopic method: Mechanical spectroscopic method used a mechanical technique known as spectrometry to ascertain how glycoprotein and polyacrylic acid reacted when mixed with a gel and to evaluate how their interaction was affected by factors such as pH and polymer chain size(70).

        
           
       
Figure 4: In vitro methods for evaluation of mucoadhesion property

1. Radio isotope method using gamma scintigraphy method: This technique provides information of oral dosage forms in the various parts of GI tract, time and place of disintegration of the dosage forms, place of drug absorption, the impact of food, disease and size of the dosage form on the in-vivo quality of the dosage forms. The main advantage of gamma scintigraphy over radiological examinations is the possibility to visualize the time-course of the formula’s passage through the GIT with a relatively low dose of irradiation to the subjects. Since microspheres after oral administration arrived at the mentioned location, the degree of entry into the GIT, distribution, and retention time of the mucoadhesive microspheres in the GIT system can be determined with GSC. Typical range: 30-90% retention after 4-6 hours Ideal range: >70% retention after 6 hours(69).

2 X-ray studies: Barium sulphate loaded tablet was used to assess the bioadhesive character of the natural polymer and the mean residence time in the stomach. Two healthy rabbits looking grossly normal with a body weight of 2.5 kg are selected and the tablet is given to them orally. X-ray photograph is taken after a definite amount of time or periods have elapsed(69) figure 6 Typical range: 2-8 hours gastric retention time Ideal range: 4-6 hours gastric retention time.

3.In vivo evaluation of mucoadhesive study: In vivo evaluation of mucoadhesive properties involves assessing the performance of mucoadhesive materials directly within living organisms to determine their effectiveness in adhering to mucosal surfaces under physiological conditions. This method typically involves applying the mucoadhesive material to specific mucosal sites in animal models or human subjects, such as the gastrointestinal tract or buccal cavity. After application, the retention time and behaviour of the material are monitored through various means, such as imaging techniques or direct observation, to evaluate how well it adheres and remains in place. This approach provides valuable insights into the material's real-world performance, safety, and potential efficacy for drug delivery systems. Typical range: 2-12 hours retention time Ideal range: >6 hours retention time(16).

4.Isolated loop technique (Rat Gut loop studies): Male Wistar rats weighing about 300 g are anesthetized and sacrificed by cervical dislocation The suspension is filled lite is removed and into of small intestine (about 15 cm in length) and sealed. These tubes are then allowed to incubate in saline at 37°C for 1 hr After this they are sphere and the number of percentages(69).

From the difference of the counts to the tissue is calculated from the magnetic resonance imaging and Fluorescence detection.(72)

5.Use of pharmacoscintigraphy: As a tool to study drug delivery to the eye, Gamma-scintigraphy offers information on deposition, dispersion of the formulation and its ‘movement’. Together with this kind of studies the determination of drug concentrations in blood or urine samples, pharmacoscintigraphy yields data regarding to the sites of drug release and absorption. Typical range: 20-80% retention after 1 hour Ideal range: >60% retention after 1 hour(73)

6. Magnetic resonace imaging and fluorescence detection: Magnetic resonance imaging (MRI) has proven to be a non-invasive technique is easily accessible for in visualization and localization of solid oral dosage forms in the rat gastrointestinal tract. Compared to the other imaging techniques MRI enables depiction of the different contrasts of the anatomical structures with higher resolution.(74)

7. Quantitative GIT distribution fluorescence microscopy: Using fluorescence microscopy investigations were carried out on distribution and penetration of microsphere formulations. GIT tissue sections prepared for the experiment were blotted with tissue paper after the excision. Altogether, the findings of quantitative GI distribution study presented higher percentage of retention of mucoadhesive microspheres lodged in the upper GIT Typical range: 20-80% retention in upper GIT Ideal range: >60% retention in upper GIT.(75)