Introduction

Harsh gastric environment and first-pass metabolism are the two major reasons for reduced bioavailability of the drugs through oral route. Peptide/protein drugs are especially vulnerable to the oral route of delivery due to obvious physiological reasons. Second, oral route requires multiple dosage administration, which is cumbersome for the patients. Parenteral route, on the other hand, is invasive and is not patient compliant. It is associated with acute pain and local site morbidity. Second, parenteral route is especially troubling for the patients who need regular injections such as those suffering from insulin-dependent diabetes or those requiring certain other regular injectable drugs such as glucagon-like peptide-1 (GLP-1) analog and amylin [Reference Shojaei1, Reference Bagan, Paderni, Termine, Campisi, Lo Russo and Di Fede2]. Swallowing tablet or capsule is usually problematic for dysphagic patients, geriatric people, and pediatric population. Children and geriatric patients very often are particular about the taste of the formulation they are consuming. Dysphagic patients, irrespective of age, are very prone to aspiration of the drug formulation, which might lead to another medical emergency [Reference Stegemann, Gosch and Breitkreutz3, Reference Mennella, Spector, Reed and Coldwell4]. Therefore, to avoid such difficulties and to reach patient compliance, alternative routes of drug administration have been explored in last few years. Buccal and sublingual routes are most patient complaint and guarantee a completely noninvasive route of delivery.

Transbuccal drug delivery has gained a lot of attention and importance in last few years for its easy accessibility and high vascularity. High vascularity ensures systemic absorption of the drug without encountering any harsh physiological environment and first-pass metabolism. These features ensure a high bioavailability as compared to the most common oral route. The oral mucosa is composed of stratified squamous epithelium (outermost layer) followed by basal membrane below which is the innermost layer called lamina propria. The buccal mucosa epithelium is almost 40–50 cell layers thick while the sublingual mucosa has lesser layers. The thickness of the buccal mucosa is reported to be 500–800 μm. Since the buccal and sublingual mucosae do not experience mechanical stress, they have nonkeratinized epithelia as opposed to that of gingiva and hard palate which are keratinized. The high vascularity of buccal mucosa is a great attraction for transmucosal drug delivery since the blood vessels directly drain into the internal jugular vein, enabling fast systemic delivery [Reference Barua, Kim, Jo, Seo, Park, Lee, Yun, Oh and Lee5, Reference Sudhakar, Kuotsu and Bandyopadhyay6].

Presence of mucus is another key anatomical component of buccal mucosa that is an important requirement for any transbuccal drug formulation. The mucus is primarily composed of water (∼95–99%), but the macromolecular ingredient that provides the signature viscosity is a class of glycoprotein known as mucin that accounts for 1–5% of mucus. Mucins are large protein with a molecular weight ranging from 0.5 MDa to almost 20 MDa. These mucins are the key components that are responsible for making adhesion with the various mucoadhesive polymers used in mucosal drug delivery. The mucus in the oral cavity is secreted as a part of saliva by the salivary glands. It has been reported that almost 70% of mucins in saliva are secreted by the minor salivary glands. Saliva is mostly water with around 1% inorganic and organic components including a few enzymes. The salivary pH range is from 5.5 to 7 and that depends upon the flow rate. At high flow rate, as reported, the bicarbonate and sodium level increase resulting in an increased pH toward 7. The mucin carries the overall negative charge at the physiological pH due to the presence of sialic acid moieties and sulfate residues that play an important role in mucoadhesion [Reference Madhav, Shakya, Shakya and Singh7]. Mucins, in terms of origin and presence, are classified into monomeric (attached to the epithelial surface) and oligomeric (secreted). The latter are responsible for imparting the rheological behavior to the saliva and mucus. The major mucin oligosaccharides present are N-acetylgalactosamine, N-acetylglucosamine, sialic acid, and fucose while sulfate groups are also present prominently. There are a total 19 mucin genes identified in humans (MUC) among which MUC2, MUC5AC, MUC5B, and MUC6 have cysteine-rich motifs in their N- and C-terminal domain that account for the oligomerization [Reference Thornton8, Reference Bansil and Turner9]. Linden et al. [Reference Linden, Sutton, Karlsson, Korolik and Mcguckin10] have reported the physiological presence of various mucins and their classification as secreted or cell surface attached. MUC2, MUC5AC, MUC5B, MUC6, and MUC19 are secretory and form gel while MUC7 is a secretory nongel-forming mucin. MUC1, MUC3A/B, MUC4, MUC12, MUC13, MUC15, MUC16, MUC17, and MUC20 are the cell surface–attached mucins. Among these mucins, MUC1 has been reported to be predominantly present on the buccal epithelium while MUC5B and MUC7 are the primarily secreted mucins in the saliva [Reference Zalewska, Zwierz and Gindzieñski11]. The daily saliva production in healthy humans is 1–1.5 L. The total stimulated saliva flow rate ranges from 1 to 3 mL/min while the saliva flow rate of unstimulated saliva is 0.25–0.35 mL/min. The normal salivary pH ranges from 6 to 7, but it changes as per the salivary flow (almost 5.8 during low flow and 7.8 during high flow) [Reference De Almeida, Grégio, Machado, De Lima and Azevedo12].

Porcine (pigs) are considered one of the most suitable animal models for in vitro and in vivo testing of mucoadhesive buccal formulation because of very close anatomical and physiological similarity to that of human buccal mucosa. Kulkarni et al. [Reference Kulkarni, Mahalingam, Pather, Li and Jasti13] have reported that the porcine buccal mucosa resembles human counterpart in ultrastructure (nonkeratinized), enzyme localization and activity, and permeability. Since human buccal samples are not easily available for in vitro and in vivo testing, porcine mucosa remains the best feasible model due to easy availability. Apart from pig, other large animals that have been used for buccal drug formulation testing are monkey, apes, and dogs due to highly similar structure and function of the respective tissue. For initial in vitro studies, the buccal tissue needs to be harvested and the animals need to be sacrificed for the same. The processing of the buccal tissue after harvesting is even tedious [Reference Janet Hoogstraate and Bodd14]. Cui et al. [Reference Cui and Mumper15], however, reported in one of their articles that the buccal mucosa of rabbit is very similar to that of humans among all the laboratory rodents commonly available. In an important article by Thirion-Delalande et al. [Reference Thirion-Delalande, Gervais, Fisch, Cuiné, Baron-Bodo, Moingeon and Mascarrel16], the team mentioned that the rabbit buccal mucosa is mostly nonkeratinized similar to that of humans. The buccal epithelium of rabbit is 30–40 layers thick which is almost exact as that of monkeys and very similar to that of humans (40–50 layers as mentioned previously). Even the connective tissue density and vascularization were similar to that of monkeys. Rabbits are more accessible than dogs and monkeys when comparing the availability of large and small animals. But the primary problem and question that remain here is sacrificing the animal even for simple in vitro studies such as mucoadhesion.

To address the issue of difficult in access to the suitable animal for mucoadhesion study and further preparation of excised buccal tissue, synthetic polymeric structures were fabricated to mimic the buccal mucosa to test the mucoadhesion of drug formulations. In this regard, as per our literature search, Khutoryanskiy et al. [Reference Hall, Khutoryanskaya and Khutoryanskiy17, Reference Cook, Smith and Khutoryanskiy18, Reference da Silva, Khutoryanskiy, Bruschi and Cook19] have carried out a considerable amount of work. They fabricated mucosa mimetic-surface by two initial approaches viz. layer-by-layer deposition of interpolymeric complex of methylcellulose and polyacrylic acid (PAA) on a chemically modified glass surface at a temperature around 120 °C and copolymerization of 2-hydroxyethyl methacrylate (HEMA) with other polymers such as N-vinylpyrrolidone, sorbitol methacrylate, N-acryloyl-glucosamine (AGA), and 2-hydroxyethyl acrylate. When compared with a natural mucosal tissue, the complex hydrogel of HEMA-AGA rendered almost similar mucoadhesion to the drug formulation. Apart from developing a complete polymer-based mucosa-mimetic material, Jacobsen et al. [Reference Jacobsen, van Deurs, Pedersen and Rassing20, Reference Jacobsen, Nielsen, Brøndum-Nielsen, Christensen, Olin, Tommerup and Rassing21] have developed filter-grown human buccal carcinoma cell line TR146.

Mucoadhesive polymers have been widely used to develop various mucosal drug delivery systems. Forms such as tablets, gels, nanoparticles, patches, and films have been synthesized and tested till date. Various natural, seminatural, and synthetic polymers have been widely validated to have strong mucoadhesive properties. Some commonly used polymers among the lot such as chitosan, alginate, PAA, sodium carboxymethyl cellulose (Na-CMC), hyaluronic acid (HA), and thiomers have shown promising characteristics to develop drug formulations [Reference Khutoryanskiy22, Reference Russo, Selmin, Baldassari, Gennari, Caviglioli, Cilurzo, Minghetti and Parodi23]. Chitosan oligosaccharide (COS) is the hydrolyzed product of chitosan with various properties different from that of chitosan. The most important aspect is its water solubility that was lacking with chitosan. COS has largely been an unexplored polymer with usage reports primarily as nanoparticles or as coating on other nanoparticles [Reference Muanprasat and Chatsudthipong24, Reference Zhang, Huang, Sun, Xing, Yamamoto and Gao25].

In this work reported, we have fabricated two novel things that, to the best of our knowledge, have not been reported till date. The first one is COS-based blended films as a proposed vehicle for buccal drug delivery while the second one is a very low-cost eggshell membrane (ESM)–based mucosa-mimetic surface using bovine submaxillary mucin (BSM). COS is a water-soluble version of chitosan which makes it easy to handle and is more appropriate than chitosan for biomedical applications. Second, we have utilized its reported mucoadhesive nature to formulate a blended patch that can be used to deliver drugs through buccal mucosa. Another attraction of the work is the fabrication of artificial buccal mucosa–mimetic surface using low-cost ESM which is normally a food waste. The mucosa-mimetic surface was fabricated using the commercially available BSM that mimicked the mucin present along the buccal mucosal surface. The COS-poly vinyl alcohol (PVA)–blended patch displayed much better mucoadhesive nature as than the COS and alginate blended patch.