The oral cavity performs several essential functions, enabling nutrient intake and speech; furthermore, it represents a primary route for drug administration. Although this route is readily accessible and non-invasive, the treatment of oral diseases remains challenging due to the complex and highly dynamic environment. In this setting, saliva flow, the swallowing reflex, and continuous exposure to external factors, such as food and beverages, rapidly clear applied formulations, significantly limiting their residence time at the site of action. In this study, on the basis of prior research performed in the same laboratory, liposomes were formulated and characterized with the aim of developing an oral spray for the treatment of xerostomia. Although xerostomia by itself is not a dangerous condition, it may lead to complications such as difficulty in swallowing, inflammation, infections, and dental caries. Liposomes were prepared via the thin-film hydration method employing Soy-phosphatidylcholine (Soy-PC) in combination with either dioleoyl-3-trimethylammonium-propane chloride salt (DOTAP) or Egg-Phosphatidylglycerol (Egg-PG) to produce positively and negatively charged liposomes, respectively. The liposomes were then coated with varying concentrations of alginate or chitosan, to identify the most stable formulation with the potential to enhance mucoadhesive properties. Furthermore, the influence of the hydration medium was evaluated by comparing a standard phosphate buffer (PB, pH 6.8) with a phosphate buffer containing 15% glycerol (PBG, pH 6.8) to exploit the intrinsic moisturizing and humectant properties of the latter. Also, the effect of adding a viscosity enhancer, specifically Hydroxyethyl cellulose (HEC) and Hydroxypropyl methylcellulose (HPMC) at different concentrations (0.5% w/v or 1% w/v), was evaluated. The formulations were characterized in terms of hydrodynamic diameter, polydispersity index (PDI), and Zeta potential immediately after preparation and over time to evaluate their stability. Preliminary studies were also conducted to assess the effect of viscosity changes on the sprayability and delivered dose of the final product at different temperature (4°C and 22°C). The results demonstrated that both positively and negatively charged liposomes, when coated with alginate and chitosan respectively, remained stable over a two-month evaluation period in both PB and PBG media. In contrast, uncoated liposomes tended to destabilize after around 4 weeks. Chitosan was selected for further development due to its superior mucoadhesive and antibacterial properties. Regarding the hydration medium, PBG (pH 6.8) was preferred as it leveraged the intrinsic humectant and lubricating properties of glycerol, which are essential for treating xerostomia, without compromising the colloidal integrity of the vesicles. The addition of viscosity enhancers resulted in a concentration-dependent increase in viscosity; however, this addition also induced modifications in the physicochemical liposomal characteristics that compromised their stability. HPMC 1% w/v emerged as the optimal candidate, providing the maximum increase in viscosity while maintaining a stable, homogeneous, and monodisperse population. This significantly higher viscosity was directly reflected in the sprayability tests, where HPMC 1% produced a more concentrated and controlled spray pattern compared to all other formulations. Notably, the delivered dose remained practically unchanged despite temperature variations between 4°C and 22°C, demonstrating the robustness and reliability of the final formulation under different storage conditions.
La cavità orale permette lo svolgimento di molteplici funzioni fisiologiche essenziali, tra cui l’assunzione dei nutrienti e l’articolazione del linguaggio, rappresentando al contempo una via preferenziale per la somministrazione di agenti terapeutici. Nonostante tale via risulti agevolmente accessibile e non invasiva, il trattamento delle patologie orali permane complesso. Ciò è dovuto principalmente alle peculiari condizioni dell’ambiente orale, dove il costante flusso salivare, il riflesso della deglutizione e l’esposizione a fattori esogeni determinano una rapida rimozione dei prodotti applicati, limitandone drasticamente il tempo di permanenza nel sito d’azione. Nel presente studio, sulla base di evidenze pregresse ottenute presso il medesimo laboratorio, sono stati formulati e caratterizzati sistemi liposomiali volti allo sviluppo di uno spray orale per il trattamento della xerostomia. Sebbene quest'ultima non costituisca di per sé una condizione patologica critica, essa può favorire l’insorgenza di complicanze quali disfagia, processi infiammatori, infezioni e carie dentali. I liposomi sono stati prodotti mediante la tecnica thin-film hydration, impiegando fosfatidilcolina di soia (Soy-PC) in associazione a 1,2-dioleoil-3-trimetilammonio-propano (DOTAP) o fosfatidilglicerolo d’uovo (Egg-PG) per l’ottenimento di liposomi dotati, rispettivamente, di carica superficiale positiva e negativa. Al fine di studiarne la stabilità, i sistemi sono stati rivestiti con polimeri (alginato o chitosano) a diverse concentrazioni (0.5% p/v o 1% p/v). Successivamente, è stata studiata l’influenza del mezzo di idratazione, confrontando un tampone fosfato standard (PB, pH 6.8) con un tampone fosfato addizionato di glicerolo al 15% (PBG, pH 6.8). Quest'ultimo è stato scelto per le sue proprietà umettanti. È stato altresì valutato l’impatto di agenti viscosizzanti, nello specifico idrossietilcellulosa (HEC) e idrossipropilmetilcellulosa (HPMC), a diverse concentrazioni (0,5% p/v o 1% p/v). La caratterizzazione ha previsto la determinazione del diametro idrodinamico, dell’indice di polidispersione (PDI) e del potenziale Zeta, sia in fase post-preparativa che durante studi di stabilità a lungo termine. Infine, sono state condotte indagini preliminari per valutare l'influenza della viscosità sulle proprietà di nebulizzazione e sull’uniformità della dose erogata a diverse temperature (4°C e 22°C). I risultati hanno evidenziato come i liposomi rivestiti con alginato o chitosano mantengano una stabilità ottimale per un arco temporale di due mesi in entrambi i mezzi di idratazione (PB e PBG). Al contrario, i sistemi non rivestiti hanno mostrato fenomeni di instabilità già dopo circa quattro settimane. Il chitosano è stato confermato come il polimero d'elezione per le finalità applicative del sistema, grazie alle sue proprietà mucoadesive e antibatteriche. Relativamente al mezzo di idratazione, il PBG è risultato preferibile, in quanto capace di coniugare l’azione lubrificante del glicerolo con il mantenimento dell’integrità colloidale dei sistemi liposomiali. L’incorporazione degli agenti viscosizzanti ha indotto un incremento della viscosità proporzionale alla concentrazione; tuttavia, tale processo ha comportato alterazioni strutturali che hanno compromesso la stabilità di alcune formulazioni. L'HPMC all'1% p/v è risultato il candidato ottimale, in quanto ha indotto il maggiore incremento della viscosità nella sospensione senza compromettere la stabilità colloidale del sistema, assicurando una distribuzione dimensionale omogenea dei liposomi. Tali proprietà si sono riflesse nei test di nebulizzazione, dove l'HPMC all'1% ha prodotto un pattern di spruzzo più concentrato ma comunque riproducibile. Inoltre, la dose erogata è risultata costante anche al variare della temperatura.
Preparazione e caratterizzazione di liposomi rivestiti da polimeri finalizzati ad alleviare i sintomi della Xerostomia
CALLONI, CHIARA MARIA
2024/2025
Abstract
The oral cavity performs several essential functions, enabling nutrient intake and speech; furthermore, it represents a primary route for drug administration. Although this route is readily accessible and non-invasive, the treatment of oral diseases remains challenging due to the complex and highly dynamic environment. In this setting, saliva flow, the swallowing reflex, and continuous exposure to external factors, such as food and beverages, rapidly clear applied formulations, significantly limiting their residence time at the site of action. In this study, on the basis of prior research performed in the same laboratory, liposomes were formulated and characterized with the aim of developing an oral spray for the treatment of xerostomia. Although xerostomia by itself is not a dangerous condition, it may lead to complications such as difficulty in swallowing, inflammation, infections, and dental caries. Liposomes were prepared via the thin-film hydration method employing Soy-phosphatidylcholine (Soy-PC) in combination with either dioleoyl-3-trimethylammonium-propane chloride salt (DOTAP) or Egg-Phosphatidylglycerol (Egg-PG) to produce positively and negatively charged liposomes, respectively. The liposomes were then coated with varying concentrations of alginate or chitosan, to identify the most stable formulation with the potential to enhance mucoadhesive properties. Furthermore, the influence of the hydration medium was evaluated by comparing a standard phosphate buffer (PB, pH 6.8) with a phosphate buffer containing 15% glycerol (PBG, pH 6.8) to exploit the intrinsic moisturizing and humectant properties of the latter. Also, the effect of adding a viscosity enhancer, specifically Hydroxyethyl cellulose (HEC) and Hydroxypropyl methylcellulose (HPMC) at different concentrations (0.5% w/v or 1% w/v), was evaluated. The formulations were characterized in terms of hydrodynamic diameter, polydispersity index (PDI), and Zeta potential immediately after preparation and over time to evaluate their stability. Preliminary studies were also conducted to assess the effect of viscosity changes on the sprayability and delivered dose of the final product at different temperature (4°C and 22°C). The results demonstrated that both positively and negatively charged liposomes, when coated with alginate and chitosan respectively, remained stable over a two-month evaluation period in both PB and PBG media. In contrast, uncoated liposomes tended to destabilize after around 4 weeks. Chitosan was selected for further development due to its superior mucoadhesive and antibacterial properties. Regarding the hydration medium, PBG (pH 6.8) was preferred as it leveraged the intrinsic humectant and lubricating properties of glycerol, which are essential for treating xerostomia, without compromising the colloidal integrity of the vesicles. The addition of viscosity enhancers resulted in a concentration-dependent increase in viscosity; however, this addition also induced modifications in the physicochemical liposomal characteristics that compromised their stability. HPMC 1% w/v emerged as the optimal candidate, providing the maximum increase in viscosity while maintaining a stable, homogeneous, and monodisperse population. This significantly higher viscosity was directly reflected in the sprayability tests, where HPMC 1% produced a more concentrated and controlled spray pattern compared to all other formulations. Notably, the delivered dose remained practically unchanged despite temperature variations between 4°C and 22°C, demonstrating the robustness and reliability of the final formulation under different storage conditions.| File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.14239/34323