In vitro evaluation of stearylamine cationic nanoemulsions for improved ocular drug delivery

Oil-in-water nanoemulsions (NEs) represent one of the formulation approaches to improve eye-related bioavailability of lipophilic drugs. The potential of cationic NEs is pronounced due to the electrostatic interaction of positively charged droplets with negatively charged mucins present in the tear film, providing prolonged formulation residence at the ocular surface. The aim of this study was to develop a cationic ophthalmic NE with cationic lipid stearylamine (SA) as a carrier of a positive charge. The addition of a nonionic surfactant provided the dual electro-steric stabilization of NEs and enabled tuning of SA concentration to achieve an optimal balance between its interaction with mucins and biocompatibility. Physicochemical characterization, stability profile, in vitro mucoadhesion study and biocompatibility study employing 3D HCE-T cell-based model of corneal epithelium pointed out the NE with 0.05 % (m/m) SA as the leading formulation. Minimizing SA content while retaining droplet/mucin interactions is of great importance for efficacy and safety of future ophthalmic drug products

Development of functional cationic nanoemulsions for treatment of dry eye disease

Bolest suhog oka je multifaktorijalna bolest koju karakterizira nestabilnost i hiperosmolarnost suznog filma te upala površine oka. Kationske nanoemulzije tipa ulje u vodi (U/V) predstavljaju napredak u liječenju bolesti suhog oka služeći kao tehnološka platforma za uklapanje slabo topljivih djelatnih tvari, omogućujući pritom njihovo produljeno zadržavanje na površini oka te istodobnu nadoknadu i stabilizaciju narušenog suznog filma. Cilj ovog doktorskog rada je razvoj funkcionalne kationske nanoemulzije za liječenje bolesti suhog oka. U tu su svrhu pripravljena dva tipa nanoemulzija korištenjem mikrofluidizatora: primarne kationske nanoemulzije sa stearilaminom i sekundarne kationske nanoemulzije s kitozanom. Primarne kationske nanoemulzije s rastućim udjelima stearilamina male su veličine kapljica (< 100 nm), primjerene disperznosti (PDI ≤ 0,25), pozitivnog zeta-potencijala (3,1-25,5 mV), prikladne pH vrijednosti, male viskoznosti i površinske napetosti 31,3-35 mN m-1. Pripravljene nanoemulzije stabilne su tijekom petomjesečne pohrane. Nakon miješanja s disperzijom mucina zapažene su promjene u veličini kapljica i zeta-potencijalu koje ukazuju na interakciju s mucinom. Takve promjene nisu bile izraženije s povećanjem udjela stearilamina na više od 0,05 % (m/m). Vijabilnost 3D HCE-T modela iznosila je najmanje 90 % nakon izlaganja pripravljenim formulacijama. Sekundarne kationske nanoemulzije pripravljene s manjim (NC1: 0,05 %, m/m) i većim (NC2: 0,3 %, m/m) udjelom kitozana veličine su kapljica približno 180 nm, veće homogenosti (PDI < 0,2) i pozitivnog zeta-potencijala (18,7 i 30 mV). Iako je formulacija NC1 pokazala bolju stabilnost, ibuprofen je uspješno uklopljen u obje formulacije (INC1 i INC2). Fizičkokemijska svojstva (pH, viskoznost, osmolarnost i površinska napetost) pripravljenih formulacija unutar su raspona prikladnog za primjenu na oko. Formulacija INC1 stabilnija je od formulacije INC2 nakon jednomjesečne pohrane. Bakteriološka filtracija prikladna je metoda sterilizacije pripravljenih formulacija. Oslobađanje ibuprofena iz pripravljenih formulacija značajno je brže nego iz uljne otopine i suspenzije ibuprofena. Reološka karakterizacija formulacija pomiješanih s disperzijom mucina pokazala je njihov mukoadhezivni karakter koji, ipak, nije izraženiji kod formulacije INC2 s većim udjelom kitozana. Ispitivanja na 3D HCE-T modelu i ex vivo modelu rožnice svinje pokazala su izuzetnu biokompatibilnost INC1 formulacije. Uzimajući u obzir sve rezultate, nanoemulzija s 0,05 % (m/m) stearilamina i INC1 formulacija ističu se kao vodeće formulacije s velikim potencijalom za liječenje bolesti suhog oka.Dry eye disease is a multifactorial disease characterized by tear film instability and hyperosmolarity and ocular surface inflammation. Oil-in-water (O/W) cationic nanoemulsions represent a progress in dry eye disease treatment serving as a technological platform for incorporation of poorly soluble drugs, enabling prolonged residence time at the ocular surface and, at the same time, replenishment and stabilization of compromised tear film. The aim of this doctoral thesis is development of a functional cationic nanoemulsion for dry eye disease treatment. For this purpose two nanoemulsion types were prepared using microfluidizer: primary cationic nanoemulsions with stearylamine and secondary cationic nanoemulsions with chitosan. Primary cationic nanoemulsions with increasing stearylamine weight fraction are characterized with small droplet size (< 100 nm), low PDI (≤ 0.25), positive zeta-potential (3.1-25.5 mV), appropriate pH, low viscosity and surface tension 31.3-35 mN m-1. The nanoemulsions are stable after 5-month storage. Changes in nanoemulsion droplet size and zeta-potential were observed after mixing with mucin dispersion, which indicates interactions with mucin. However, the changes were not more pronounced with the increase in stearylamine weight fraction above 0.05 % (w/w). Viability of 3D HCE-T model was at least 90 % after exposure to the prepared formulations. Secondary cationic nanoemulsions prepared with low (NC1: 0.05 %, w/w) and high (NC2: 0.3 %, w/w) chitosan weight fraction are characterized with droplet size of around 180 nm, greater homogeneity (PDI < 0.2) and positive zeta-potential (18.7 and 30 mV). Although the formulation NC1 showed greater stability, ibuprofen was successfully incorporated in both formulations (INC1 and INC2). The physico-chemical properties (pH, viscosity, osmolarity and surface tension) of the prepared formulations are within the range acceptable for ophthalmic application. The formulation INC1 is more stable than the formulation INC2 after 1-month storage. Filtration is an appropriate sterilization method for the prepared formulations. Ibuprofen release from the prepared formulations is significantly faster than from ibuprofen oil solution and suspension. Rheological characterization of the formulations mixed with mucin dispersion showed their mucoadhesive character which, however, is not more pronounced with the formulation INC2 with higher chitosan weight fraction. The assays performed using 3D HCE-T model and ex vivo porcine cornea model showed remarkable biocompatibility of the formulation INC1. Taking everything into account, the nanoemulsion with 0.05 % (w/w) stearylamine and INC1 formulation point out as the lead formulations holding great potential for the treatment of dry eye disease

Development of functional cationic nanoemulsions for treatment of dry eye disease

Bolest suhog oka je multifaktorijalna bolest koju karakterizira nestabilnost i hiperosmolarnost suznog filma te upala površine oka. Kationske nanoemulzije tipa ulje u vodi (U/V) predstavljaju napredak u liječenju bolesti suhog oka služeći kao tehnološka platforma za uklapanje slabo topljivih djelatnih tvari, omogućujući pritom njihovo produljeno zadržavanje na površini oka te istodobnu nadoknadu i stabilizaciju narušenog suznog filma. Cilj ovog doktorskog rada je razvoj funkcionalne kationske nanoemulzije za liječenje bolesti suhog oka. U tu su svrhu pripravljena dva tipa nanoemulzija korištenjem mikrofluidizatora: primarne kationske nanoemulzije sa stearilaminom i sekundarne kationske nanoemulzije s kitozanom. Primarne kationske nanoemulzije s rastućim udjelima stearilamina male su veličine kapljica (< 100 nm), primjerene disperznosti (PDI ≤ 0,25), pozitivnog zeta-potencijala (3,1-25,5 mV), prikladne pH vrijednosti, male viskoznosti i površinske napetosti 31,3-35 mN m-1. Pripravljene nanoemulzije stabilne su tijekom petomjesečne pohrane. Nakon miješanja s disperzijom mucina zapažene su promjene u veličini kapljica i zeta-potencijalu koje ukazuju na interakciju s mucinom. Takve promjene nisu bile izraženije s povećanjem udjela stearilamina na više od 0,05 % (m/m). Vijabilnost 3D HCE-T modela iznosila je najmanje 90 % nakon izlaganja pripravljenim formulacijama. Sekundarne kationske nanoemulzije pripravljene s manjim (NC1: 0,05 %, m/m) i većim (NC2: 0,3 %, m/m) udjelom kitozana veličine su kapljica približno 180 nm, veće homogenosti (PDI < 0,2) i pozitivnog zeta-potencijala (18,7 i 30 mV). Iako je formulacija NC1 pokazala bolju stabilnost, ibuprofen je uspješno uklopljen u obje formulacije (INC1 i INC2). Fizičkokemijska svojstva (pH, viskoznost, osmolarnost i površinska napetost) pripravljenih formulacija unutar su raspona prikladnog za primjenu na oko. Formulacija INC1 stabilnija je od formulacije INC2 nakon jednomjesečne pohrane. Bakteriološka filtracija prikladna je metoda sterilizacije pripravljenih formulacija. Oslobađanje ibuprofena iz pripravljenih formulacija značajno je brže nego iz uljne otopine i suspenzije ibuprofena. Reološka karakterizacija formulacija pomiješanih s disperzijom mucina pokazala je njihov mukoadhezivni karakter koji, ipak, nije izraženiji kod formulacije INC2 s većim udjelom kitozana. Ispitivanja na 3D HCE-T modelu i ex vivo modelu rožnice svinje pokazala su izuzetnu biokompatibilnost INC1 formulacije. Uzimajući u obzir sve rezultate, nanoemulzija s 0,05 % (m/m) stearilamina i INC1 formulacija ističu se kao vodeće formulacije s velikim potencijalom za liječenje bolesti suhog oka.Dry eye disease is a multifactorial disease characterized by tear film instability and hyperosmolarity and ocular surface inflammation. Oil-in-water (O/W) cationic nanoemulsions represent a progress in dry eye disease treatment serving as a technological platform for incorporation of poorly soluble drugs, enabling prolonged residence time at the ocular surface and, at the same time, replenishment and stabilization of compromised tear film. The aim of this doctoral thesis is development of a functional cationic nanoemulsion for dry eye disease treatment. For this purpose two nanoemulsion types were prepared using microfluidizer: primary cationic nanoemulsions with stearylamine and secondary cationic nanoemulsions with chitosan. Primary cationic nanoemulsions with increasing stearylamine weight fraction are characterized with small droplet size (< 100 nm), low PDI (≤ 0.25), positive zeta-potential (3.1-25.5 mV), appropriate pH, low viscosity and surface tension 31.3-35 mN m-1. The nanoemulsions are stable after 5-month storage. Changes in nanoemulsion droplet size and zeta-potential were observed after mixing with mucin dispersion, which indicates interactions with mucin. However, the changes were not more pronounced with the increase in stearylamine weight fraction above 0.05 % (w/w). Viability of 3D HCE-T model was at least 90 % after exposure to the prepared formulations. Secondary cationic nanoemulsions prepared with low (NC1: 0.05 %, w/w) and high (NC2: 0.3 %, w/w) chitosan weight fraction are characterized with droplet size of around 180 nm, greater homogeneity (PDI < 0.2) and positive zeta-potential (18.7 and 30 mV). Although the formulation NC1 showed greater stability, ibuprofen was successfully incorporated in both formulations (INC1 and INC2). The physico-chemical properties (pH, viscosity, osmolarity and surface tension) of the prepared formulations are within the range acceptable for ophthalmic application. The formulation INC1 is more stable than the formulation INC2 after 1-month storage. Filtration is an appropriate sterilization method for the prepared formulations. Ibuprofen release from the prepared formulations is significantly faster than from ibuprofen oil solution and suspension. Rheological characterization of the formulations mixed with mucin dispersion showed their mucoadhesive character which, however, is not more pronounced with the formulation INC2 with higher chitosan weight fraction. The assays performed using 3D HCE-T model and ex vivo porcine cornea model showed remarkable biocompatibility of the formulation INC1. Taking everything into account, the nanoemulsion with 0.05 % (w/w) stearylamine and INC1 formulation point out as the lead formulations holding great potential for the treatment of dry eye disease

Development of functional cationic nanoemulsions for treatment of dry eye disease

Bolest suhog oka je multifaktorijalna bolest koju karakterizira nestabilnost i hiperosmolarnost suznog filma te upala površine oka. Kationske nanoemulzije tipa ulje u vodi (U/V) predstavljaju napredak u liječenju bolesti suhog oka služeći kao tehnološka platforma za uklapanje slabo topljivih djelatnih tvari, omogućujući pritom njihovo produljeno zadržavanje na površini oka te istodobnu nadoknadu i stabilizaciju narušenog suznog filma. Cilj ovog doktorskog rada je razvoj funkcionalne kationske nanoemulzije za liječenje bolesti suhog oka. U tu su svrhu pripravljena dva tipa nanoemulzija korištenjem mikrofluidizatora: primarne kationske nanoemulzije sa stearilaminom i sekundarne kationske nanoemulzije s kitozanom. Primarne kationske nanoemulzije s rastućim udjelima stearilamina male su veličine kapljica (< 100 nm), primjerene disperznosti (PDI ≤ 0,25), pozitivnog zeta-potencijala (3,1-25,5 mV), prikladne pH vrijednosti, male viskoznosti i površinske napetosti 31,3-35 mN m-1. Pripravljene nanoemulzije stabilne su tijekom petomjesečne pohrane. Nakon miješanja s disperzijom mucina zapažene su promjene u veličini kapljica i zeta-potencijalu koje ukazuju na interakciju s mucinom. Takve promjene nisu bile izraženije s povećanjem udjela stearilamina na više od 0,05 % (m/m). Vijabilnost 3D HCE-T modela iznosila je najmanje 90 % nakon izlaganja pripravljenim formulacijama. Sekundarne kationske nanoemulzije pripravljene s manjim (NC1: 0,05 %, m/m) i većim (NC2: 0,3 %, m/m) udjelom kitozana veličine su kapljica približno 180 nm, veće homogenosti (PDI < 0,2) i pozitivnog zeta-potencijala (18,7 i 30 mV). Iako je formulacija NC1 pokazala bolju stabilnost, ibuprofen je uspješno uklopljen u obje formulacije (INC1 i INC2). Fizičkokemijska svojstva (pH, viskoznost, osmolarnost i površinska napetost) pripravljenih formulacija unutar su raspona prikladnog za primjenu na oko. Formulacija INC1 stabilnija je od formulacije INC2 nakon jednomjesečne pohrane. Bakteriološka filtracija prikladna je metoda sterilizacije pripravljenih formulacija. Oslobađanje ibuprofena iz pripravljenih formulacija značajno je brže nego iz uljne otopine i suspenzije ibuprofena. Reološka karakterizacija formulacija pomiješanih s disperzijom mucina pokazala je njihov mukoadhezivni karakter koji, ipak, nije izraženiji kod formulacije INC2 s većim udjelom kitozana. Ispitivanja na 3D HCE-T modelu i ex vivo modelu rožnice svinje pokazala su izuzetnu biokompatibilnost INC1 formulacije. Uzimajući u obzir sve rezultate, nanoemulzija s 0,05 % (m/m) stearilamina i INC1 formulacija ističu se kao vodeće formulacije s velikim potencijalom za liječenje bolesti suhog oka.Dry eye disease is a multifactorial disease characterized by tear film instability and hyperosmolarity and ocular surface inflammation. Oil-in-water (O/W) cationic nanoemulsions represent a progress in dry eye disease treatment serving as a technological platform for incorporation of poorly soluble drugs, enabling prolonged residence time at the ocular surface and, at the same time, replenishment and stabilization of compromised tear film. The aim of this doctoral thesis is development of a functional cationic nanoemulsion for dry eye disease treatment. For this purpose two nanoemulsion types were prepared using microfluidizer: primary cationic nanoemulsions with stearylamine and secondary cationic nanoemulsions with chitosan. Primary cationic nanoemulsions with increasing stearylamine weight fraction are characterized with small droplet size (< 100 nm), low PDI (≤ 0.25), positive zeta-potential (3.1-25.5 mV), appropriate pH, low viscosity and surface tension 31.3-35 mN m-1. The nanoemulsions are stable after 5-month storage. Changes in nanoemulsion droplet size and zeta-potential were observed after mixing with mucin dispersion, which indicates interactions with mucin. However, the changes were not more pronounced with the increase in stearylamine weight fraction above 0.05 % (w/w). Viability of 3D HCE-T model was at least 90 % after exposure to the prepared formulations. Secondary cationic nanoemulsions prepared with low (NC1: 0.05 %, w/w) and high (NC2: 0.3 %, w/w) chitosan weight fraction are characterized with droplet size of around 180 nm, greater homogeneity (PDI < 0.2) and positive zeta-potential (18.7 and 30 mV). Although the formulation NC1 showed greater stability, ibuprofen was successfully incorporated in both formulations (INC1 and INC2). The physico-chemical properties (pH, viscosity, osmolarity and surface tension) of the prepared formulations are within the range acceptable for ophthalmic application. The formulation INC1 is more stable than the formulation INC2 after 1-month storage. Filtration is an appropriate sterilization method for the prepared formulations. Ibuprofen release from the prepared formulations is significantly faster than from ibuprofen oil solution and suspension. Rheological characterization of the formulations mixed with mucin dispersion showed their mucoadhesive character which, however, is not more pronounced with the formulation INC2 with higher chitosan weight fraction. The assays performed using 3D HCE-T model and ex vivo porcine cornea model showed remarkable biocompatibility of the formulation INC1. Taking everything into account, the nanoemulsion with 0.05 % (w/w) stearylamine and INC1 formulation point out as the lead formulations holding great potential for the treatment of dry eye disease

Permeability and biocompatibility evaluation of olopatadine hydrochloride viscous ophthalmic solutions using in vitro 3D corneal model

Olopatadine is a selective H1-receptore antagonist that inhibits release of histamine and proinflammatory mediators [1]. The poor ocular bioavailability, as the main limitation of efficient ocular therapy, could be overcome by increasing residence time and enhancing corneal penetration. ...Saopštenje štampano u celin

Towards the development of a biorelevant in vitro method for the prediction of nanoemulsion stability on the ocular surface

Ophthalmic oil-in-water nanoemulsions (NEs) are a complex technological platform, representing an advancement in the treatment of dry eye disease. In addition to enabling the incorporation of poorly soluble active pharmaceutical ingredients (APIs), NEs provide prolonged residence time of APIs and other formulation components and consequent replenishment and stabilization of the compromised tear film. Ophthalmic NEs have been on the market for over 20 years, but considering their complexity, as well as the complex nature of the ocular surface, they are still a poorly understood advanced dosage form. The objective of this study was to develop a biorelevant in vitro method that would be able to predict the behavior of ophthalmic NEs after application. With that goal, NE formulations differing in critical material attributes and critical formulation variables were employed and subjected to simulated tear turnover and blinking. By gradually increasing the complexity of the in vitro method, we were able to detect key parameters influencing NE stability. The undertaken study presents a step forward in the development of in vitro tools that are fundamental to the reliable, cost and time-effective development of innovative and generic topical ophthalmic NEs

Formulation of olopatadine hydrochloride viscous eye drops – physicochemical, biopharmaceutical and efficacy assessment using in vitro and in vivo approaches

The aim of this work was the formulation and the comprehensive evaluation of the viscous eye drops using vehicles containing medium chain chitosan (0.5% w/v), hydroxypropyl guar gum (0.25% w/v) and their com-bination as carriers for olopatadine (0.1% w/v). Physicochemical properties (appearance, clarity, pH, osmolality, viscosity and drug content) of the tested formulations were within acceptable ranges for the ophthalmic prep-arations, while DSC and FT-IR techniques demonstrated the compatibility between olopatadine and polymers. The drug permeability was successfully estimated in vitro using both HCE-T cell-based models (Model I and Model II) and the parallel artificial membrane permeability assay (PAMPA), considering the impact of chitosan as a permeation enhancer. The MTT cytotoxicity assay demonstrates that the tested formulations (diluted 10-fold in HBSS pH 5.5) were non-toxic and well tolerated. An ocular itch test on mice was carried out with the formulation containing the combination of polymers comparable with a commercially available olopatadine eye drops without viscosity enhancers. The tested eye drops produced a slightly higher anti-pruritic/analgesic-like effect than the commercial preparation. It could be assumed that the use of this viscous ophthalmic vehicle due to its advanced mucoadhesive properties and good safety profile is a feasible strategy to improve the efficacy of olopatadine

Functional ibuprofen-loaded cationic nanoemulsion: Development and optimization for dry eye disease treatment

Inflammation plays a key role in dry eye disease (DED) affecting millions of people worldwide. Non-steroidal anti-inflammatory drugs (NSAIDs) can be used topically to act on the inflammatory component of DED, but their limited aqueous solubility raises formulation issues. The aim of this study was development and optimization of functional cationic nanoemulsions (NEs) for DED treatment, as a formulation approach to circumvent solubility problems, prolong drug residence at the ocular surface and stabilize the tear film. Ibuprofen was employed as the model NSAID, chitosan as the cationic agent, and lecithin as the anionic surfactant enabling chitosan incorporation. Moreover, lecithin is a mixture of phospholipids including phosphatidylcholine and phosphatidylethanolamine, two constituents of the natural tear film important for its stability. NEs were characterized in terms of droplet size, polydispersity index, zeta-potential, pH, viscosity, osmolarity, surface tension, entrapment efficiency, stability, sterilizability and in vitro release. NEs mucoadhesive properties were tested rheologically after mixing with mucin dispersion. Biocompatibility was assessed employing 3D HCE-T cell-based model and ex vivo model using porcine corneas. The results of our study pointed out the NE formulation with 0.05% (w/w) chitosan as the lead formulation with physicochemical properties adequate for ophthalmic application, mucoadhesive character and excellent biocompatibility

Synergism of polysaccharide polymers in antihistamine eye drops: Influence on physicochemical properties and in vivo efficacy

The incorporation of polymers into drug delivery vehicles has been shown to be a useful approach to prolong the residence time of drugs in the precorneal tear film and to improve penetration into biological membranes. The main objective of this research was to formulate novel viscous eye drops with ketotifen as the active ingredient, containing the polysaccharides: chitosan (MCH), hydroxypropyl guar gum (HPG) and hyaluronic acid (SH) alone and in combination as functional polymers. DSC and FT-IR techniques showed the compatibility between ketotifen and polymers. Physicochemical and rheological analysis at ambient and simulated physiological conditions, as well as the evaluation of mucoadhesive properties showed that vehicles containing combinations of polymers have suitable physicochemical and functional properties with demonstrated synergism between combined polymers (MCH and HPG i.e. SH and HPG). The drug permeability was successfully estimated in vitro using HCE-T cell-based models. MTT cytotoxicity assay demonstrates that the tested formulations were non-toxic and well tolerated. In vivo preclinical study on mice revealed that both vehicles containing mixed polymers enhanced and prolonged the antipruritic/analgesic-like effect of ophthalmic ketotifen. Based on these results, both combinations of polysaccharide polymers, especially SH-HPG, could be considered as potential new carriers for ketotifen for ophthalmic use