Razvoj metode za določanje sproščanja zdravilne učinkovine iz očesnega gela in vitro

Lavrič, Suzana (author), Žakelj, Simon (mentor), Krese, Ana (comentor)

Človeško oko je zelo občutljiv organ z zapleteno fiziologijo. Okvara vida je lahko posledica različnih bolezni in močno vpliva na kakovost življenja. Za zdravljenje očesnih okvar je na voljo veliko zdravil, vendar je njihova biološka uporabnost nizka, zaradi številnih absorpcijskih barier v očesu. Za ovrednotenje sproščanja zdravilne učinkovine (ZU) iz farmacevtske oblike (FO) se danes v največji meri uporablja in vitro teste. Osrednji namen magistrske naloge je bil razviti ustrezno metodo sproščanja ZU iz očesnega gela, ki bo imela ustrezno diskriminatornost in robustnost. S tem namenom smo v prvi stopnji izvajali teste raztapljanja učinkovine iz FO na različnih aparaturah. In vitro teste smo najprej izvajali na USP aparaturi 2 – skupaj z majhnimi vesli in Enhancer® celicami, kjer smo ovrednotili vpliv dodatka različnih vrst in koncentracij surfaktantov v osnovno raztopino PBS pH 7,4. Nato smo izvedli test raztapljanja v steklenih čašah – skupaj s steklenimi kroglicami in stresalnikom, kjer smo poskušali čim bolje ponazoriti fiziološke pogoje v očesu. Na koncu pa smo in vitro teste sproščanja izvedli še na USP aparaturi 4 s pretočnimi celicami, kjer smo v nadaljevanju v drugi stopnji z namenom optimizacije metode izvedli še dodatne analize številnih variacij. Po izvedenih testih smo vzorcem določili vsebnost sproščene učinkovine s pomočjo visoko zmogljive tekočinske kromatografije (HPLC). Dobljene profile sproščanja smo med seboj primerjali in vrednotili količino raztopljene učinkovine v določenem času in pa variabilnost rezultatov. S pomočjo uporabe različnih aparatur smo opazili, da izbira le te vpliva na dobljene profile sproščanja učinkovine iz očesnega gela. Ugotovili smo, da dobimo najbolj optimalne profile sproščanja oz. najboljše rezultate z uporabo USP aparature 4. Ta metoda se je namreč izkazala za najbolj diskriminatorno, ponovljivo (tj. nizka variabilnost rezultatov) in robustno. Poleg tega s krajšim časom analize omogoča, da se v čim večji meri približamo fiziološkim pogojem v očesu. Za končno vrednotenje preiskovanih vzorcev smo izbrali sledečo najbolj ustrezno kombinacijo parametrov: medij za raztapljanje: PBS pH 7,4 + 0,5 % Tween 80; volumen medija: 160 mL; temperatura: 35°C, pretok: 8 mL/min; volumen vzorca: 0,2 mL; čas analize: 45 minut; standardna količina steklenih kroglic, Glassfiber PTFE filter.

• Language: Slovenian
• Keywords: oftalmološki gel, očesne pregrade, in vitro testi sproščanja, primerljivost profilov, USP aparatura 4
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2023
• PID: 20.500.12556/RUL-150561
• Publication date in RUL: 20.09.2023

Secondary language

• Language: English
• Title: Development of an in vitro method for drug release evaluation from an ophthalmic gel

Abstract

The human eye is a very sensitive organ with a complex physiology. Visual impairment can be the result of various diseases and greatly affects on the quality of life. Many drugs are available for the treatment of ocular defects, but their bioavailability is low, due to the many ocular absorption barriers. Today, the most common way to evaluate the release of the drug from the pharmaceutical form are in vitro dissolution tests. The main purpose of the Master's thesis was to develop an appropriate in vitro method for evaluating of drug release from an ophthalmic gel, which will have adequate discriminatory capability and robustness. With this aim, in the first stage we performed drug dissolution tests from pharmaceutical forms on various dissolution devices. In vitro tests were first performed on USP Apparatus 2 - together with small paddles and Enhancer® cells, where we evaluated the impact of the addition of different types and concentrations of surfactants in the base solution of PBS pH 7,4. Then we performed a dissolution test in glass beakers - together with glass beads and a shaker, where we tried to illustrate the physiological conditions in the eye as best as possible. In the end, in vitro release tests were performed on USP apparatus 4 with flow cells, where in the second stage, in order to optimize the method, we performed additional analyzes of numerous variations. After preformed tests, we determined the content of the released drug in the samples using high-performance liquid chromatography (HPLC). The obtained release profiles were compared with each other, then the amount of dissolved drug at a certain time points and also the variability of the results were evaluated. With the help of the various devices used, we noticed that the choice of the device affects the obtained drug release profile from the ophthalmic gel. We found that the optimal results were obtained using the USP apparatus 4. Namely, this method proved to be the most discriminatory, reproducible (i.e. low variability of results) and robust. In addition, shorter analysis time allows us to get as close as possible to the physiological conditions in the eye. For the final evaluation of the investigated samples, we chose the following most appropriate combination of parameters dissolving medium: PBS pH 7.4 + 0.5 % Tween 80; medium volume: 160 mL; temperature: 35°C, flow rate: 8 mL/min; volume of sample: 0.2 mL; analysis time: 45 minutes; standard amount of glass beads, Glassfiber PTFE filter.

• Keywords: ophthalmic gel, eye barriers, in vitro release tests, dissolution profile similarity, USP apparatus 4