Development of stability-indicating capillary electrophoresis method fo the analysis of febuxostat

Febuksostat je selektivni inhibitor ksantin oksidaze, snažan lijek koji se koristi u terapiji gihta, a u Republici Hrvatskoj registriran je 2012. godine. Još uvijek nema službenu monografiju niti u jednoj Farmakopeji. U ovom diplomskom radu započet je razvoj kapilarnoelektroforetske metode za analizu utjecaja kiseline na febuksostat ekstrahiran iz ljekovitog oblika, tableta Adenuric®, u sklopu stres studija nužnih za razvoj stabilitetno indikativne metode. Kao najprikladnija metoda za analizu odabrana je micelarna elektrokinetička kromatografija zbog mogućnosti da se uz dodatak surfaktanta natrijevog dodecil sulfata (SDS-a) razdvoje i nabijeni i neutralni spojevi. Ispitani su različiti radni puferi te se kao najprikladniji pokazao radni pufer sastava 20 mM boratnog pufera pH 9,3 i 50 mM SDS-a, bez dodatka organskog otapala. Kao prikladan unutarnji standard odabran je ketoprofen, budući da mu dodatak kiseline nije uzrokovao promjene u visini i površini pika, za razliku od pravastatina koji je uz dodatak kiseline laktonizirao i epimerizirao. Koncentracija klorovodične kiseline u iznosu od 0,1 M pokazala se prikladnom za analizu, dok je koncentracija u iznosu od 1 M uzrokovala pretjeranu razgradnju febuksostata. Vrijeme izloženosti febuksostata klorovodičnoj kiselini koncentracije 0,1 M u trajanju od 2 sata i 50 minuta dovelo je do prevelike razgradnje febuksostata, stoga je potrebno odrediti prikladno vrijeme izloženosti kiselini, kao i volumen te koncentraciju natrijevog hidroksida potrebnog za zaustavljanje kisele hidrolize. To prelazi okvire ovog diplomskog rada te ostaje kao izazov u budućim istraživanjima.Febuxostat is a selective inhibitor of xanthin oxidase, a potent drug used in gout therapy, and is registered in the Republic of Croatia in 2012. Febuxostat still does not have an official monograph in any Pharmacopoeia. In this graduate thesis the development of a capillary electrophoretic method for the analysis of the effect of acid on febuxostat extracted from the Adenuric® tablet was started, within stress studies necessary for the development of the stability-indicating method. Micellar electrokinetic chromatography was chosen as the most suitable method, due to the possibility that with the addition of a surfactant (sodium dodecile sulphate, SDS) both neutral and charged compounds can be differentiated. Several working buffers were tested and the one composed of 20 mM borate buffer pH 9.3 and 50 mM SDS without the addition of organic solvent was shown as the most appropriate one. Ketoprofen was chosen as a suitable internal standard, since the addition of acid did not cause changes in height and area of its peak, unlike pravastatin which, by addition of acid, lactonized and epimerized. The concentration of hydrochloric acid in the amount of 0.1 M was found to be suitable for analysis, while the concentration in the amount of 1 M caused excessive degradation of febuxostat. The exposure time of hydrochloric acid to 0.1 M for 2 hours and 50 minutes resulted in excessive degradation of febuxostat, so it is necessary to determine the appropriate exposure time to the acid as well as the volume and concentration of sodium hydroxide necessary to stop acid hydrolysis. It goes beyond the frameworks of this graduate work and remains a challenge in future research