Browsing by Subject "ocular pigment"

Certain drugs accumulate into pigmented tissues due to their binding to melanin, a macromolecule inside pigmented cells. Melanin can affect the drug's pharmacokinetics by acting as a drug reservoir. Binding can also cause toxic effects by accumulating compounds to pigmented cells. This thesis focuses on ocular melanin. The literature review covers the most common methods used in the study of ocular melanin binding and concentrates on in vitro methods and the analysis and usability of the results in pharmacokinetic modeling. The aim of the experimental part was to study melanin binding of a set of compounds in vitro with melanin isolated from the retinal pigment epithelium (RPE) and choroid of porcine eyes and with primary porcine RPE cells and then construct a pharmacokinetic model of melanin binding with STELLA® software and simulate it with the in vitro results. The compounds chosen for the study; nadolol, timolol, chloroquine, methotrexate, carboxydichlorofluorescein (CDCF) and dexamethasone, are small molecules with diverse physicochemical properties (octanol/water partitioning coefficient (logP), pKa, acid/base status). Some are also efflux substrates. The in vitro binding with melanin was studied at pH 7.4 and in addition at pH 5 for the acidic compounds, since the pH inside melanosomes where melanin is located is acidic. Porcine RPE cells were used to study the amount of uptake and rate of elimination of the set of compounds. The effect of efflux was also evaluated with a general efflux inhibitor probenecid. All the basic compounds bound to melanin in vitro. The acidic compounds did not seem to bind at pH 7.4 but bound at pH 5. Chloroquine, as expected, had the highest binding. In the cell studies, the uptake of chloroquine was significant, at least partly due to melanin binding. The other compounds were taken into the cells to a much smaller extent. The efflux inhibitor did not seem to affect the results. The results of the binding study were used in the models constructed of melanin binding and cellular pharmacokinetics. The constructed model was a very simple one not taking into account many factors affecting cellular pharmacokinetics. The results of both the in vitro studies and the model give a good idea of the importance of melanin binding in ocular drug delivery. The model can be used in the future as a base for more comprehensive models of the effect of melanin binding on ocular pharmacokinetics.