*2.9. Pharmacokinetics*

The amount of clarithromycin diffused into the aqueous humor of the rabbit eyes after ophthalmic administration was determined to compare the ocular bioavailability between CL10 and control (solution). In vivo pharmacokinetic investigations were performed in New Zealand Albino rabbits (2–3 kg) with two groups (*n* = 6). The experiments were carried out by strictly following the guidelines for animal care at Nirma University (IP/PCEU/FAC/21/029; dated 9 June 2017). Single topical instillation (60 µL of 0.25% *w*/*v* drug) of SLN was dropped in one eye of individual rabbit in the first group, while the second eye remained untreated. A similar strength and volume of control solution was instilled into the second group of rabbits. Both eyelids of all rabbits were lightly closed for 2 min to increase the contact of drug with the corneal membrane. Before aqueous humor withdrawal, individual animals were anaesthetized by intramuscular administration of xylazine and ketamine [36]. The aqueous humor was collected using 29-gauge insulin syringe needle [37], and 20 µL of sample was mixed with acetonitrile and centrifuged, and the organic layer was assessed for clarithromycin content by HPLC.
