W/O microemulsions for ocular delivery: Evaluation of ocular irritation and precorneal retention
Introduction
Ocular conditions are usually treated by topical application of drug solutions administered as eye drops. These conventional dosage forms account for around 90% of the available ophthalmic formulations, mainly due to their simplicity and convenience [1]. However, extensive precorneal loss caused by rapid drainage and high tear turnover are amongst the main drawbacks associated with topical ocular drug delivery. Only 1–5% of the topically applied drug penetrates the cornea/sclera and reaches the intraocular tissue with the remainder of the instilled dose undergoing non-productive absorption via the conjunctiva or drainage via the nasolacrimal apparatus. This results in drug loss into the systemic circulation and provides undesirable systemic side effects [2]. Many strategies have been implemented to overcome such delivery challenges. These included the use of thermosetting in situ gelling polymer-based systems [3], nanoparticles, liposomes and niosomes [4], [5], [6], [7]. On the other hand, microemulsions offer a promising alternative. They are thermodynamically stable and optically isotropic colloidal systems with excellent wetting and spreading properties. Moreover, they are comprised of aqueous and oily components and therefore can accommodate both hydrophilic as well as lipophilic drugs.
We have previously reported on the formulation and physico-chemical characterisation of two microemulsion-forming pseudoternary systems [8]. The first is characterised by a phase diagram in which, the single-phase, oil-rich microemulsion (ME) region is followed upon increasing water concentration by a region where liquid crystalline (LC) and coarse dispersed systems form. In the second, the oil-rich and water-rich ME systems merge together forming a continuous one-phase region upon increasing water concentration and the phase diagram is devoid of any LC systems. Systems comprising the two non-ionic surfactants sorbitan mono-laurate (Crill 1) and polyoxyethylene 20 sorbitan mono-oleate (Crillet 4 super) along with oily component ethyl oleate (Crodamol EO) and water as well as systems formulated with 1-hexanol, 1-octanol, 1,2-propandiol, 1,2-pentandiol and 1,2-octandiol as cosurfactants can be classified under the first category whereas systems formulated with 1-propanol, 1-butanol and 1,2-hexandiol as cosurfactants can be classified under the second category [8].
A prototype of the first category is the cosurfactant-free system and its phase behaviour is illustrated in Fig. 1. In this system, the LC region lies next to the ME region and towards the water apex. Thus, fluid ME systems that can undergo a phase-transition to viscous systems containing lamellar liquid crystals upon dilution with water can be formulated using these excipients (arrow, Fig. 1). It is postulated that the phase transition of a cosurfactant-free droplet w/o ME to a more viscous LC system could be induced in situ once such systems are instilled into the eye. Such transition could serve to prolong precorneal retention and hence have application in ophthalmic drug delivery. In contrast to the conventional in situ gel forming systems, the phase transition ME reported herein are oil-based, hence particularly suited for the solubilisation and delivery of lipophilic actives. Further, since these ME systems also contain aqueous domains, they may also have an application in the formulation of hydrophilic or amphiphilic bioactives for ophthalmic drug delivery.
Before testing this hypothesis it was necessary to establish the ocular irritation potential of the developed systems as well as the components used in their formulation. A lack of any ocular adverse effects such as irritation, burning, stinging and tearing is an essential requirement of any system intended for ocular use. This was also considered an essential prerequisite to any in vivo testing as the colloidal systems under investigation contained potential irritant chemicals such as non-ionic surfactants, fatty acid esters, n-alcohols, and 1,2-alkanediols.
Irritation testing of the formulations and the ingredients used to formulate these systems was carried out using a modified HET-CAM test (hen's egg chorioallantoic membrane test) [9]. The chorioallantoic membrane (CAM) is a highly vascular embryonic membrane formed by the fusion of the allantoic wall with the chorion. As a highly vascular and stratified tissue, it responds to injury with an inflammatory reaction similar to that produced by the conjunctiva. Thus the CAM has been suggested as a model for the conjunctiva of the eye [10]. The usefulness of the HET-CAM test to assess the irritation potential of surfactant based and hydro-alcoholic formulations is well established and it is reported that protocols with short exposure times seem to provide more accurate results compared to others with longer exposure time [11]. In addition, the HET-CAM test has been reported as a useful in vitro assay for assessing the eye irritation properties of cosmetic formulation and their ingredients with high agreement of the results with in vivo data [12].
Following irritation testing, a precorneal clearance study using gamma scintigraphy was conducted to evaluate the precorneal retention of the non-irritant formulations and test the hypothesis that dilution of the ME system and subsequent phase transition upon ocular administration by resident tear fluid would enhance retention. The investigated ME systems have a fluid consistency, thus enable ease of administration and the resulting phase-transition may facilitate prolonged retention by virtue of the high viscosity of the formed LC systems. It is well established that nasolacrimal drainage is the major factor behind precorneal drug loss that leads to poor ocular bioavailability and systemic exposure with possible side effects and potential toxicity [13]. Thus it was anticipated that the in situ phase transition would serve to increase ocular retention and subsequently, improve ocular bioavailability. The aims of this work were therefore to:
- •
establish the physiological compatibility of the components used to formulate the previously described pseudoternary systems so as to select formulations that could be evaluated in vivo and;
- •
investigate whether a phase-transition w/o ME based formulation indeed demonstrates prolonged preocular retention.
Section snippets
Materials
Ethyl oleate was used as the oil component (Crodamol EO). Sorbitan mono-laurate was used as surfactant component 1 (Crill 1) and polyoxyethylene 20 sorbitan mono-oleate as surfactant component 2 (Crillet 4 super). Crodamol EO, Crill 1 and Crillet 4 super were obtained form BTB Oleochemicals (Auckland, New Zealand). 1-Propanol, 1-butanol, 1-hexanol and 1-octanol were purchased from BDH (Poole, Dorset, UK) and were used as alcohol cosurfactants. 1,2-Propandiol (May and Baker, Dagenham, England)
Preparation of pseudoternary systems
Systems lying on the sampling path marked by the arrow in Fig. 1, Fig. 2 were prepared by adding water to the oil : surfactant blend/cosurfactant, at increments of 1–5% w / w. The resultant systems were then vortex mixed for 5 min and left to equilibrate overnight at ambient temperature. They were then characterised and classified based on their visual appearance and behaviour using phase-contrast and polarized light microscopy [16].
Irritation testing using a modified HET-CAM test
Freshly collected fertile hen's eggs (White Leghorn) were
Results
Crill 1, Crillet 4 super and Crodamol EO were found to be practically non-irritant when applied to the surface of the CAM. All three components gave a score of 0.0 (n = 4). On the other hand, aliphatic n-alcohols with carbon chain length of 3–8 were ranked as strong irritants while ethanol was ranked as a moderate irritant (Fig. 4a). 1,2-alkanediols with carbon chain length 5–8, previously reported as non-toxic substitutes to n-alcohols [14], [15] were found to be strong irritants while those
Discussion
Reports on the non-ionic surfactant Crill 1 (sorbitan mono-laurate) have shown it to be non-irritant [17], on the other hand reports on the irritation of Crillet 4 super (polyoxyethylene 20 sorbitan mono-oleate also known commercially as Tween 80) have been contradictory with rankings varying from minimally irritant to non-irritant [18], [19], [20]. Moreover, Sterzel et al. showed oleic acid as well as fatty acid esters with carbon chain length of 18 atoms to be non-irritant, based on four
Conclusion
Water-in-oil ME and LC containing systems may be formulated using a blend of the two non-ionic surfactants, Crill 1 and Crillet 4 super, the oily component Crodamol EO and water. The w/o ME systems containing 5% and 10% water were retained in the preocular area significantly longer than a solution whereas a lamellar LC system was retained significantly longer than any other evaluated system. The rapid clearance of the w/o ME formulated with 10% water compared to the LC system indicates that
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