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IntroductionIntroductionIntroductionIntroduction

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1. Introduction

The eye is a unique organ, both anatomically and physiologically, containing several

widely varied structures with independent physiological functions that render the organ

highly impervious to foreign substances. The cornea and crystalline lens are the only

tissues in the body besides cartilage that have no blood supply. The inner and outer

blood-retinal barriers which have no cellular components, separate the retina and the

vitreous body from the systemic circulation therefore reduce the diffusion of molecules.

This complexity of the eye imposes challenges in front of the pharmaceutical scientist to

circumvent the protective barriers of the eye without causing any permanent tissue

damage [1].

1.1 Anatomy of Eye [2]

The human eye is an anatomically and physiology unique and a sensitive organ. It offers

many barriers to the entry of any foreign material. The eyes are protected by the blinking

eyelids at the anterior and at the posterior are housed in a protective bony cavity, the

orbit. The eyeball is ablate spheroid in shape.

Fig. 1.1. Anatomy of eyeball

The eyeball is composed of three layers:

i. Outer fibrous layer (Sclera and Cornea),

ii. Middle vascular layer, the uveal tract (Choroid, Ciliary body & Iris) and

iii. The innermost nervous layer (Retina).

i. Outer fibrous layer

The outer fibrous cornea

anterior and the sclera at the posterior segment. The white and opaque sclera a matrix

composed of proteoglycans and collagen fibres, at the anterior portion is extended to a

clear transparent mucosal surface, the conjunctiva. The most an

part of the eye is the transparent, dome

refractive medium of the eye offering resistance to the passage of most drugs.

The cornea is composed of five layers:

a. Stratified squamous epithelium,

b. Bowman’s membrane,

c. Thick, elastic network of connective tissue, the stroma (Substantia propria),

d. The fibrous collagen rich Descemet’s membrane, and

e. Endothelium (posterior epithelium).

Fig. 1.2. The cornea

hydrophilic barriers

The Bowman’s membrane ends at the cornea sclera junction, the limbus. In the limbus is

present the Canal of Schlemm that drains aqueous humor (secreted in the chamber

The innermost nervous layer (Retina).

Outer fibrous layer

The outer fibrous cornea-sclera layer comprises of the cornea and c

anterior and the sclera at the posterior segment. The white and opaque sclera a matrix

composed of proteoglycans and collagen fibres, at the anterior portion is extended to a

clear transparent mucosal surface, the conjunctiva. The most anterior portion of the outer

part of the eye is the transparent, dome-shaped window, the cornea. It forms the strongest

refractive medium of the eye offering resistance to the passage of most drugs.

The cornea is composed of five layers:

Stratified squamous epithelium,

Bowman’s membrane,

Thick, elastic network of connective tissue, the stroma (Substantia propria),

The fibrous collagen rich Descemet’s membrane, and

Endothelium (posterior epithelium).

The corneal structure showing the sequential hydrophobic and

hydrophilic barriers

The Bowman’s membrane ends at the cornea sclera junction, the limbus. In the limbus is

present the Canal of Schlemm that drains aqueous humor (secreted in the chamber

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sclera layer comprises of the cornea and conjunctiva at the

anterior and the sclera at the posterior segment. The white and opaque sclera a matrix

composed of proteoglycans and collagen fibres, at the anterior portion is extended to a

terior portion of the outer

shaped window, the cornea. It forms the strongest

refractive medium of the eye offering resistance to the passage of most drugs.

Thick, elastic network of connective tissue, the stroma (Substantia propria),

phobic and

The Bowman’s membrane ends at the cornea sclera junction, the limbus. In the limbus is

present the Canal of Schlemm that drains aqueous humor (secreted in the chamber

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between the lens and cornea) from the eye into veins of the sclera therefore, preventing

the rise of intraocular pressure. The hydrophobic epithelium and the hydrophilic stroma

prevent the entry of the hydrophilic and hydrophobic molecules respectively making

cornea the rate limiting membrane for the absorption of drugs.Conjunctiva is the thin,

transparent mucous membrane extending from the lateral margin of the cornea, across the

sclera, covering the internal surface of the eyelids. It is composed of a stratified

squamous columnar epithelium which contains many goblet cells, that rests on a lamina

propria of loose connective tissue.

ii. The uveal tract

The uveal tract also known as the vascular tunic consists of the choroid, ciliary body and

iris. The choroid forms the posterior five-sixths of the vascular tunic. It is the brown

coloured membrane pigmented by the melanocytes which prevents scattering of light rays

within the eye. The ciliary body is composed of ciliary muscles, thickened ring of tissue

which functions for the visual accommodation by controlling the extension of lens. Iris is

the circular, muscular, lightly pigmented diaphragm with a pupil in the center. The

contraction and expansion of the muscles of the iris adjusts the size of the pupil in

response to the light entering the eyeball.

iii. The innermost nervous layer

It is the retina consisting of the photoreceptor cells, bipolar neurons, ganglions and the

pigment cells. The retina is also known as tunica nervosa due to the presence of the

neurons. It is composed of two layers the pigmented layer which is a single layer of

melanocytes and a neural layer which is a sheet of nervous tissue. It contains three main

types of neurons: photoreceptor cells, bipolar cells and the ganglion cells. There are two

types of photoreceptor cells; rod cells responsible for coloured vision and cone cells to

impart vision in bright light. The ganglion cells function to process the electrochemical

information transmitted from the photoreceptors.

The lens is a transparent, avascular, biconvex structure, suspended by the suspensory

ligaments of the lens. It has three components:

i. The lens capsule, produced by anterior lens cells.

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ii. A sub capsular epithelium, a cuboidal layer of cells that is only present on the

anterior surface of the lens.

iii. Lens fibres, derived from the sub capsular epithelial cells, which lose their nuclei

and organelles to become filled with proteins called crystallins.

A semi solid substance called the vitreous humor fills the posterior chamber of the

eyeball behind the lens in direct contact with the retina. It is responsible for the

transmission of light, provides support to the posterior surface of the lens and helps to

maintain the intraocular pressure.

1.2 The Nasolacrimal Apparatus

It is responsible for the production of tears which lubricates the cornea and conjunctiva.

The apparatus consists:

i. Meibomian glands (also known as sebaceous tarsal palpebral glands) secrete

anterior layer composed of lipids, which retard the evaporation of the underlying

water layers, serve to stabilize the film by increasing the surface tension and

lubricate the eyelids.

ii. Lacrimal gland and the accessory lacrimal glands (Krause and Wolfring glands)

secrete the middle aqueous layer which provides oxygen to the corneal

epithelium, and

iii. The goblet cells of the conjunctiva secrete the posterior layer which is a mixture

of mucins (glycoproteins) for hydrating the cornea. The stability of the tear film

depends on the integrity of mucin. In the absence of mucin, the cornea becomes

non wettable and the tear film becomes unstable and is subjected to breaking, with

the formation of dry areas.

The secreted tears flow through the superior and inferior punctum lachrimal followed by

their passage into the superior punctum canaliculus, lachrimal sac. The eyeball can

accommodate approximately 7 µL of fluid. Instillation of more than 10 µL leads to reflex

blinking of the eyelids. The reflex blinking of the eyes in response to the entry of any

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foreign matter like the topically administered formulation leads to tear secretion and

drainage through the nasolacrimal duct to the nasal cavities.

Fig. 1.3. Anatomy of nasolacrimal apparatus

Through the highly vascular mucosa of the nasal epithelium, it is absorbed into the

systemic circulation. Tears dilute the drug remaining in the cul-de-sac, which reduces the

transcorneal flux of the drug.

1.3 Mechanisms and route of transport of drug molecules

The membrane, whether it is plasma membrane or a membrane encompassing cellular

organelles, imposes a barrier to the free movement of molecules. There are two general

processes of drug absorption across barrier membranes. The first type is called "the

transcellular transport process," where drug molecules have to go through the barrier cells

to reach the circulation. Transcellular transport is typically a two step process, starting

with drug uptake into the cells, and ending with drug efflux out of the cells. The second

type is called “the paracellular transport process," where the drug molecules travel

between the cells (or in the gaps) to reach the circulation.

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Fig. 1.4. Transcellular and paracellular pathways for drug transport across

membrane

The transport of most of the molecules is contributed by both pathways. Lipophilic

molecules and molecules with specialized transport processes prefer the transcellular

route; whereas hydrophilic molecules lacking membrane transport processes prefer the

paracellular route. The mechanism of transcellular transport includes simple diffusion,

facilitated diffusion, active transport, endocytosis and exocytosis. Simple or passive

diffusion is a process that does not require cellular energy, but requires a chemical

gradient for the molecule to be transported. Facilitated diffusion is similar to simple

diffusion not requiring energy and follows chemical gradient, but it requires a membrane

transporter or carrier to facilitate the transport; two primary mode of facilitated

transport have been recognized in the biological system [3,4].

The paracellular pathway is an aqueous route involving diffusion of the molecule

between adjacent cells restricted by the presence of a series of junctional strands known

as tight junction, gap junction & desmosomes.

Mechanism of Ocular (Topical) Drug Absorption

Drugs administered by instillation must penetrate the eye and do so primarily through the

cornea and followed by the non-corneal routes. These non-corneal routes involve drug

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diffusion across the conjunctiva and sclera and appear to be particularly important for

drugs that are poorly absorbed across the cornea.

Corneal permeation: The penetration of drugs across the corneal membrane occurs from

the pre-corneal space. Thus, the mixing and kinetic behaviour of drug disposition in tears

has a direct bearing on efficiency of drug absorption into the inner eye. The productive

absorption of most ophthalmic drugs results from diffusional process across the corneal

membrane. The efficiency of absorption process is a function of rate and extent at which

the transport processes occur. The flux of any drug molecule across a biological

membrane depends on the physicochemical properties of the permeating molecule and its

interaction with the membrane. The extent to which the transport or absorption process

occurs is also a function of physiological mechanism of pre-corneal fluid drainage or

turnover.

Factors affecting corneal transport:

Three factors are considered responsible for determining the transport efficiency

of a particular drug species across the corneal membrane.

1) The physiochemical properties of drug substance (e.g., ionization constant,

aqueous solubility, oil/water partition coefficient).

2) The formulation in which drug is prepared (e.g., pH of solution, types and

concentration of buffers, viscosity inducing agents and stabilizers).

3) The corneal structure and integrity.

Non-corneal permeation: Primary mechanism of drug permeation in the sclera is likely to

be diffusion across the intercellular aqueous media as in the case of structurally similar

corneal stroma. Therefore the possibility of partitioning mechanism cannot be eliminated.

Although, like cornea, the conjunctiva is composed of an epithelial layer covering an

underlying stroma, the conjunctival epithelium offers substantially less resistance than

does the corneal epithelium [5, 6].

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Fig. 1.5. Mechanism and routes of drug transportation

1.4 Approaches to ocular drug delivery

Topical administration is more direct, but conventional preparations of ophthalmic drugs,

such as solutions or suspensions are relatively inefficient as therapeutic systems. A large

proportion of the topically applied drug is immediately diluted in the tear film and excess

fluid spills over the lid margin and the remainder is rapidly drained into the nasolacrimal

duct. A portion of the drug is not available for therapeutic action since it binds to the

surrounding extra orbital tissues. In view of these losses frequent topical administration is

necessary to maintain adequate drug levels. This results in transient periods of over and

under-dosing. Three factors have to be considered when drug delivery to the eye is

attempted.

Firstly, how to cross the cornea (external to ocular) to reach the site of action; secondly,

how to localize the pharmacodynamic action at the eye and minimize drug action on

other tissues and finally, how to prolong the duration of drug action such that the

frequency of drug administration can be reduced [7].

Initial attempts to overcome the poor bioavailability of topically instilled drugs (in

solution forms) typically involved the use of ointments based on mixtures of white

petrolatum and mineral oil. Ointment ensures good drug bioavailability by increasing the

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contact time with the eye, minimizing the dilution of tears, and resisting nasolachrymal

drainage. But these vehicles has major disadvantage of providing blurred vision, they are

nowadays mainly used for either night time administration or for treatment on the outside

edges of the eyelids [8].

Now use of suspensions ophthalmic delivery system relies on the assumption that

particles may persist in conjuctival sac. The efficiency of suspension has shown high

variability, which occurred as a result of inadequate dosing, probably mainly due to the

lack of patient’s compliance in adequately shaking the suspension before administration

[9]. These disadvantages led to other formulation approaches being investigated or

simply into the development of dosage formulation that are divided into the ones that can

affect the precorneal parameters, and those that provide controlled and continuous

delivery to the pre and intraocular tissues.

The system discussed involves:

a) The newer system involved include, phase transition hydrogel systems, use of

cyclodextrins to increase solubility of various drug, vesicular systems, particulate

system, and chemical delivery systems such as the prodrugs.

b) The developed and under-developed controlled /continuous drug delivery systems

including ocular inserts, collagen shields, ocular fits, disposable intraocular lenses,

other new ophthalmic drug delivery systems, and

c) The newer trends directed towards the combination of drug delivery technologies

for improving the therapeutic response of a non-efficacious drug.

1.4.1 Conventional ocular delivery system

Eye drops/lotion - Eye drops may be solutions (aqueous and oily) are comparatively

convenient, safe, immediately active and acceptable to patients. Depending on the disease

condition, eye drops may contain steroids, prostaglandins, anesthetics, antihistamines,

sympathomimetics, parasympatholytics, β-receptor blockers, parasympathomimetics, and

non-steroidal anti-inflammatory drugs (NSAID). Eye drops also contain various inactive

ingredients such as viscosity enhancers (hydroxypropyl methylcellulose (HPMC),

carboxy methylcellulose (CMC), polyvinyl alcohol (PVA), and carbopol), preservatives

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(benzalkonium chloride and chlorobutanol), demulcents (polyethylene glycol 400 and

polypropylene glycol), buffers (disodium phosphate, sodium borate, sodium citrate) and

tonicity adjusting agents (sodium chloride).

Fortunately, many therapeutic agents used in eye products are water-soluble compounds

or can be formulated as water-soluble salts, and a sufficiently high solution concentration

can be achieved in the administered dose. In comparison with more sophisticated

multiphase systems, solution products are preferred because they are generally easier to

manufacture and potentially provide better dose uniformity and ocular bioavailability.

The absorption of drugs from the eye is dependent upon the complex interplay of

physiological, physiochemical and formulation factors. Formulation of an optimal

ophthalmic dosage form requires a balancing act between the ocular irritation, corneal

permeation and stability. In this context, effect of formulation factors on corneal

permeation from aqueous drops and oily drops have been investigated earlier for different

NSAIDs like diclofenac [10,11], ketorolac [12,13] and flurbiprofen [14] and

fluoroquinolones like gatifloxacin [15], moxifloxacin [16,17].

Aqueous or oily suspension eye-drop formulations may be considered for drugs that are

poorly water soluble, or because of poor aqueous drug stability. The drug particle size

must be reduced to less than 10 µm levels to avoid irritation of the eye surface, leading to

blinking and excessive lachrymation. Suspensions may also be used to overcome

chemical instability of the drug, but at the same time may pose physical instability

problems. For example, there may be an increase in particle size with time (Ostwald

ripening), or difficulties in resuspension after periods of storage. The latter may result in

problems with homogeneity and dose uniformity. Although eye drops in the solution and

suspension forms have been the most common formulation approach for the treatment of

anterior segment diseases yet they have several drawbacks.

In a study it was reported that a large number of patients found it difficult to instill the

drop [18]. The secretion and drainage of tears at the rate of 1µL/min leads to tear

turnover at the rate of 16% which dilutes the drug solution and eventually increases its

drainage [19, 20]. The rates of drainage increases with increase in volume of the solution

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instilled in the eye resulting in reduced bioavailability of the drug [21]. The amount of

drug absorbed cannot be estimated due to the limited volume holding capacity of the cul-

de-sac. The most widely used preservative benzalkonium chloride causes peeling of the

corneal epithelium cells at their borders, inhibits the growth of the cells and enlarges the

intercellular spaces in the superficial cells of the cornea [22-24]. Corneal calcification can

occur due to the phosphate buffer present in the eye drops [25, 26].

Eye ointments are sterile semi-solid preparations intended for application to the

conjunctiva. They are attractive because of their increased contact time and better

bioavailability compared to solutions but their use is limited as they interfere with vision

and are not suitable for use during the day time [27-29].

1.4.2 Hydrogel

Aqueous gel (hydrogels) consists of high molecular weight, cross linked polymers that

form a three dimensional network in water. Hydrogels are based on the addition of

hydrocolloids to aqueous drug solutions. Currently, two groups of hydrogels are

distinguished, namely preformed and in situ forming gels.

Preformed hydrogels can be defined as simple viscous solutions which do not undergo

any modifications after administration. Preformed hydrogels for topical administration in

the eye can be based on natural, synthetic, or semisynthetic polymers (like cellulose

derivatives, polyvinyl alcohol, carbomer, hyaluronic acid).

The use of preformed hydrogels still has drawbacks that can limit their interest for

ophthalmic drug delivery or as tear substitutes. They do not allow accurate and

reproducible administration of quantities of drugs and, after administration; they often

produce blurred vision, crusting of eyelids, and lachrymation. A new approach is to try to

combine advantages of both solutions and gels, such as accuracy and facility of

administration of the former and prolonged residence time of the latter. Thus, in situ

hydrogels can be instilled as eye drops and undergo an immediate gelation when in

contact with the eye [30].

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In situ gels

In situ gels are the low viscosity solutions that undergo phase transition in the

conjunctival cul-de-sac to form viscoelastic gels due to conformational changes of

polymers in response to the physiological environment change (pH, ion activated or

temperature). The viscous gels reduce the drainage and improve the contact time than the

conventional eye drops. Gellan gum, Gelrite®

, xanthan gum, chitosan, sodium alginate,

carbopol, pluronic, poloxamers and pluronic copolymers are reported to exhibit the sol-

gel transition [31-35].

1.4.3 Ocular inserts

Solid erodible (soluble) or non-erodible (insoluble) inserts have been commercially

available for some time as a means of prolonging the release of drugs in to the eye. The

Ocusert™ (nonerodible system developed by Alza Corporation) and SODI™ (Soluble

Ophthalmic Drug Insert - erodible system) are commercially available formulations.

Ocular inserts are the solid ophthalmic devices usually of 8 mm diameter with drug

dispersed in the polymer reservoir or matrix system intended to increase the precorneal

contact time, increase bioavailability and maintain a therapeutic drug concentration in the

target tissues. The release rate and the mechanism can be controlled by the use of

different natural and synthetic polymers like sodium alginate, collagen, gelatin, Eudragit

RS-100 and RL-100, p-HEMA, polyethylene oxide, cellulose acetate phthalate and

hydroxypropylmethyl cellulose (HPMC) [36-41].

To improve the intraocular retention of inserts, mucoadhesive thiolated ocular inserts

comprising of polyacrylic acid-cysteine conjugate as polymeric matrix, containing either

diclofenac sodium or diclofenac-tris (hydroxymethyl)-amino methane were prepared and

evaluated. In-vitro release studies conducted using simulated lachrymal fluid revealed a

significantly lesser release of drug from diclofenac-tris inserts compared with diclofenac

sodium inserts. The inserts were successful in providing a controlled drug release with

release rate approximating zero order release kinetics [42].

In another study reservoir type ocular inserts of ketorolac were formulated using HPMC

or methylcellulose and povidone as polymeric films, and ethylcellulose film as rate

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controlling membrane [43]. The ocular inserts casted using HPMC (4%) and ethyl

cellulose (3%) was found to sustain ketorolac tromethamine release by zero order kinetics

for 22 h.

Ocular inserts of indomethacin were fabricated using a combination of low and high

molecular weight polyvinyl alcohol. The ex-vivo permeation of drug from ocular inserts

was evaluated through excised goat cornea, and it was observed that physical cross

linking of inserts by freeze-thaw cycling and increase in the proportion of low molecular

weight polyvinyl alcohol decreased the apparent corneal permeability of indomethacin

[44].

In another study, ocular inserts/films of aceclofenac were prepared using glycero-gelatin

polymer and evaluated for physicochemical parameters and drug release profiles. Ocular

irritation of the developed formulation was also checked by HET-CAM test and efficacy

of the developed formulation against prostaglandin-induced ocular inflammation in rabbit

eye was determined. The results of the study showed that the cross-linked ocular insert

proved to be significantly better in various evaluating parameters such as tensile strength,

swelling potential, drug release and pharmacodynamic activity [45].

1.4.4 Contact lenses

Use of soft contact lenses as a carrier for drug delivery is gaining attention. Sedlacek, in

the year 1965 described [46] the use of contact lenses to increase the residence time

(typically days) of the drug in the eye, patient comfort and to limit the drug loss by

drainage, lacrimation or conjunctival absorption [47]. Medicated soft contact lenses can

be used to treat ocular diseases in addition to the correction of refractive deficiencies.

Contact lenses loaded by soaking technique [48] are not therapeutically effective due to a

very low uptake of drug by the lens and a rapid release might lead to low residence time.

The other limitations like the limited drug incorporation in lens matrix, incompetent to

offer slow and extended drug release, wastage of large fraction of drug during loading

procedure have limited its use in ODDS. The uptake and release of the drug depends on

the material of the lens (as a function of ionicity, water and silicone content), loading

solution, time during which is placed in the loading solution and molecular weight of the

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drug incorporated [49]. Novel techniques molecular imprinting and supercritical solvent

impregnations have surmounted all these issues. Molecular imprinting produces synthetic

macromolecular networks template-medicated polymerization mechanisms with specific

affinity, capacity and selectivity for the diffusion of drug molecules. A therapeutically

relevant amount of drug can be loaded and released over an extended period of time and

this technique is applicable to daily-wear and extended-wear contact lenses. In the

supercritical fluid impregnation technique, drug is impregnated and dispersed in a

polymer matrix by dissolving in compressed high volatile fluids (like carbon dioxide) at

temperatures and pressures near or above their critical temperatures and pressures and

bringing the resultant mixture in contact with polymer matrix [50-52].

A method for preparation of soft contact lenses of poly (hydroxyethyl methacrylate)

(PHEMA) using molecular imprinting technique for norfloxacin delivery has been

described [51]. PHEMA hydrogels are used as soft contact lenses. Hydrogel is a

polymeric material which swells in water and retain large quantity of water within its

structure but does not dissolve. Due to high water content hydrogel is soft and rubbery

and offers low frictional irritation to the body tissue. PHEMA hydrogels on immersion in

drug solution takes up the drug which can be applied to eye. PHEMA is synthesized by

polymerization. If HEMA is polymerized with a functional monomer like acrylic acid

which can interact with cationic drug by non-covalent interaction, the affinity of the lens

for drug could be increased. However, if the lens is synthesized by conventional

polymerization method, a random distribution of functional monomer is obtained. In the

molecular imprinting technology, the drug is polymerized with the monomers, so that the

functional monomers arrange themselves around the drug molecule, according to their

interaction capacity. The polymerization and cross-linking fix the spatial sequence and

after removal of the template molecule, recognition cavities complementary in shape and

functionality of the drug are obtained. In the said study, acrylic acid and ethyleneglycol

dimethacrylate (cross-linker) were dissolved in 2- hydroxyethyl methacrylate (HEMA).

Norfloxacin was added to the solution and 2-2'-azo-bis (isobutyronitrle) was added as a

polymerization initiator. The solution was put in a mould and exposed to 50°C for 12h

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and 70°C for 24h. After polymerization, the gel was immersed in boiling water for 15

min to remove unreacted monomers and treated with HCL and water to remove

norfloxacin and subsequently the polymer was dried. Similarly, a non-imprinted polymer

was made without addition of norfloxacin.The imprinted polymer when immersed in

norfloxacin solution loaded greater amounts of norfloxacin than the non-imprinted ones.

Imprinted polymeric hydrogel, synthesized with norfloxacin: acrylic acid in 1:3 and 1:4

molar ratios, showed greatest ability to control the release process sustained for >24h.

1.4.5 Collagen Shield

Collagen is the most abundant primary fibrous protein present in the animal tissues. The

fibrils of the connective tissue in skin, bones, eyes, tendons and ligaments mainly

comprise of collagen. Stroma, which forms 90% of the cornea, and the Bowman’s layer

are lamellas of collagen, which imparts mechanical strength to the eyeball [53, 54].

Collagen derived from porcine sclera and molded into clear thin pliable sheets was

fabricated as a potential vehicle to promote the healing of the de epithelialized corneal

conditions, to eliminate the painful use of contact lenses and to achieve a sustained

delivery of drugs [55, 56]. Bio-Cor® marketed by Bausch and Lomb was the first

commercially available porcine collagen shield. Glucan/collagen therapeutic eye shields

were the patented products of Biosource Genetics Corporation used to deliver glucan

through the collagen shields [57]. A pre-soaked collagen shield of ofloxacin applied

before surgery appeared safe and provided higher concentration of ofloxacin in aqueous

humor (0.96 µg/mL) of the human eye compared to 0.3% w/v ofloxacin aqueous

drop(0.29 µg/mL) [58].

1.4.6 Microspheres

Microspheres are monolithic particles possessing a porous or solid polymer matrix,

whereas microcapsules consist of a polymeric membrane surrounding a solid or a liquid

drug reservoir. The application of such systems in the ophthalmic field has been

extensively reviewed by Kreuter and colleagues [59-60]. Upon topical instillation of a

particulate suspension in the cul-de-sac, the drug is slowly released in the lachrymal pool

by dissolution and mixing, diffusion, or mechanical disintegration or erosion of the

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polymeric matrix (60). The upper size limit for microparticles for ophthalmic

administration is about 5-10 µm. The ophthalmic administration of particles of higher

size can result in an itching sensation and can induce lachrymation, with the possible

consequence of reducing drug bioavailability. Modification with the use of mucoadhesive

and viscous polymers can be used to improve the corneal retention and bioavailability of

the formulation [61].

1.4.7 Nanoparticles

Nanoparticles are polymeric colloidal particles ranging in size from 10 to 1000 nm.

Depending on the method of the preparation, nanospheres or nanocapsules can be

obtained. Nanospheres have a matrix-like structure, wherein active compounds can be

firmly adsorbed onto their surface, entrapped, or dissolved in the matrix. Nanocapsules

have a polymeric shell and an inner core. In this case the active substances are not only

dissolved in the core but may also be adsorbed onto the nanocapsule surface. Colloidal

systems in the sub-micron range can be incorporated in the eye drops as an alternative to

the conventional dosage forms [62, 63].

Pignatello et al formulated nanoparticles with Eudragit RL and RS using a modified

quasi- emulsion solvent diffusion technique and solvent evaporation method [64]. Upon

administration of these nanoparticles in rabbits, sustained release and increased

absorption of the incorporated NSAIDs (ibuprofen and flurbiprofen) were observed.

Furthermore, no signs of inflammation or discomfort were detected in the rabbits' eyes,

suggesting a local bio tolerance of these nanoparticles [65, 66].

To increase the ocular availability, ketorolac was entrapped in N-isopropylacrylamide,

vinyl pyrrolidone and acrylic acid based copolymeric nanoparticles. The particles were

spherical and had a size of 35 nm. In-vitro permeation studies through excised cornea

revealed twofold higher permeation of drug from nanoparticle formulation, compared

with an aqueous suspension of the drug. Pharmacodynamic evaluation of nanoparticle

formulation in PGE2-induced ocular inflammation in rabbits showed a significantly

higher ocular anti-inflammatory activity for 5 h compared with the aqueous suspension,

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which has been attributed to the small size of the particles and mucoadhesiveness. The

nanoparticle formulation did not show any corneal damage during in-vitro study [67].

Adibkia et al also studied a NSAID-loaded Eudragit nanoparticle formulation. The

piroxicam based Eudragit RS100 nanoparticles were prepared by solvent

evaporation/extraction technique. These nanoparticles were used to control inflammatory

symptoms in rabbits with endotoxin-induced uveitis, where a microsuspension of

piroxicam was used as control. The in-vivo examinations revealed that the inflammation

could be inhibited by the nanoparticle formulation more significantly than the pure

piroxicam microsuspension, improving ocular delivery for local inhibition of

inflammation [68].

In another study with PLGA, Vega et al applied an experimental design to optimize a

formulation of flurbiprofen-loaded nanoparticles. The particle surface was coated with

Poloxamer 188 (BASF SE, Ludwigshafen, Germany), resulting in zeta potential values of

approximately -25 mV and a particle size of 230 nm. Drug entrapment efficiencies of

over 90% were obtained, and in-vitro release studies showed a burst release, followed by

a slower release, completed after 90 min. In-vivo studies performed in rabbits

demonstrated that the formulations did not induce toxicity or irritation. Nanoparticle

formulations were compared with the commercial eye drops (Ocoflur Allergan, Belgium)

after induction of inflammation by instillation of sodium arachidonate. The results

indicated a very good anti-inflammatory efficacy for flurbiprofen-loaded PLGA

nanoparticles providing controlled and continuous drug delivery [69].

1.4.8 Solid lipid nanoparticles

SLNs were first patented by Muller and Lucks (1996). Since then they have attracted

scientists as a widely used, stable, non-toxic, and reliable particulate drug delivery

vehicle. Lipid nanoparticles consist of solid lipid spheres, with average diameters

between 50 and 1000 nm, dispersed in aqueous medium [70-72]. These particles are

similar to those described as polymeric nanoparticles, being in this case composed of a

solid lipid core matrix, stabilized by surfactants. As such, these lipid nanoparticles will

have high encapsulation efficiency and loading capacity for lipophilic molecules.

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Surfactants used for their production include biological membrane lipids (lecithin, pure

phospholipids), bile salts (e.g., sodium taurocholate), biocompatible nonionic molecules

(e.g., ethylene oxide/propylene oxide copolymers, sorbitan esters, and fatty acid

ethoxylates). When optimized, lipid nanoparticles show high physical stability, protection

of incorporated labile drugs against degradation and excellent in-vivo tolerability.

However, these systems generally exhibit a low drug payload and have the tendency to

expulsion of drug during storage due to the transition of highly ordered lipid particles

[73, 74].These disadvantages can be alleviated by using structured lipid matrices in SLN

formulations and surface modification of the particles [75]. The sodium diclofenac loaded

lipid nanoparticles were prepared by Attama et al using the combination of homolipid

from goat (goat fat) and a phospholipid, applying the hot high-pressure homogenization

method [76]. Administration of this formulation in bioengineered human cornea showed

a sustained release of the drug. Furthermore, permeation of sodium diclofenac through

the cornea construct was improved by surfacing nanoparticles with phospholipid, which

showed the better performance of this formulation for ocular application.

1.4.9 Liposomes

Liposomes are membrane-like vesicles consisting of one or more concentric bilayers,

produced from natural nontoxic phospholipids and cholesterol, alternating aqueous and

lipophilic compartments. Because of their size, amphiphilic properties, and

biocompatibility, liposomes are promising systems for drug delivery. They can be

classified according to structural features as multilamellar vesicles, small unilamellar

vesicles, and large unilamellar vesicles, depending on their size and the number of lipid

bilayers. With respect to their electrical charge, liposomes can have a positive, negative,

or neutral surface charge, depending on their chemical composition (type of

phospholipids, drug substances, and other lipid molecules). They can accommodate both

hydrophilic and lipophilic drugs, respectively, into the aqueous compartment or the lipid

bilayer [77].

The use of phospholipid vesicles to increase the precorneal residence time and as carriers

for water soluble and lipid soluble drugs has been reported [78, 79]. Positively charged

19

liposomes showed a higher binding affinity to the anionic corneal and conjunctival

mucoglycoproteins and better entrapment efficiency than the neutral and negatively

charged vesicles [80]. Mixed brain gangliosides were incorporated into the membranes of

phosphatidylcholine liposomes to provide receptor sites for wheat germ agglutinin, a

lectin that binds strongly to corneal epithelium, led to enhanced topical drug flux [81].

Collasomes, liposomes coupled to collagen matrices and PAMAM (Polyamido amine)

dendrimers-coated puerarin liposomes were prepared to increase the adhesion capacity

and penetration of liposomes [82, 83]. In another study, pre-corneal retention of

diclofenac sodium has also been reported to be increased by use of liposomes. The

cationic liposomes of diclofenac sodium prepared by reverse phase evaporation were

observed to provide a 211% increase in aqueous humor concentration of drug compared

with conventional aqueous eye drop formulation [84]. Niosomes were found to be better

vesicle system to deliver hydrophilic and lipophilic drugs to eyes due to their better

stability than the liposomes [85, 86]. Large disc shaped vesicles, discomes were derived

from niosomes by the addition of a non-ionic surfactant Solulan C24 [87]. The large size

of discomes fit in cul-de-sac thereby preventing its drainage into the systemic pool as

well as disc shape [88].

1.4.10 Ocular minitablets

These are the bioerodible tablets weighing about 6 mg and 8 mm (4×2mm) in diameter.

Minitablets with matrix system were prepared by direct compression of the powdered

blend of drug, polymer and other excipients. The drug release occurs due to swelling of

the polymer followed by dissolution in the tear fluid. High drug concentration can be

maintained in tear fluid using minitablets for a prolonged time period. On insertion, the

tablet absorbs the tear fluid and sustains the drug release on being hydrated [89, 90].

1.4.12 Ocular iontophoresis

It is an advanced non-invasive technique to avoid the complications associated with

administration of frequent, high dose sub-conjunctival injections or eye drops which

require hospitalization of the patient [91-93]. Direct current (DC) of low intensity 0.2 mA

to 4 mA for 10-20 min is applied that drives charged molecules across the cornea, sclera

20

and the adjacent tissues [94-98]. It is limited to drug molecules which can ionize and

have low molecular weight. Singh et al., proposed a new efficient technique

MacroesisTM (patent filed by Buckeye Pharmaceuticals, Beachwood, OH, U.S.) which

uses alternating current instead of direct current. Therapeutic concentrations were

achieved in the ocular tissues in less time period and it is possible to deliver the drug to

the posterior segment of the eyeball [99].

1.5 Ophthalmic inflammatory disorders and treatment

The most common disease affecting the eye is inflammation. Inflammatory conditions of

the eye are designated according to the tissue affected. Conjunctiva and cornea are

constantly exposed to various types of physical, chemical, microbial (bacteria, fungi,

viruses), and allergic agents and hence prone to develop acute, sub-acute, and chronic

inflammations. Conjunctivitis, keratitis, retinitis, uveitis, endophthalmitis, dacryocystitis,

dacryoadenitis and iritis are the inflammatory disorders of the eye parts. The disease

ranges in severity from mild forms, which can still interfere significantly with quality of-

life, to severe cases characterized by potential impairment of visual function. Ocular

inflammation is also a common result of cataract surgery, producing pain and

photophobia in many patients and potentially leading to serious complications including

increased intraocular pressure, posterior-capsule opacification, cystoid macular edema,

and decreased visual acuity.

Inflammation is manifested as a cellular and vascular response to the injury, infection,

ischemia and excessive or inappropriate operation of immune mechanism. The response

is amplified by activation of inflammatory cells and production of chemical mediators

like acidic lipids e.g. prostaglandins, thromboxanes, leukotrienes, vasoactive amines,

cytokines etc. The acidic lipids are produced through arachidonic acid metabolism.

Arachidonic acid is released from the phospholipids component of cell membrane by the

action of phospholipase A2. The arachidonic acid is fed into the cyclooxygenase and

lipoxygenase pathways, resulting in production of pro-inflammatory prostaglandins and

leukotrienes [100].

21

Ocular actions of PGs are manifested in three ways [101]. Firstly, they act on intraocular

pressure (IOP). PGE1 & E2 increase the IOP by local vasodilation and increased

permeability of blood aqueous barrier. On the other hand PGF2α lowers the IOP which is

attributed to increased uveoscleral out flow. Secondly they act on iris smooth muscle to

cause miosis. Thirdly, PGs cause vasodilation and increase the vascular permeability

resulting in increased aqueous humor protein concentration.

The pharmacological approach of management of inflammation involves administration

of anti-inflammatory agents. Topical administration of drugs is the most preferred route

for management of ocular inflammations as it provides higher ocular drug concentrations,

avoiding the systemic side effects associated with the oral administration. However due

to the physiologic constraints of the eye only few of the anti-inflammatory agents which

possess certain physicochemical properties can be formulated into a suitable dosage form

effective for the management of ocular inflammations. Corticosteroids used to be the

mainstay of topical therapy in the management of ocular inflammations [102], but their

anti-inflammatory effect was outweighed by serious adverse effects like elevation of

intraocular pressure, progression of cataracts, increased risk of infection and worsening

of stromal melting [103]. Corticosteroids, the potent anti-inflammatory agents elicit their

action by blocking the enzyme phospholipase A2 to inhibit arachidonic acid production,

thereby preventing the synthesis of all the PGs, thromboxanes and eicosanoids. On the

other hand non-steroidal anti-inflammatory drugs (NSAIDs) exert their anti-inflammatory

action by inhibiting the enzymes cyclooxygenase (COX 1 & COX 2).

As a class, NSAIDs have proven to be a safe and effective alternative to corticosteroids in

the topical management of ocular inflammations [104]. Currently these drugs are used

topically very widely in inhibition of intra-operative miosis, management of post-

operative inflammation, treatment of seasonal allergic conjunctivitis, prevention and

treatment of cystoid macular edema and in the control of pain after photo refractive

keratectomy [105]. NSAIDs have also been found to be useful in decreasing bacterial

colonization of contact lenses and prevent bacterial adhesion to human corneal epithelial

cells [106].

22

NSAIDs comprise of several chemically heterogeneous class of drugs which possess

potent cyclooxygenase inhibitory activity. However, the topical use of NSAIDs in

ophthalmology is limited to relatively water soluble salicylic acid, indole acetic acid, aryl

acetic acid, aryl propionic acid and enolic acid derivatives. Most of the NSAIDs are

weakly acidic drugs, which ionize at the pH of the lachrymal fluid and therefore have

limited permeability through the anionic cornea which has an isoelectric point (pI) of 3.2

[107]. Reducing the pH of the formulation increases the unionized fraction of the drug

which enhances permeation. Being acidic, NSAIDs are inherently irritant and reducing

the pH of formulation further increases their irritation potential, as well as decreasing

their aqueous solubility. In addition, the anionic nature of NSAIDs lends to the formation

of insoluble complexes with cationic quaternary ammonium preservatives, such as

benzalkonium chloride [10, 14]. Thus; it has proved difficult to formulate topical NSAID

formulations that are comfortable when applied topically to the eye.

There are four NSAIDs currently approved by the US Food and Drug Administration for

the treatment of postoperative inflammation after cataract surgery, namely kerotolac,

bromfenac, diclofenac and nepafenac.

Aceclofenac, 2-[[2-[2- [(2, 6-dichloro phenyl) amino] phenyl] acetyl] oxy] acetic acid, is

a NSAID of the phenyl acetic acid group which is structurally related to diclofenac. It

possesses good anti-inflammatory and analgesic activities, while maintaining better

gastric tolerance in comparison with other NSAIDs such as indomethacin and diclofenac.

Aceclofenac acts as such by inhibiting the secretion of tumor necrosis factor (TNF-α) and

interleukin-1 along with preferential selective cyclooxygenase-2 (COX-2) inhibition after

conversion into active metabolite [108-110]. The present study gives an insight into

various approaches used in topical ocular delivery of aceclofenac.

23

Drug profile [110-112]

Aceclofenac

Chemical Structure

Drug Category : Anti-inflammatory

Chemical Name : 2-[(2, 6-Dichlorophenylamino) phenyl]acetoxy acetic acid

Molecular Formula : C16H13Cl2NO4

Molecular Weight : 354.2

Melting Point : 149° to 150 °C

Description : A white to almost white crystalline powder.

Solubility : It is practically insoluble in water; soluble in alcohol and

methyl alcohol; freely soluble in acetone and

dimethylformamide.

Partition Coefficient : 2.9

Elimination Half Life : 4 h

Dissociation Constant : 4.7 [pKa]

24

Pharmacokinetics

Absorption : Aceclofenac is well absorbed after oral administration.

Metabolites : It is metabolised in hepatocytes and microsomes to 4-

hydroxyaceclofenac, which can undergo further

conjugation. Other metabolites also identified include

diclofenac and 4-hydroxydiclofenac, and their

hydroxylated derivatives.

Distribution : Aceclofenac penetrates into synovial fluid and

concentrations here reach approximately 57% of those

in plasma.

Volume of distribution : Apparent volume of distribution is approx. 25 L

Protein binding : >99% plasma protein bound.

Clearance : 70% of an administered dose is excreted in urine as

glucuronides of aceclofenac and diclofenac

Mechanism of action : Preferential selective cyclooxygenase-2 (COX-2)

inhibition after conversion into active metabolite,

aceclofenac acts as such by inhibiting the secretion of

tissue necrosis factor (TNF-α) and interleukin-1.

Dosage : A usual dose of 100 mg twice daily is administered with

a reduction to 100 mg daily for patients with hepatic

impairment.

Adverse Effects : Dizziness, vertigo, puritus, rash and dermatitis

Drug Interactions : Interaction with anticoalulants, cyclosporins, diuretics,

quinolones,digoxin,methotrexate and lithium

Usage and adminisrations : It is used in the management of osteoarthritis,

rheumatoid arthritis, and ankylosing spondylitis, in

usual oral doses of 100 mg twice daily. Reduced doses

should be used in patients with hepatic impairment.

Proprietary names : Airtal; Barcan; Biofenac; Difucrem; Falcol; Gerbin.

25

polymer profile [113]

Eudragit RL 100 / Eudragit RS 100

Nonproprietary Name

"Ammonio Methacrylate Copolymer Type A“Ph. Eur.

"Ammonio Methacrylate Copolymer Type B“Ph. Eur.

"Ammonio Methacrylate Copolymer, Type A and B" USP/NF

"Aminoalkylmethacrylate Copolymer RS" JPE

Commercial form

Solid substances Eudragit RL 100 (Type A) and Eudragit RS 100 (Type B)

Chemical structure

Eudragit RL 100- Poly (ethyl acrylate, methyl methacrylate,

trimethylammonioethyl methacrylate chloride) 1 : 2 : 0.2.

Eudragit RS 100- Poly(ethyl acrylate, methyl methacrylate,

trimethylammonioethyl methacrylate chloride) 1 : 2 : 0.1.

Eudragit RL 100 and Eudragit RS 100 are copolymers of acrylic and methacrylic acid

esters with a low content in quaternary ammonium groups. The ammonium groups are

present as salts and make the polymers permeable. The average molecular weight is

approx. 150,000.

26

Structural formula of Eudragit RS and Eudragit RL

Description

Eudragit RL 100 and Eudragit RS 100: colourless, clear to cloudy granules with a faint

amine-like odour. Eudragit RS and Eudragit RL are copolymers synthesized from acrylic

acid and methacrylic acid esters, with Eudragit RL (Type A) having 10% of functional

quaternary ammonium groups and Eudragit RS (Type B) having 5% of functional

quaternary ammonium groups. The ammonium groups are present as salts and give rise to

pH independent permeability of the polymers. Both polymers are water-insoluble, and

films prepared from Eudragit RL are freely permeable to water, whereas, films prepared

from Eudragit RS are only slightly permeable to water.

Solubility

1g of the substances dissolves in 7g aqueous methanol, ethanol and isopropyl alcohol

(containing approx. 3 % water), as well as in acetone, ethyl acetate and methylene

chloride to give clear to cloudy solutions. The substances are practically insoluble in

petroleum ether, 1 N sodium hydroxide and water.

Safety

Polymethacrylate copolymers are widely used as film-coating materials in oral

pharmaceutical formulations. They are also used in topical formulations and are generally

regarded as nontoxic and nonirritant materials. A daily intake of 2 mg/kg body-weight of

27

Eudragit (equivalent to approximately 150mg for an average adult) may be regarded as

essentially safe in humans.

Applications in Pharmaceutical Formulation or Technology

Eudragit RL 100 and Eudragit RS 100 are used to form water-insoluble film coats for

sustained-release products. Eudragit RL films are more permeable than those of Eudragit

RS, and films of varying permeability can be obtained by mixing the two types together.

28

References

1. Mitra AK, editor. Ophthalmic drug delivery systems. 2nd

edition. Marcel Dekker

Inc., New York, USA, 2003. p. 1-15.

2. Davson, H. (1969). In "The Eye" (H. Davson, ed.), Vol. 1, 2nd ed. Academic Press,

New York and London

3. Grass GM, Cooper ER, Robinson JR. Mechanisms of corneal drug penetration III:

Modeling of molecular transport. J Pharm Sci 2006; 77(1):24 - 6.

4. Grass GM, Robinson JR. Mechanism of corneal drug penetration I: In vivo and In-

vitro kinetics. J Pharm Sci 1988; 77(1):3-14.

5. Chastain JE, Precorneal, corneal and post corneal factors, in: A.K. Mitra (Ed.),

Ophthalmic Drug Delivery Systems, Marcel Dekker, New York, (2003) pp. 59–82

6. Maurice DM and Mishima S. Ocular pharmacokinetics, in: M.C. Sears (Ed.),

Handbook of Experimental Pharmacology, Pharmacology of the Eye, vol. 69,

Springer-Verlag, Berlin-Heidelberg (1984) pp. 19-116

7. Washington N, Washington C. and wilson,C.G. physiological pharmaceutics,

Barriers to drug absorbtion ,2nd edition, (2001) pp; 254-255

8. Lachman L, Liberman HA, ; The theory and practice of industrial pharmacy;

Verghees publishing house, Bombay , 3rd edition, (1991) pp 548

9. Hecht G, Roehrs RE, Cooper ER, Hiddemen JW, VanDuzee BF (1990) Design and

evaluation of ophthalmic pharmaceutical products. In: Banker GS, Rhodes CT

(ed.), Modern Pharmaceutics, Vol. 40, 2 nd ed., Marcel Dekker, New York, pp.

539-603.

10. Ahuja M, Dhake AS, and and Majumdar DK. Effect of formulation factors on In-

vitro permeation of diclofenac from experimental and marketed aqueous eye drops

through excised goat cornea. Yakugaku Zasshi 2006; 126:1369-1375.

11. Ahuja M, Sharma SK and Majumdar DK. In-vitro corneal permeation of diclofenac

from oil drops. Yakugaku Zasshi. 2007; 127(10):1739-1745.

29

12. Malhotra M, Majumdar DK. In-vitro transcorneal permeation of ketorolac

tromethamine from buffered and unbuffered aqueous ocular drops. Indian J Exp

Biol. 1997, 35:941Y947.

13. Malhotra M and Majumdar DK. In-vitro transcorneal permeation of ketorolac from

oil based ocular drops and ophthalmic ointment. Indian J. Exp. Biol. 1997;

35:1324-1330.

14. Gupta M and Majumdar DK. Effect of concentration, pH and preservatives on In-

vitro transcorneal permeation of ibuprofen and flurbiprofen from non-buffered

aqueous drops. Indian J. Exp. Biol. 1997;35:844-849.

15. Rathore MS, Majumdar DK. Effect of formulation factors on In-vitro transcorneal

permeation of gatifloxacin from aqueous drops. AAPS PharmSciTech 2006; 7:57.

16. Pawar PK, Majumdar DK. Effect of formulation factors on In-vitro permeation of

moxifloxacin from aqueous drops through excised goat, sheep, and buffalo corneas.

AAPS PharmSciTech 2006; 7:E13.

17. Pawar PK, Majumdar DK. In-vitro permeation characteristics of moxifloxacin from

oil drops through excised goat, sheep, buffalo and rabbit corneas. Pharmazie 2007;

62:853-7.

18. Winfield AJ, Jessiman D, Williams A, Esakowitz L. A study of the causes of non-

compliance by patients prescribed eyedrops. Br J Ophthalmol 1990; 74:477-80.

19. Mishima S, Gasset A, Klyce SD Jr, Baum JL. Determination of tear volume and

tear flow. Invest Ophthalmol 1966; 5:264-76.

20. Schoenwald RD, Boltralik JJ. A bioavailability comparison in rabbits of two

steroids formulated as high viscosity gels and reference aqueous preparations.

Invest Ophthalmol Vis Sci 1979; 18:61-6.

21. Benedetto DA, Shah DO, Kaufman HE. The instilled fluid dynamics and surface

chemistry of polymers in the pre ocular tear film. Invest Ophthalmol 1975; 14:887-

902.

30

22. Pfister RR, Burstein N. The effects of ophthalmic drugs, vehicles, and preservatives

on corneal epithelium: a scanning electron microscope study. Invest Ophthalmol

1976; 15:246-59.

23. Samples JR, Binder PS, Nayak S. The effect of epinephrine and benzalkonium

chloride on cultured corneal endothelial and trabecular meshwork cells. Exp Eye

Res 1989; 49:1-12.

24. Lidgate DM, Fu RC, Fleitman JS. Corneal permeability of ketorolac tromethamine

when formulated with tobramycin. Drug Dev Ind Pharm 1989; 15:1779-95.

25. Schrage NF, Schlossmacher B, Aschenbernner W, Langefeld S. Phosphate buffer in

alkali eye burns as an inducer of experimental corneal calcification. Burns 2001;

27:459-64.

26. Schrage NF, Frentz M, Reim M. Changing the composition of buffered eye-drops

prevents undesired side effects. Br J Ophthalmol 2010; 94:1519-22.

27. Hardberger R, Hanna C, Boyd CM. Effects of drug vehicles on ocular contact time.

Arch Ophthalmol 1975; 93:42-5.

28. Shchipanova AI, Maĭchuk IuF, Nikolaeva IS, Pershin GN. Pharmacokinetics and

antiviral activity of riodoksol in an ointment and in ophthalmic drug films in ocular

herpes. Farmakol Toksikol 1981; 44:439-42.

29. Greaves JL, Wilson CG, Birmingham AT. Assessment of the precorneal residence

of an ophthalmic ointment in healthy subjects. Br J Clin Pharmacol 1993; 35:188-

92.

30. Edith Mathiowitz. In: Encyclopedia of controlled drug delivery, A Wiley-

lnterscience Publication.

31. Srividya B, Cardoza RM, Amin PD. Sustained ophthalmic delivery of ofloxacin

from a pH triggered in situ gelling system. J Control Release 2001; 73:205-11.

32. Gupta H, Velpandian T, Jain S. Ion- and pH-activated novel in-situ gel system for

sustained ocular drug delivery. J Drug Target 2010; 18:499-505.

33. Balasubramaniam J, Pandit JK. Ion-activated in situ gelling systems for sustained

ophthalmic delivery of ciprofloxacin hydrochloride. Drug Deliv 2003; 10:185-91.

31

34. Lin HR, Sung KC. Carbopol/pluronic phase change solutions for ophthalmic drug

delivery. J Control Release 2000; 69:379-88.

35. Ma WD, Xu H, Nie SF, Pan WS. Temperature-responsive, Pluronic-g-poly(acrylic

acid) copolymers in situ gels for ophthalmic drug delivery: rheology, In-vitro drug

release, and in vivo resident property. Drug Dev Ind Pharm 2008; 34:258-66.

36. Bensinger R, Shin DH, Kass MA, Podos SM, Becker B.. Pilocarpine Ocular Inserts.

Invest Ophthalmol 1976; 15:1008-10.

37. Sultana Y, Aqil M, Ali A. Ocular inserts for controlled delivery of pefloxacin

mesylate: preparation and evaluation. Acta Pharm 2005; 55:305-14.

38. Hsiue GH, Guu JA, Cheng CC. Poly (2-hydroxyethyl methacrylate) film as a drug

delivery system for pilocarpine. Biomaterials 2001; 22:1763-9.

39. Charoo NA, Kohli K, Ali A, Anwer A. Ophthalmic delivery of ciprofloxacin

hydrochloride from different polymer formulations: In-vitro and in vivo studies.

Drug Dev Ind Pharm 2003; 29:215-21.

40. Di Colo G, Burgalassi S, Chetoni P, Fiaschi MP, Zambito Y, Saettone MF. Gel-

forming erodible inserts for ocular controlled delivery of ofloxacin. Int J Pharm

2001; 215:101-11.

41. Deshpande PB, Dandagi P, Udupa N, Gopal SV, Jain SS, Vasanth SG. Controlled

release polymeric ocular delivery of acyclovir. Pharm Dev Technol 2010; 15:369-

78.

42. Hornof M, Weyenberg W, Ludwig A and Bernkop- Schnurcha A. Mucoadhesive

ocular insert based on thiolated poly acrylic acid): development and in vivo

evaluation in humans. J. Control Rel. 2003;89:419-428.

43. Jayaprakash P S, James CC, Rajan N S M G, Saisivam S and Nagarajan M. Design

and evaluation of ketorolac tromethamine ocuserts. Indian J. Pharm. Sci. 2000;

62:334-338.

44. Pandit JK, Harikumar SL, Mishra D N and Balasubramaniam J. Effect of physical

cross-linking on In-vitro and ex vivo permeation of indomethacin from polyvinyl

alcohol ocular inserts. Indian J. Pharm. Sci. 2003; 65:146-151.

32

45. Manish Mathurm, and Ritu Mehra Gilhotra. Glycerogelatin-based ocular inserts of

aceclofenac: Physicochemical, drug release studies and efficacy against

prostaglandin E2-induced ocular inflammation. 2011; 18(1):54-64.

46. Sedlacek J. Possibility of application of eye drugs with the aid of gel-contact lenses.

Cesk Oftalmol 1965; 21: 509-12.

47. Singh K, Nair AB, Kumar A, Kumria R. Novel approaches in formulation and drug

delivery using contact lenses. J Basic Clin Pharm 2011;2:87-101.

48. Hillman JS. Management of acute glaucoma with Pilocarpine-soaked hydrophilic

lens. Br J Ophthalmol 1974; 58:674-9.

49. Karlgard CC, Wong NS, Jones LW, Moresoli C. In-vitro uptake and release studies

of ocular pharmaceutical agents by silicon-containing and p-HEMA hydrogel

contact lens materials. Int J Pharm 2003; 257:141-51.

50. Costa VP, Mara EM, Braga A, Duarte ARC, Duarte CMM, Leite EOB, Gil MH, de

Sousa HC. Development of therapeutic contact lenses using a supercritical solvent

impregnation method. J Supercrit Fluids 2010; 52:306-16.

51. Alvarez-Lorenzo C, Yañez F, Barreiro-Iglesias R, Concheiro A. Imprinted soft

contact lenses as norfloxacin delivery systems. J Control Release 2006; 113:236-

44.

52. Hiratani H, Fujiwara A, Tamiya Y, Mizutani Y, Alvarez-Lorenzo Cl. Ocular

release of timolol from molecularly imprinted soft contact lenses. Biomaterials

2005; 26:1293-8.

53. Baer E, Cassidy JJ, Hiltner A. Hierarchial structure of collagen composite system:

lessons from biology. Pure & Appl Chem 1991; 63:961-73.

54. Komai Y, Ushiki T. The three-dimensional organization of collagen fibrils in the

human cornea and sclera. Invest Ophthalmol Vis Sci 1991; 32:2244-58.

55. Shaker GJ, Ueda S, LoCascio JA, Aquavella JV. Effect of a Collagen Shield on Cat

Corneal Epithelial Wound Healing. Invest Ophthalmol Vis Sci 1989; 30:1565-8.

56. Palmer RM, McDonald MB. A corneal lens/shield system to promote postoperative

corneal epithelial healing. J Cataract Refract Surg 1995; 21:125-6.

33

57. Erwin LR. Glucan/collagen therapeutic eye shields. US4946450 (1990).

58. Taravella MJ, Balentine J, Young DA, Stepp P.Collagen shield delivery of

ofloxacin to the human eye. J Cataract Refract Surg1999; 25:562-5.

59. Zimmer AK, Zerbe H, Kreuter J. Evaluation of pilocarpine-loaded albumin

particles as drug delivery systems for controlled delivery in the eye I. In-vitro and

in vivo characterization. J Control Release 1994; 32:57-70.

60. Zimmer AK, Chetoni P, Saettone MF, Zerbe H , Kreuter J. Evaluation of

pilocarpine-loaded albumin particles as controlled drug delivery systems for the eye

II. Co-administration with bioadhesive and viscous polymers. J Control Release

1995; 33:31-46

61. Bin Choy Y, Park JH, Prausnitz MR. Mucoadhesive microparticles engineered for

ophthalmic drug delivery. J Phys Chem Solids 2008; 69:1533-36.

62. Calvo P, Vila-Jato JL, Alonso MJ. Comparative In-vitro evaluation of several

colloidal systems, nanoparticles, nanocapsules, and nanoemulsions, as ocular drug

carriers. J Pharm Sci 1996; 85:530-6.

63. Calvo P, Alonso MJ, Vila-Jato JL, Robinson JR. Improved ocular bioavailability of

indomethacin by novel ocular drug carriers. J Pharm Pharmacol 1996;48:1147-52.

64. Pignatello R, Bucolo C, Puglisi G. Ocular tolerability of Eudragit RS100 and

RL100 nanosuspensions as carriers for ophthalmic controlled drug delivery. J

Pharm Sci 2002; 91:2636-41.

65. Pignatello R, Bucolo C, Ferrara P, Maltese A, Puleo A, Puglisi G. Eudragit RS100

nanosuspensions for the ophthalmic controlled delivery of ibuprofen. Eur J Pharm

Sci 2002; 16:53-61.

66. Pignatello R, Bucolo C, Spedalieri G, Maltese A, Puglisi G. Flurbiprofen loaded

acrylate polymer nanosuspensions for ophthalmic application. Biomaterials 2002;

23:3247-55.

67. Gupta AK, Madan S, Majumdar DK, Maitra A. Ketorolac entrapped in polymeric

micelles: preparation, characterization and ocular anti-inflammatory studies. Int. J.

Pharm. 2000; 209:1-14.

34

68. Adibkia K, Siahi MR, Nokhodchi A, Javadzedeh AR, Barzegar-Jalali M, Barar J,

Mohammadi G, Omidi Y. Piroxicam nanoparticles for ocular delivery:

physicochemical characterization and implementation in endotoxin-induced uveitis.

J Drug Target, 2007; 15(6):407-16.

69. Vega E, Egea MA, Valls O, Espina M, Garcia ML. Flurbiprofen loaded

biodegradable nanoparticles for ophthalmic administration. J Pharm Sci 2006;

95:2393-405.

70. Muller RH, Mader K, Gohla S. Solid lipid nanoparticles (SLN) for controlled drug

delivery-a review of the state of the art. Eur J Pharm Biopharm 2000; 50:161-77.

71. Muller RH, Petersen RD, Hommoss A, Pardeike J. Nanostructured lipid carriers

(NLC) in cosmetic dermal products. Adv Drug Deliv Rev 2007; 59:522-30.

72. Muller RH, Radtke M, Wissing SA. Solid lipid nanoparticles (SLN) and

nanostructured lipid carriers (NLC) in cosmetic and dermatological preparations.

Adv Drug Deliv Rev 2002; 54 (Suppl 1):S131-55.

73. Souto EB, Mehnert W, Muller RH. Polymorphic behaviour of Compritol 888 ATO

as bulk lipid and as SLN and NLC. J Microencapsul 2006; 23: 417-33.

74. Souto EB, Muller RH. Investigation of the factors influencing the incorporation of

clotrimazole in SLN and NLC prepared by hot high pressure homogenization. J

Microencapsul 2006; 23:377-88.

75. Attama AA, Schicke BC, Muller-Goymann CC. Further characterization of

theobroma oil-beeswax admixtures as lipid matrices for improved drug delivery

systems. Eur J Pharm Biopharm 2006; 64:294-306.

76. Attama AA, Reichl S, Muller-Goymann CC. Diclofenac sodium delivery to the

eye: In-vitro evaluation of novel solid lipid nanoparticle formulation using human

cornea construct. Int J Pharm 2008; 355:307-13.

77. Niesman MR. The use of liposomes as drug carriers in ophthalmology. Crit Rev

Ther Drug Carrier Syst. 1992;9(1):1-38.

78. Smolin G, Okumoto M, Feiler S, Condon D. Idoxuridine-liposome therapy for

herpes simplex keratitis. Am J Ophthalmol 1981; 91:220-5.

35

79. Schaeffer HE, Krohn DL. Liposomes in topical drug delivery. Invest Ophthalmol

Vis Sci 1982; 22:220-7.

80. Taniguchi K, Itakura K, Yamazawa N, Morisaki K, Hayashi S, Yamada Y Morisaki

K, Hayashi S, Yamada Y Morisaki K, Hayashi S, Yamada Y Morisaki K, Hayashi

S, Yamada Y. Efficacy of a liposome preparation of anti-inflammatory steroid as an

ocular drug-delivery system. J Pharmacobiodyn 1988; 11:39-46.

81. Schaeffer HE, Breitfeller JM, Krohn DL. Lectin-mediated attachment of liposomes

to cornea: influence on transcorneal drug flux. Invest Ophthalmol Vis Sci 1982;

23:530-3.

82. Kaufman HE, Steinemann TL, Lehman E, Thompson HW, Varnell ED, Jacob-

LaBarre JT, Gebhardt BM. Collagen-based drug delivery and artificial tears. J Ocul

Pharmacol 1994; 10:17-27.

83. Liu Y, Sun K, Yao W, Liang N, Mu H, Liang R, Yao C. Corneal penetration of

PAMAM dendrimers-coated puerarin liposomes. Zhongguo Zhong Yao Za Zhi

2010; 35:30-4.

84. Sun KX, Wang AP, Huang LJ, Liang RC and Liu K. Preparation of diclofenac

sodium liposomes and its ocular pharmacokinetics. Yao Xue Xue Bao 2006;

41:1094-1098.

85. Aggarwal D, Garg A, Kaur IP. Development of a topical niosomal preparation of

acetazolamide: preparation and evaluation. J Pharm Pharmacol 2004; 56:1509-17.

86. Abdelbary G, El-Gendy N. Niosome-encapsulated gentamicin for ophthalmic

controlled delivery. AAPS PharmSciTech 2008; 9:740-7.

87. Vyas SP, Mysore N, Jaitely V, Venkatesan N. Discoidal niosome based controlled

ocular delivery of timolol maleate. Pharmazie 1998; 53:466-9.

88. Abdelkader H, Ismail S, Kamal A, Alany RG. Design and evaluation of controlled-

release niosomes and discomes for naltrexone hydrochloride ocular delivery. J

Pharm Sci 2011; 100:1833-46.

36

89. Weyenberg W, Vermeire A, Remon JP, Ludwig A. Characterization and in vivo

evaluation of ocular bioadhesive minitablets compressed at different forces. J

Control Release 2003; 89:329-40.

90. Weyenberg W, Bozdag S, Foreman P, Remon JP, Ludwig A. Characterization and

in vivo evaluation of ocular minitablets prepared with different bioadhesive

Carbopol-starch components. Eur J Pharm Biopharm 2006; 62:202-9.

91. Fishman PH, Jay WM, Rissing JP, Hill JM, Shockley RK. Iontophoresis of

gentamicin into aphakic rabbit eyes. Sustained vitreal levels. Invest Ophthalmol Vis

Sci 1984; 25:343-5.

92. Hobden JA, Rootman DS, O'Callaghan RJ, Hill JM. Iontophoretic application of

tobramycin to uninfected and Pseudomonas aeruginosa-infected rabbit corneas.

Antimicrob Agents Chemother 1988; 32:978-81.

93. Hobden JA, O'Callaghan RJ, Hill JM, Reidy JJ, Rootman DS, Thompson HW.

Tobramycin iontophoresis into corneas infected with drug-resistant Pseudomonas

aeruginosa. Curr Eye Res 1989; 8:1163-9.

94. Grossman RE, Chu DF, Lee DA. Regional ocular gentamicin levels after

transcorneal and transscleral iontophoresis. Invest Ophthalmol Vis Sci 1990;

31:909-16.

95. Hobden JA, Reidy JJ, O'Callaghan RJ, Insler MS, Hill JM. Ciprofloxacin

iontophoresis for aminoglycoside-resistant pseudomonal keratitis. Invest

Ophthalmol Vis Sci 1990; 31:1940-4.

96. Pescina S, Ferrari G, Govoni P, Macaluso C, Padula C, Santi P, Nicoli S. In-vitro

36permeation of bevacizumab through human sclera: effect of iontophoresis

application. J Pharm Pharmacol 2010; 62:1189-94.

97. Molokhia SA, Jeong EK, Higuchi WI, Li SK. Transscleral iontophoretic and

intravitreal delivery of a macromolecule: study of ocular distribution in-vivo and

postmortem with MRI. Exp Eye Res 2009; 88:418-25.

98. Vollmer DL, Szlek MA, Kolb K, Lloyd LB, Parkinson TM. In-vivo transscleral

iontophoresis of amikacin to rabbit eyes. J Ocul Pharmacol Ther 2002; 18:549-58.

37

99. Singh RP, Mathews ME, Kaufman M, Riga A. Transcleral delivery of

triamcinolone acetonide and ranibizumab to retinal tissues using macroesis. Br J

Ophthalmol 2010; 94:170-3.

100. Harry J and Mission G. Clinical Ophthalmic Pathology, Principles of Diseases of

the Eye and Associated Structures, Butterworth Heinemann, Oxford, UK, 2001.

101. Chang J H and Chung H. Non-steroidal anti-inflammatory drug and endotoxin

induced uveitis. Korean. J. Ophthalmol. 1993; 7:35-42.

102. Polansky J and Weinreb R. Steroids as anti-inflammatory agent. In M. Sears (ed.),

Pharmacology of the Eye, Springer, New York, 1984, pp. 460-583.

103. Raizman M. Corticosteroid therapy of eye diseases; fifty years later. Arch.

Ophthalmol. 1996; 114:1000-1001.

104. Waterbury L, Kunysz EA and Bewerman R. Effect of steroidal and non steroidal

anti-inflammatory agents on corneal wound healing. J. Ocul. Pharmacol. 1987;

3:43-54.

105. Schalnus R. Topical nonsteroidal anti-inflammatory therapy in ophthalmology.

Ophthalmologica 2003; 217:89-98.

106. Bandare BM, Sankaridurg PR and Willcox MD. Nonsteroidal anti-inflammatory

agents decrease bacterial colonization of contact lenses and prevent adhesion to

human corneal epithelial cells. Curr. Eye Res. 2004; 29:245-251.

107. Rojanasakul Y and Robinson JR. Transport mechanisms of the cornea:

characterization of barrier permselectivity. Int. J. Pharm. 1989; 55:237-246.

108. Grau M, Montero JL, Guasch J, Felipe A, Carrasco E, Julia S. The pharmacological

profile of aceclofenac, a new non steroidal anti-inflammatory and analgesic drug.

Agents Actions Suppl. 1991; 32: 125-129.

109. Hinz B, Auge D, Rau T, Rietbrock S, Brune K, Werner U. Simultaneous

determination of aceclofenac and three of its metabolites in human plasma by high

performance liquid chromatography. Biomed. Chromatogr. 2003; 17:268-275.

110. Dooley M, Spencer CM, Dunn CJ. Aceclofenac: a reappraisal of its use in the

management of pain and rheumatic disease. Drugs 2001; 61: 1351-78.

38

111. Neil M.J,editor. The Merck index: an Encyclopedia of chemicals, drugs and

biologicals. 14th ed. Whitehouse station, NJ, USA, Merck Research Laboratories

Division of Merk &Co., 2006, 582

112. Martindale. The Complete Drug Reference. 33th

edition, Pharmaceutical Press

2002.

113. Rowe RC, Sheskey PJ, Owen SC. Polymethacrylates In: Handbook of

Pharmaceutical Excipients, 5th

ed.,2009, 553-560