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Ophthalmic Drug Dosage Forms: Characterisation and Research Methods

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Hindawi Publishing Corporation

e Scientific World Journal


Volume 2014, Article ID 861904, 14 pages
http://dx.doi.org/10.1155/2014/861904

Ophthalmic Drug Dosage Forms: Characterisation


and Research Methods

Przemys Baw Baranowski, Bohena Karolewicz, Maciej Gajda, and Janusz Pluta
Department of Drug Form Technology, Wroclaw Medical University, Borowska 211A, 50-556 Wroclaw, Poland

Correspondence should be addressed to Przemysław Baranowski; przemyslaw.baranowski@umed.wroc.pl

Received 20 December 2013; Accepted 4 March 2014; Published 18 March 2014

Academic Editors: A. Concheiro and M. Ozyazici

Copyright © 2014 Przemysław Baranowski et al. This is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

This paper describes hitherto developed drug forms for topical ocular administration, that is, eye drops, ointments, in situ gels,
inserts, multicompartment drug delivery systems, and ophthalmic drug forms with bioadhesive properties. Heretofore, many
studies have demonstrated that new and more complex ophthalmic drug forms exhibit advantage over traditional ones and are
able to increase the bioavailability of the active substance by, among others, reducing the susceptibility of drug forms to
defense mechanisms of the human eye, extending contact time of drug with the cornea, increasing the penetration through the
complex anatomical structure of the eye, and providing controlled release of drugs into the eye tissues, which allows reducing
the drug application frequency. The rest of the paper describes recommended in vitro and in vivo studies to be performed for
various ophthalmic drugs forms in order to assess whether the form is acceptable from the perspective of desired properties and
patient’s compliance.

1. Introduction time, which in turn contributes to smaller application fre-


quency [1–5].
Ophthalmic drug forms have been one of the most important One of the first modifications to conventional forms of
and widely developed areas of pharmaceutical technology for ophthalmic drugs was introducing polymers to formulation,
dozens of years. T he main reason of continuingly strong which enabled longer contact time of active ingredient and the
interest of scientists in these drug forms is the problem of a low corneal surface, thus increasing its bioavailability. Next
bioavailability of medicinal substance after the application to possibility to modify the ophthalmic forms active ingredients’
the eyeball. It is caused by, amongst other reasons, the bioavailability involved introducing excipients to formula-
complicated anatomical structure of the eye, small absorptive tion, which enhanced drugs’ penetration into the eyeball.
surface and low transparency of the cornea, lipophilicity of These excipients included chelating agents, surfactants, and
corneal epithelium, metabolism, enzymolysis, bonding of the cyclodextrins, which, along with active ingredients, form
drug with proteins contained in tear fluid, and defence inclusion complexes. This increases solubility, permeability,
mechanisms, that is, tear formation, blinking, and flow of the and bioavailability of poorly soluble drugs [1–4].
substance through nasolacrimal duct [1– 3]. Low capacity of The newer drug forms, on which in recent years research
conjunctival sac, that is, approximately 30 L without blinking has been conducted in order to achieve a controlled release of
[4], and the aforementioned defence mechanisms cause drug to eyeball tissues, include multicompartment carrier
decrease in drug concentration in the place of application and systems, inserts, collagen shields, contact lenses, and the so-
shorten the time during which the active ingredient stays in the called in situ gels [1–3, 5]. The advantages of using these new
place of absorption. T he pri-mary purpose for the development drug forms of controlled release are, among others, increasing
of ophthalmic drug forms is to achieve the required drug bioavailability of substance through extending the time of its
concentration in the place of absorption and sustaining it for contact with cornea—which can be achieved by effective
appropriately long adhesion to the corneal surface, the possibility of targeted
2 The Scientific World Journal

therapy preventing the loss of drug to other tissues, increasing formulation’s viscosity. Substances which have
ensuring patient’s comfort when applying the drug form such effect include hydrophilic polymers of high molecular
and during the whole therapy, and increasing resistance to weight which do not diffuse through biological membranes
eye defence mechanisms, like tearing [4]. and which form three-dimensional networks in the water.
This paper constitutes an overview and characterization Examples of such polymers include polyvinyl alcohol, polox-
of the hitherto developed ophthalmic drug forms. amers, hyaluronic acid, carbomers, and polysaccharides, that
is, cellulose derivatives, gellan gum, and xanthan gum. The
2. Topical Ophthalmic Drug Forms aforementioned carbomer is used in liquid and semisolid
formulations as a suspending substance or a substance which
2.1. Liquid Ophthalmic Drug Forms increases viscosity, whereas hyaluronic acid is used as a
polymer, forming biodegradable and biocompatible matrix,
2.1.1. Eye Drops. Eye drops are accessible in the forms of
which enables extending time periods of drug release [4, 8].
waterand oil solutions, emulsions, or suspensions of one or
The research has proved that maximum increase of
more active ingredients, which may contain preservatives if
stored in multiuse packaging. These forms are sterile and penetration through the cornea by a solution in the form of eye
isotonic. The optimum pH for eye drops equals that of tear drops takes place when the viscosity falls into the range of 15
fluid and is about 7.4. In deciding whether to buffer the drug to 150 mPas. An example of “extreme” use of substances
in this form, one should take into account the stability of increasing viscosity is forming gels, which would enable
active ingredient and the tissue tolerance to the preparation [7– reducing the frequency of drug application to once daily. It has
9]. If the pH value gets outside the range of 4–8 which is been proved that synthetic polyoxyethylene-polyoxypropylene
tolerated by eye, the patient may feel discomfort, there may be block copolymer (poloxamer 407) is suit-able for use as a
irritation, and the drug bioavailability can decrease because of carrier in ophthalmic formulation with pilocarpine, which
increased tearing [10]. stimulates the active ingredient. The main disadvantage of this
formulation is blurring of vision, which negatively affects its
2.1.2. Ophthalmic Solutions. Ophthalmic solutions are acceptability among patients [4, 8].
sterile,aqueous solutions used for, among other things, Presently, hydrophilic polymers are employed in many
cleansing and rinsing eyeballs. They may contain ophthalmic products, though rather as compounds that exhibit
excipients, which, for example, regulate osmotic pressure, mucoadhesive properties than for increasing viscosity
the pH, and viscosity of the preparation. They may also [4]. These forms contain polymers which connect through
contain preservatives if stored in multiuse packaging [7]. noncovalent bonds with conjunctival mucin and usually are
macromolecular hydrocolloids with many hydrophilic groups
2.1.3. Microemulsions. Microemulsions are promising (carboxyl, hydroxyl, amide, and sulfate) able to form
drugforms, inexpensive to produce, and easy to sterilize electrostatic connections, which enables longer contact with
and stable, providing the possibility to introduce larger eye surface. Mucoadhesive dosage form is characterized by
amounts of active ingredient. In vivo research and clinical higher bioavailability in comparison to conventional forms
examination of healthy volunteers proved extended time [5]. Examples of polymers which were examined in the
periods of effective-ness and increased bioavailability of direc-tion of mucoadhesion and increasing substance
drugs applied in these forms. The mechanism of action bioavail-ability in ophthalmic preparations include
involves the adsorption of nanodrops constituting a polyacrylic acid, hyaluronic acid, sodium carboxymethyl
reservoir of the drug and the inner phase of microemulsion cellulose, and chi-tosan. Other compounds which extend
on the corneal surface, which limits the overflow [5]. the time period of contact with eye surface are lectins,
Active ingredients for which microemulsions have been which were also examined in the direction of selective drug
developed include difluprednate [11], cyclosporine A [12], binding to a specified corneal area [4, 5, 8].
flurbiprofen axetil, and the prodrug of flurbiprofen [13]. Two preparations, NyoGel (Novartis) with timolol
maleate and Pilogel (Alcon Laboratories) with pilocarpine
2.1.4. Modifications of Liquid Ophthalmic Dosage Forms. hydrochloride, contain cross-linked polyacrylic acids which
Inthe course of technological research on dosage forms, exhibit mucoadhesive properties, Carbomer and Carbopol,
many ways have been proposed as to how to extend the respectively [14].
time period of contact of liquid dosage forms with eye
tissues, as well as to increase the active ingredient 2.1.6. Addition of Penetration Increasing Substances. The pur-
absorption to these tissues. These modifications include the pose of using penetration increasing substances in oph-thalmic
addition of substances which increase viscosity, drugs is to enhance their corneal absorption by modifying the
introducing the drug penetration enhancing substances to continuity of corneal epithelium structure. Research has shown
formulation, using pro-drugs or cyclodextrins [4, 5, 7–10]. that such properties are displayed by chelating agents,
preservatives (like benzalkonium chloride), surfactants, and
2.1.5. Addition of Substances Increasing Viscosity/Adhesion. bile acid salts. However, these substances displayed local
Extending the time period of contact with cornea and toxicity, which caused restrictions in their use in ophthalmic
improving bioavailability of substances may be obtained by drug forms technology [3, 4].
The Scientific World Journal 3

2.1.7. Prodrugs. Modifying drug properties by effects, because of which they are mainly applied night-
developingprodrugs also enables increasing drug permeability time [8].
through the cornea. This method involves modification of
chemical structure, which gives the active ingredient new
2.3. Solid Ophthalmic Drug Forms
properties, that is, selectivity and site specificity [4]. Examples
of medic-inal substances for which prodrugs were developed 2.3.1. Contact Lenses Coated with Drugs. This drug form
include epinephrine, phenylephrine, timolol, and pilocarpine canabsorb on its surface water-soluble substances, released
[4, 15]. Dipivefrine, a diester of pivalic acid and epinephrine, after applying the drug over the eyeball for a longer period of
displays seventeenfold higher permeability through the cornea time. The first and most widely used polymer in the
than epinephrine, which is caused by its six hundredfold production of lenses was the cross-linked poly(2-hydroxyethyl
higher lipophilicity at pH 7.2. Therefore, a smaller dose of methacry-late) with small amount of ethylene glycol
dipive-frine applied over the eyeball has similar therapeutic dimethylacrylate [4, 5]. In recent years, research has been
effect to epinephrine. In comparison to conventional eye drops conducted on employing silicon-based lenses [17–20]. Interest
containing 2% epinephrine, eye drops with dipivefrine 0.1% in contact lenses still grows, which is confirmed by increase in
display only slightly smaller activity lowering the intraocular the number of articles on its use published in recent years.
pressure with significant reduction of side effects [15]. Examples of drugs whose pharmaceutical availability from
lenses was researched include timolol [17], ciprofloxacin [18],
2.1.8. Cyclodextrins. Cyclodextrins are cyclic oligosaccha- dexamethasone [19], and cyclosporine [20].
rides able to form inclusion complexes with active ingredi-
ents, thus increasing the solubility in water of hydrophobic 2.3.2. Ocular Inserts. Inserts are solid or semisolid dosageforms
compounds without changing their molecular structure [3, 16]. without disadvantages of traditional ophthalmic drug forms [5,
As carriers, they enable keeping hydrophobic drugs in solution 21]. They are less susceptible to defence mechanisms like outflow
and transport them to biomembranes surface. In the case of through nasolacrimal duct, show the ability to stay in conjunctival
ophthalmic drugs, optimal bioavailability of the active sac for a longer period, and are more stable than conventional
ingredient is obtained at the appropriate concentration of dosage forms. Their undoubtable advantages over conventional
cyclodextrins (<15%) in aqueous eye drops solution [4]. The forms are also accurate dosing, the possibility of slow substance
most often used cyclodextrin in developing forms applied over release with constant speed, and limiting its systemic absorption.
the eyeball is 2-hydroxypropyl- -cyclodextrin, which does not Moreover, using them enables reduction of the drug application
show irritating effects. Eye drops containing drug inclusion frequency, as well as adverse effects and blurring of vision
complexes, namely, dexamethasone or pilocarpine with 2- occurrence [8, 21]. Polymeric materials most often employed in
hydroxypropyl- -cyclodextrin, are well tolerated and ensure developing inserts include, for example, methylcellulose [22] and
increased bioavailability in comparison to conven-tional ones its derivatives, that is, hydroxypropyl methylcellulose (HPMC) [8,
[3]. 21–23], ethylcellulose [22, 24, 25], polyvinylpyrrolidone (PVP K-
90) [8, 21, 25], polyvinyl alcohol [8, 23], chitosan
2.2. Semisolid Ophthalmic Drug Forms [21] and its derivatives, like carboxymethyl chitosan [22],
2.2.1. In Situ Gels (or Sol-to-Gel Systems). In situ gels gelatin [24, 26], and various mixtures of the aforementioned
areviscous liquids, showing the ability to undergo sol-to-gel polymers. Employed polymers indicate the division of inserts
transitions when influenced by external factors, like appro- into soluble, insoluble, and biodegradable. A well-known
insert Ocusert (Alza Corporation), built from copolymer of
priate pH, temperature, and the presence of electrolytes. This
ethylene and vinyl acetate, is an example of insoluble insert,
property causes slowing of drug drainage from the eyeball
containing pilocarpine as an active ingredient [5, 8, 27]. The
surface and increase of the active ingredient bioavailability.
main factors limiting the employment of inserts in the therapy
Polymers employed in developing these drug forms include
are still patients’ unwillingness to abandon traditional dosage
gellan gum, poloxamer, and cellulose acetate phthalate, forms, the feeling of foreign body in the eye, and sporadic
whereas active ingredients used in the course of research on in failures in using and introducing inserts, such as unnoticed
situ gels include ciprofloxacin hydrochloride, timolol maleate, excretion from the eye [4, 5, 21, 27].
fluconazole, ganciclovir, and pilocarpine [3–5, 8].
2.3.3. SODI (Soluble Ophthalmic Drug Inserts). SODI
2.2.2. Eye Ointments. Ointments are semisolid dosage
aresoluble eye inserts in the form of small oval wafers,
formsfor external use, usually consisting of solid or semisolid produced from acrylamide, -vinylpyrrolidone, and ethyl
hydrocarbon base of melting or softening point close to human acrylate. After their application to conjunctival sac, they are
body temperature. After applying the ointment to the eye, it moist-ened by tear fluid, and then they soften and adhere to
decomposes into small drops, which stay for a longer time eyeball surface. This dosage form was originally developed
period in conjunctival sac, thus increasing drug’s for astronauts to apply it in the state of weightlessness.
bioavailability. Eye ointments have certain disadvantages— Drug is released from SODI in a pulsational, uncontrolled
although they are well tolerated and safe, they cause, among manner, and the dosage form ensures its prolonged effect.
other things, blurring of vision and sometimes have irritating Active ingredients employed in the course of research on
4 The Scientific World Journal

SODI include neomycin, kanamycin, atropine, pilocarpine, 2.3.8. Minitablets. Minitablets are biodegradable, solid
dexamethasone, sulfapyridine, and tetracaine [4, 28, 29]. drugforms, that, after application to conjunctival sac, transit
into gels, which extends the time period of contact between
2.3.4. Minidiscs/OTS (Ocular Therapeutic System). Minidiscis active ingredient and the eyeball surface, which in turn
a profiled, convex outside, concave from the side of contact increases the active ingredient’s bioavailability [34].
with eye surface, dosage form similar to a contact lens with 4- The advantages of minitablets include easy application
5 mm diameter. Main copolymers from which minidiscs are to conjunctival sac, resistance to defence mechanisms like
developed are - -bis(4-methacryloxy)-butyl tear-ing or outflow through nasolacrimal duct, longer
poly(dimethylsiloxane) and poly(hydroxyethyl methacry-late). contact with the cornea caused by presence of
This dosage form may be either hydrophilic or hydrophobic, mucoadhesive polymers, and gradual release of active
which enables extended time period of release of water-
ingredient from the formulation in the place of application
soluble and poorly water-soluble drugs. Active ingredients
due to the swelling of the outer carrier layers [35, 36].
employed in research on minidiscs were, among others,
The development of minitablets applied to the eyeball
sulfisoxazole and gentamicin sulfate [2, 4, 28, 30].
usually involves using polymers, that is, cellulose derivatives,
like hydroxypropyl methylcellulose (HPMC), hydroxyethyl
2.3.5. Artificial Tear Inserts. This dosage form is a long,
cellulose (HEC), sodium carboxymethyl cellulose, ethyl cel-
rod-shaped pellet, containing no preservatives and
lulose [35, 37, 38], acrylates [35], that is, polyacrylic acid and
developed from hydroxypropyl cellulose. It is available on
the market under the name Lacrisert and is employed in its cross-linked forms, Carbopol or Carbomer [34, 35, 37, 38],
treatment of the dry eye syndrome. After its introduction to chitosan [35], starch, for example, drum-dried waxy maize
conjunctival sac, the insert absorbs water from conjunctiva starch [34, 35], and excipients, that is, mannitol [35, 37, 38],
and cornea, forming a hydrophilic layer, which stabilizes performing the function of solubilizate or sodium stearyl
the tear film and moistens the cornea [2, 5]. fumarate [35, 38] and magnesium stearate [36, 37] with
lubricating properties. Minitablets are developed applying the
2.3.6. Collagen Shield. Collagen shields are developed method of direct compression or indirect method, the latter
fromporcine sclera, whose collagen displays similarities to involving tableting the earlier obtained granules. The
the one in human cornea. T he shields are stored in dry state advantage of indirect method is the dry granulation stage,
and hydrated before they are introduced to the eye. T he which increases flow properties of powders often containing
standard collagen shields, applied by an ophthalmologist, bioadhesive polymers, which enables minitablets production
are not individually suited to the patient’s eyeball and cause on a larger than laboratory scale [34]. Active ingredients from
certain discomfort due to interfering with vision. Moreover, which minitablets were developed include piroxicam [36],
they may be accidentally excreted from the eye just after timolol [35, 37], ciprofloxacin [34, 35, 38], gentamicin, and
introduction [5]. acyclovir [35].
Collagen shields were tested on animal and human
mod-els and may be carriers of antibiotics like gentamicin,
anti-inflammatory drugs like dexamethasone or antiviral 2.4. Multicompartment Drug Delivery Systems
drugs. In comparison to contact lenses and eye drops, the 2.4.1. Nanoparticles and Microparticles. Polymeric,
use of collagen shields enabled obtaining higher drug solid,multicompartment drug delivery systems are
concentration in the cornea and the aqueous humor [4, 30]. promising dosage forms for application to the eyeball. With
More recent dosage forms built from collagen are the
respect to the size of polymeric microvessels, nanoparticles
so-called collasomes, small pieces of collagen (1mm
and microparticles can be distinguished, the former’s size
×2mm ×0.1mm) suspended in a 1% methylcellulose
being from 10 nm to 1000 nm and the latter’s, in case of
vehicle.Collasomes show all advantages of collagen shields
application to the eyeball, from 1 m to 5–10 m [4, 5, 8].
without disadvantages of the latter [5, 30].
Nanoparticles are polymeric carriers, built from
biodegradable, biocompatible, natural, or synthetic polymers
2.3.7. NODS (New Ophthalmic Delivery System). NODS is
with often mucoadhesive properties [39–41]. Ingredients used
adosage form patented by Smith and Nephew
in its development, for the purpose of application to the
Pharmaceuticals Ltd, consisting of solidified paper handle eyeball, were poly(alkyl cyanoacrylate), polylactic acid,
and a flag from polyvinyl alcohol, containing the active poly(epsilon-caprolactone), poly(lactic-co-glycolic acid),
ingredient, attached to the handle with a soluble membrane. chitosan, gelatin [40–42], sodium alginate [41, 42], and
A film containing drug separates from the handle at the albumin [40–42]. These forms can be divided into
point of introduction to conjunctival sac and dissolves in the nanospheres, the solid, monolithic spheres built from dense
tear f luid, releas-ing the active ingredient. This system polymer matrix, in which the active ingredient is scattered, and
ensures delivery of specified drug dose to the eyeball and nanocapsules constituting reservoirs, built from polymer
increased bioavail-ability of active ingredient (even membrane surrounding the drug in solid or liquid form
eightfold in the case of pilocarpine) in comparison to [40]. The mechanism of drug absorption from nanospheres
conventional eye drops. NODS does not contain or nanocapsules after their application to conjunctival sac
preservatives and is sterilized with gamma rays [31–33]. involves diffusion or degradation of the polymer [8].
The Scientific World Journal 5

The pointed-out advantages of using nanoparticles as an of cholesterol and polyethylene glycol) and ethoxylated fatty
ophthalmic dosage form include increased corneal penetra- alcohols (ether of cetyl alcohol and polyethylene glycol). The
tion and a larger dissolution area, which enables improve- size of discosomes is their advantage, because of which they
ment of the active ingredient’s bioavailability in compari- do not enter the general circulation. Moreover, the disc shape
son to traditional eye drops [40]. On the other hand, the ensures better fitting of this form into the conjunctival sac
main pointed-out limitation of nanoparticles is their low [41]. A substantial research has already been conducted on
capacity [8]. niosomal drug forms for substances, that is, ganciclovir
Medicinal substances for which nanoparticle delivery [42], cyclopentolate, or timolol [41].
systems were developed include sulfacetamide,
sparfloxacin, levofloxacin, acyclovir, piroxicam, 2.4.4. Dendrimers. Dendrimers are branched,
cyclosporine A, flurbipro-fen, and pilocarpine [42]. spherical,monodisperse, three-dimensional polymer structures,
of spe-cific size, shape, and molecular mass [45–48]. They
2.4.2. Liposomes. Liposomes are phospholipid drug may be used as carriers, which enclose the active ingredient
carriersusually built of phosphatidylcholine, stearylamine, and inside the polymer structure or create, due to the presence of
vari-ous amounts of cholesterol or lecithin and -L-dipalmitoyl- many functional groups (carboxyl, hydroxyl, and amine),
phosphatidylcholine [5, 41, 43, 44]. The pointed-out advan- electrostatic or covalence bonds with the surface-bound drug
tages of these carriers are their biocompatibility, biodegrad- [46–48]. It has been proved that polyamidoamine (PAMAM)
ability, amphiphilic properties, and relative intoxicity [4, 5, dendrimers, used as carriers for ophthalmic drugs, extend the
41]. However, it is also emphasized that their stability is duration of active ingredients’ effectiveness and increase their
smaller in comparison to therapeutic systems based on bioavailability [48]. Research on using dendrimers as
polymers [5, 8, 41] and that their volume in which drug can be ophthalmic drug carriers was conducted for model sub-
contained is limited [8, 41]. Moreover, their large-scale stances: the pupil dilating tropicamide and pupil constricting
production is expensive and very difficult technologically pilocarpine nitrate. The increased bioavailability of these
[8]. Their employment in ophthalmic drug forms enables substances after application to the eyeball may be in this case
improvement of bioavailability of applied substance and its caused by enclosing the drug inside these structures, which
protection from enzymes present on the surface of corneal results in slower release of the active ingredient. It is also
epithelium [43]. It should be emphasized that effectiveness in explained by their bioadhesive properties [47, 48].
delivery of the active ingredient from liposomes depends on
many factors, that is, encapsulation efficiency, size and charge 2.5. Other Ophthalmic Drug Forms and
of liposomes, stability of liposomes in conjuncti-val sac, or af
Methods of Application
f inity to corneal surface41,[ 43]. Liposomes charged
positively, in comparison to ones charged negatively and 2.5.1. Filter Paper Strips. T hese are paper strips covered
neutrally, display higher affinity to negatively charged corneal withpigments (i.e., fluorescein or Bengal Red) and used in
surface and conjunctival mucoglycoproteins, because of which diag-nostics of corneal, conjunctival, or palpebral damage, as
they slow down the elimination of active ingredient from the well as in diagnosing the presence of microbiological
place of application [41]. In order to increase adhe-sion of infections and eyeball infection (for example with Herpes
negatively and neutrally charged liposomes to corneal or simplex virus) [5, 49, 50]. Every strip of the Fluorets
conjunctival surface, introducing liposome suspensions to preparation, sized approximately 5×15mm, contains 1 mg of
mucoadhesive gels or coating them with mucoadhesive sodium fluorescein
polymers has been proposed [4]. [49]. The strip is usually wetted with a drop of sterile saline
Active ingredients for which liposomal ophthalmic drug solution [49, 50].
forms were being developed include acyclovir, pilocarpine,
acetazolamide, chloramphenicol [43], and ciprofloxacin [44]. 2.5.2. Sprays. Sprays are rarely used ophthalmic dosageforms.
Active ingredients for which they were developed include
2.4.3. Niosomes and Discosomes. Niosomes are cycloplegics, mydriatics, and their mixtures [5, 51], that is,
chemicallystable, built of nonionic surfactants, two-layered phenylephrine-tropicamide and phenylephrine-tropicamide-
carriers used for both hydrophilic and hydrophobic particles, cyclopentolate. Before application to the eye, the distance
without the disadvantages of liposomes (chemical instability, between dosage device and the eyeball should range from 5 to
oxidative degradation of phospholipids, and expensiveness of 10 cm. T he advantage of using these forms is the possibility of
natural phospholipids) [2, 5, 28, 41]. Moreover, these applying the drug on closed eyelid, and the effectiveness of
biodegradable, biocompatible, and nonimmunogenic carriers application is approximately the same as in the case of eye
extend the time period of contact between drug and cornea, drops containing the same ingredients [51]. Results of research
which in turn increases drug’s bioavailability [41]. conducted by Martini and his associates proved that miotic
Discosomes are modified forms of niosomes, which also may effect of pilocarpine hydrochloride applied to the eyeball in the
act as carriers for ophthalmic drugs. Their size varies from 12 form of spray with the active ingredient concentration at 1 to
to 16 m. Discosomes differ from niosomes in that the former 4% is close to the effect achieved after applying eye drops of
contain the addition of nonionic surfactants, Solulan C24, a 1% concentration, with the volume of dose applied in spray
derivative of lanolin, which is a mixture of ethoxylated being 5 L, which was 6 times lower than one applied in eye
cholesterol (ether drops [52].
6 The Scientific World Journal

2.5.3. Ocular Iontophoresis. It is a noninvasive The indirect method (membrane filtration method) is used
procedureduring which ions are introduced to cells or when the character of the product enables it. For water and oil
tissues by use of direct current. When iontophoresis is used solutions, filters from cellulose nitrate are used in which size of
in pharmacother-apy, the aforementioned ions are charged pores does not exceed 0.45 m. For some prod-ucts, for
drug molecules, with positively charged molecule being example, antibiotics, specifically adjusted filters are employed.
introduced to tissue from anode and the negatively charged In the case of testing products with antimicrobial effects, the
one from cathode. Iontophoresis enables fast, safe, and membrane should be washed with chosen sterile solvent not
painless pharmacother-apy and in most cases also obtains less than 3 times, not exceeding the fivefold cycle of filter
high drug concentration in the desired area [5, 53]. Active washing for 100 mL of solvent. The entire membrane is
ingredients that were employed in the course of research on transferred to a suitable medium or is aseptically cut into two
introducing drug using iontophoresis include gentamicin, identical parts, which are transferred into two different media.
dexamethasone, ciprofloxacin, and ketoconazole [53], and In the case of solids soluble in water, the substance should be
it is emphasized that applying antibiotics using this method dissolved in a suitable solvent and the further procedure should
enhances their bactericidal activity [5, 53]. be the same as with water solutions. T he indirect method can
be also used for ointments. Ointments with fatty bases can be
diluted with isopropyl myristate if it is required, at the
3. Examinations of Ophthalmic Drug temperature not higher than 40∘ C. In exceptional situations, the
Forms Properties upper temperature limit may be 44∘ C. Afterwards, the product
is filtered as quickly as possible. For every drug form, after
Examinations which have to be performed in order to filtration and washing, the membrane is transferred to the
determine the properties may be divided into performed medium or the medium is introduced to the filtration set on the
invitro and in vivo. The former determine sterility, the membrane [9, 54].
pH,clarity of solutions, visual assessment, size of the particles,
tonicity/osmolarity, viscosity, amount of substance, amount of 3.1.2. Determining pH. The pH of solutions, drops, sus-
preservative, stability, and in vitro release [9, 13, 17, 26, 42, pensions, and in situ gels is most often determined using a
44, 48, 54, 55]. The latter include the Draize eye test and the in potentiometric method. In this method, the pH value is
vivo release [13, 26, 42, 48, 54, 55]. Other determined by measuring potential difference between
distinguishedexaminations, performed for chosen drug forms, electrodes placed in examined and reference solutions of
include analysis of ions and oxygen permeability for contact known pH or between measurement (glass) electrode and
lenses or determination of encapsulation efficiency for reference (calomel or silver chloride) electrode, both placed
multicompart-ment drug delivery systems and emulsions [13, in examined preparation [9, 44, 48, 54–56].
17, 42, 44].
3.1.3. Clarity Examination. Clarity examination involves
thevisual assessment of formulation in suitable lighting on
3.1. In Vitro Examinations
white and black background. It is performed for liquid forms,
3.1.1. Sterility Examination. The basic requirement for with the exception of suspensions. This examination applies to
drugforms applied on the eyeball is their sterility. Examination eye drops and in situ gels before and after gelling [54, 55].
of sterility involves inoculation in aseptic conditions of the Another method of clarity examination involves trans-
sample examined on two microbiological media: thioglyco- mittance measurement using a UV-Vis spectrophotometer.
late medium (fluid sodium mercaptoacetate or sodium thio- This method can be employed in research on contact lenses
glycolate), which is used for growth of aerobic and filled with active ingredients. The lenses are hydrated in
anaerobicbacteria, and medium with hydrolysate of casein and physiological saline and placed on the surface of quartz
soy (soya-bean casein digest media) used for growth of cuvette. The transmittance is measured afterwards from 200
aerobic bacteria and fungi. A thioglycolate medium with an to 1000 nm wavelength [17].
applied sample is incubated at the temperature of 30–35∘ C,
whereas a medium with hydrolysate of casein and soy with an 3.1.4. Examination of Size and Morphology of Particles.
applied sample is incubated at the temperature of 20–25∘ C for Forexamination of particles’ size multiple methods are
the time not shorter than 14 days. Two methods are employed: optical microscopy (microscopic particle count
distinguished for inoculation of examined material: direct test), light obscuration particle count test, dynamic imaging
inoculation and a method involving use of membrane filters analysis, laser diffraction particle analyzers, electron
[9, 54, 55]. microscopy (SEM, TEM, AFM), DLS (dynamic light
The direct inoculation method, as described in Phar- scattering), Coulter Counter test, and nanoparticle tracking
macopoeia, involves transferring the suitable amount of analysis (NTA).
examined preparation to the medium. If a product has
antimicrobial properties, such effect of the substance should be Optical Microscopy Method (Microscopic Particle Count
neutralized before the examination. Before their introduc-tion Test). Description of this method includes
to the medium, the ointments should be diluted with a suitable requirementsfrom both American and International
sterile solvent containing the chosen surface active agent. Pharmacopoeia. The examination is performed under
During incubation, the media with introduced samples should microscope after taking sample, rinsing, and drying it on
be observed at specified time intervals [9, 55]. microporous membrane
The Scientific World Journal 7

filter with pores’ diameter ≤1 m. This examination enables However, a single measurement does not enable observing
calculating the number of particles sized ≥10 m in examined particles of sizes within the whole range. While observing
products. The test begins from small magnification, for particles of size of lower range limit, it is not possible to
example, ×10 or ×50, at which it is possible to find particles observe particles of size of upper range limit. Flow-based
larger than 25 micrometers. After that, at ×100–×500 systems differ from one another in, among others, the
magnification, smaller particles are being searched for [9, 57, sampling method, the quality of digital image, percentage of
58]. In order to fulfill requirements of American simultaneously analysed particles, and the range of particles’
Pharmacopoeia for formulations, no more than 50 particles per concentration at which measurement is possible. The main
mL may be of size ≥10 m, no more than 5 particles per mL advantages of digital imaging method are a real-time
should be of size ≥25 m, and no more than 2 particles per mL measurement and its conditions, in which particles remain
may be of size ≥ 50 m [59]. On the other hand, the suspended in the liquid. It allows imaging of very irregular
requirements of International Pharmacopoeia stipulate that, for shapes of particles and observing dynamic behaviour of
every 10 m of solid active ingredient, no more than 20 particles under conditions of changing size distribution [57].
particles should be of maximum size larger than 25 m and no
more than 2 of these particles should be of maximum size Laser Diffraction Particle Analyzers. The examination
larger than 50 m. None of these particles can be of maximum involvespassing a laser beam through a sample containing
size larger than 90 m. This method cannot be employed for particles of different shapes, which scatter the light, and the
particles’ analysis in difficult to filtrate solutions of high direction and intensity of scattered light are closely related to
viscosity [9]. the size of particles in examined sample. The diffraction of
light can be described mathematically using the Fraunhofer or
Light Obscuration Particle Count Test. The examination Mie theory. The standard laser light diffraction analyzers
isperformed using a device which counts particles contained in employ detectors whose particle size measurement range is
liquid and employs a light obscuration sensor with a suitable from 0.5 to 2000 m. Using a suitable technology (PIDS—
system dosing the sample to provide controlled portions of polarization intensity differential scattering) enables reduction
sample for analysis. The suspended particles in liquid sample, of the lower range limit of measuring instrument to even about
floating between light source and the sensor, cause changes in 17 nm. One of the biggest disadvantages of laser beam
signal, which are correlated with size of the particles. The scattering technology is the large sample volume. However,
nature of the system which detects and counts particles causes the size of sample is largely related to the concentration of
air bubbles, as well as drops of immiscible liquids, to block particles—as it grows, the required sample volume falls.
sufficient amount of light, because of which they may be Analysis of sample with the use of laser diffraction analyzers
recorded together with suspended particles. The influence of often requires large dilution of samples. It is also important to
these factors on the measurement should be neutralised by its point out that in most of scattered light measurements, the size
suitable technique. This method has certain limitations for of particle of examined sample is determined by calculating
formulations that do not exhibit lucidity and viscosity close to the equivalent spherical diameter, regardless of actual particle
water. Moreover, colour formulations, as well as these with shape [57].
high viscosity, exhibiting changes from shear stress or forming
air or gas bubbles in the moment of contact with the sensor, Electron Microscopy (SEM, TEM, AFM).
for example, products containing bicarbonate buffer, also Advancedmicroscopic methods, such as transmission electron
generate wrong results. For such formulations, in order to microscopy (TEM), scanning electron microscopy (SEM), and
measure size of particles, the membrane microscopy method is atomic force microscopy (AFM), enable high-quality imaging
used. The equipment used for examinations of chosen of particles in nanometer resolution. TEM and SEM require,
formulation should have the maximum range of detected however, strong samples processing. On the other hand, AFM
concentration (maximum number of particles per mL) larger enables capturing the topology of particles’ surface on the
than predicted concentration of examined formulation, image in nanometer resolution. All three methods are
whereas the dynamic range of equipment, that is, range of appropriate largely for specialistic application because of high
sizes of particles for which the size and amount may be equipment costs, low efficiency, and changing conditions of
precisely specified, must include the smallest size of particle sample examination [57, 60–65].
which may be found in examined formulation [59].
DLS (Dynamic Light Scattering) or Photon Correlation
Dynamic Imaging Analysis. This examination Spectroscopy, Quasielastic Light Scattering. T he DLS
enablesmeasurement of size and shape of particles in methodmeasures fluctuations of scattered light caused by
solutions or suspensions. It involves recording the digital Brownian motion of molecules in a solution and is therefore
images of particles suspended in moving fluid, for example, related to diffusion coefficient. From the Stokes-Einstein
during mixing or flow, which enables marking the number equation, knowing the value of diffusion coefficient, it is
of particles in specified volume and specifying particle size possible to determine the hydrodynamic particle radius in
distribution. The lower range of size of particles detected examined sample. It is the radius of the sphere having the same
by optical microscope used in dynamic imaging is about 1 diffusion coefficient as the measured particle. DLS enables
m. The full particle size range possible to observe using simple and quick measurements of particles’ size in the range
this method is from about 1 m to over 1000 m. from <1 nm to even 10 m for one marking. The examinations
8 The Scientific World Journal

can be performed on solutions or suspensions of active Table 1: General conditions for stability examination [6].
ingredients without the need of modification or dilution of
formulation for very small sample volume (10–100 L). Minimum time
DLS is the only method which enables measuring of period covered
Study Storage conditions by data at
particles’ size in the solution in wide range, that is, from
submission
about 0.3 to over 1000 nm, which partly fills the gap of
25∘ C ±2∘ C/60% RH1±5% RH
submicron analysis. This method is mostly used in batch
Long term∗ or 12 months
mode with nonfractionated samples placed in cuvettes or
30∘ C ±2∘ C/65% RH ±5% RH
well plates. Its disadvantages are limitations in size
resolution, lack of shape measurement possibility, and Intermediate∗∗ 30∘ C ±2∘ C/65% RH ±5% RH 6 months
masking light scattering intensity by small particles in the Accelerated 40∘ C ±2∘ C/75% RH ±5% RH 6 months
presence of considerable amount of larger ones. DLS can 1Relative humidity.
resolve two different size groups only when their ∗
It is up to the applicant to decide whether long-term stability studies are performed
hydrodynamic diameters differ 2–5 times [13, 57, 62–66]. ∘ ∘ ∘
at 25 ±2 C/60% RH ±5% RH or 30 C ±2 C/65% RH ±5% RH.
∗∗ ∘
If 30 C

±2 C/65% RH ±5% RH is the long-term condition, there is no intermediate
Coulter Counter. T his method employs the rule whichsays
condition.
that particles placed in electric field modify the flow of
charge (current). For detecting particles, the electrical
sensing zone technique is used. The method of determining
size and amount of particles using Coulter Counter is between the active ingredient and other ingredients of the
described in ISO 13319 norm (“Determination of particle
preparation [21, 55, 62].
size distribution—electrical sensing zone methods”). The
particles’ size measuring range in this method is from about
0.4 m to 1600 m. Several markings enable detecting particles 3.1.7. Stability Examination. The purpose of stability exam-
in the whole measuring range. The main advantage of this ination is to provide information on changes in quality of
method is the fact that particles’ properties, that is, colour, active ingredient or medicinal product in time due to the
shape, composition, or refractive index, do not affect the effect of environmental factors, that is, temperature, humid-
measurement. Using Coulter Counter, it is possible to obtain ity, and light, on examined substance/product, as well as to
very precise particles’ size distribution. Before the set the date of further examination of medicinal substance
measurement, a formulation containing particles must be or expiry date of medicinal product and recommended
suspended in electrolyte, which may cause changes in storage conditions [6].
composition or number of particles [57, 67, 68]. General stability requirements for ophthalmic products, for
example, drops and ointments, are similar to those for other
Nanoparticle Tracking Analysis (NTA). NTA is a new tech- pharmaceutical products. They are harmonized through ICH
nique employed for measuring size of particles in the range (International Conference on Harmonisation) process in USA,
from about 30 to 1000 nm. It combines laser light scattering Europe, and Japan, acknowledging the contribution of a
microscopy with a CCD camera, which enables visualization European institution EMEA (European Agency for the
and recording particles in a solution. The examination, as in Evaluation of Medicinal Products) and its Committee for
the DLS method, involves determining size of the particle Proprietary Medicinal Products (CPMP), QWP (Quality
from Stokes-Einstein equation. NTA exhibits more precision Working Party), and the American institution FDA (Food and
in size distribution in comparison to DLS but requires larger Drug Administration) as well as the Japanese Ministry of
volume of the sample (about 300 L) [57, 69]. Health [71]. Generally, active ingredients should be stored in
conditions that enable assessment of their ther-mal stability
and, if applicable, also proneness to humidity. Storage
3.1.5. Examination of Content of Substance or conditions and examination period should correlate with
Preservative. The examination of drug or preservative warehousing, transport, and later use conditions [6].
content in given formulation is labeled with relevant There are many documents containing guidelines on
analytical technique, that is, spectrophotometric method or stability examinations. However, they are general and often do
HPLC [12, 21, 26, 54, 55, 61, 70]. not acknowledge special features of ophthalmic products.
Matthews and Wall, in their article, referenced (with a short
description) the documents which may constitute footholds for
3.1.6. Examination of Drug and Carrier Interaction/Compat- planning stability examinations of ophthalmic products,
ibility Using FTIR, DSC, and XRD Methods. Fouriertransform particularly those different from conventional drops and
infrared spectroscopy (FTIR) and examinations employing ointments [71]. General conditions for stability examination
differential scanning calorimetry (DSC) and X-ray are contained in Tables 1, 2, and 3.
diffractometry (XRD) are performed for, among others, pure Despite existing guidelines, the scientists often choose
substance, physical mixtures of drug and polymers used to their own conditions for stability examinations. Nagargoje
obtain formulation, and the ingredients of the formulation in with associates performed stability examinations for an in situ
order to identify potential interactions gel containing fluconazole at temperatures 4∘ C ±1∘ C, 27∘ C
±1∘ C, 45∘ C ± 1∘ C for one month period [55], and Nanjwade
The Scientific World Journal 9

Table 2: Conditions for active ingredients stored in refrigerators [6]. stirrer. In specified time intervals, samples of dissolution
medium are taken, and the medicinal substance is marked
Minimum time
Study Storage conditions period covered by
using a suitable analytical technique. The taken sample
data at submission
amount is replaced with analogical amount of a fresh solution
simulating a tear f luid or phosphate buf fer 44,[ 61, 78, 79].
Long term 5∘ C ±3∘ C 12 months
∘ ∘
Accelerated 25 C ±2 C/60% RH ±5% RH 6 months Modified Rotating Basket Method. In this method, a drugform
is placed in a set of baskets or substitutes, for example, glass
Table 3: Conditions for active ingredients stored in freezers [6]. cylindrical pipes, connected with a stirrer. The glass pipes are
covered with dialysis membrane on one side, while the other
Minimum time period
Study Storage conditions covered by data at side is attached to shafts of the apparatus. All the components
submission are put in a beaker with a water jacket, containing a buffer
solution, for example, a simulated tear fluid (STF).
Long term −20∘ C ±5∘ C 12 months
Temperature of the system may be maintained, for example, at
35∘ C ± 1∘ C, and the frequency of stirrer rotation may be at,
with associates examined eye drops stability at for example, 50 rpm. A sample of solution is taken in specified
temperatures 5∘ C, 25∘ C, 37∘ C, and 45∘ C [54]. time intervals and examined for drug amount. The taken
sample amount is supplemented with the analogical amount of
a fresh solution simulating a tear fluid in order to keep
3.1.8. Drug Release Studies. In literature, several
methodsemployed for the examination of accessibility of constant volume [80].
pharmaceutical substance from ophthalmic forms were
Modified Rotating Paddle Apparatus. In this method,
described. They include bottle method, modified rotating
diffusionchambers, used for analysis of half-solid
basket method, diffusion method with the use of Franz cell,
formulations, are placed in a paddle apparatus container. A
modified rotating paddle apparatus, or method with the use
suitable liquid is poured into the container and stirred
of flow-through device.
during test at the speed, for example, of 50 rpm, at the
Bottle Method. In this method, the examined drug formsare temperature of 37∘ C. Containers with diffusion chambers
placed in culture bottles [72, 73] or vials [21, 38, 74, 75] soaked in dissolution medium are placed in a water bath,
containing phosphate buffer at pH 7.4 [21, 38, 72–74, 76, maintaining the temper-ature at 37±0.5∘ C. Samples of
77] or artificial isotonic tear fluid [75]. Bottles and vials are buffer solution into which the substance from diffusion
usually shaken in water baths [21, 38, 72–74] (or incubated chambers is being released are taken in specified time
under magnetic stirring [76, 77]), mostly at a temperature intervals and examined for drug content [72, 81].
of 37∘ C [72–74, 76, 77], and the medium samples are taken Kao and associates employed this method for
in specified time intervals and examined for drug amount examination of substance release from nanoparticles
using a suitable analytical method [21, 38, 72–77]. introduced directly to a paddle apparatus container holding
a solution simulating tear fluid, stirred at the speed of 75
Diffusion Method with the Use of Franz Cell or Other Two- rpm at the temperature of 37∘ C. In specified time intervals,
Compartment Systems. T his method employs a two- they took solution samples, centrifuged them, and marked
chambersystem consisting of two compartments: donor and spectrophotometrically in a supernatant the amount of
receiver. A sample of examined formulation is placed in a
active ingredient [62].
donor compartment of Franz cell or other systems, while a
receiver compartment contains a thermostated dissolution Flow-Through Devices. In this technique, an apparatus inwhich
medium, for example, at the temperature of 37∘ C ± 0.5∘ C, permanent dissolution medium circulation takes place is
subjected to continuous stirring using a magnetic stirrer, employed for substance release studies. The device consists of
usually at the speed of 50 rpm. Both compartments are a cell in which the substance is dissolved (a jacketed flow-
separated with a dialysis membrane, for example, made from
through cell), a continuous duty oscillating pump, a water bath,
cellophane. During examination, in specified time intervals,
and a jacketed flask containing a dissolution medium. A drug
samples of dissolution medium are taken, and the medicinal
form is put in the jacketed flow-through cell, into which a
substance is marked using a suitable analytical technique [55,
dissolution medium is introduced afterwards. The medium
56, 65, 72].
For release tests in the described method, a glass container, circulates in closed cycle. Temperature is maintained at a level
for example, of a cylindrical shape, may be used. It is placed in close to that of human body (e.g., 33±2∘ C or 37∘ C) and the
a beaker (a receiver compartment) [44, 61, 78, 79], f illed with samples are taken in specif ied time intervals and are examined
an artif icial tear f luid78,[ 79] or phosphate buf fer at the pH of for drug content [72, 82].
7.4 [44, 61]. In the cylindrical container, constituting a donor Examinations of active ingredients release from drug
compartment, an examined drug form is placed, after which a forms may be performed also in flow-through devices with
diffusion cell membrane is put on a containers aperture. The open flow, which was described in articles written by Rao
ingredients of the compartment are continuously stirred at and Shyale [83] as well as Tanwar and associates [84].
fixed temperature using a magnetic
10 The Scientific World Journal

Other examinations performed for ophthalmic drug For moisture absorption examination, a specified
forms include viscosity examinations using viscometers number of inserts are chosen and placed in desiccator, in
[44, 48, 55, 56], osmolarity examinations using which high moisture level, for example, 75±5% RH, is
osmometers [44, 48, 56], and the light refractive index maintained. After a specified time period, inserts are taken
measurement using ellipsometers/refractometers [20, 48]. out and weighed again, and the percentage moisture
absorption is calculated from the formula [21, 70, 85]
3.2. Other Examinations Performed for Chosen Drug Forms
% Moisture Absorption
3.2.1. Examinations for In Situ Gels (Final weight −Initial Weight)×100 (2)
= Initial Weight .
Examination of Gel-Forming Ability. This examination
isperformed in order to assess the ability of formulation to
In moisture loss examination, a chosen number of
form gels on the surface of eyeball. A sample of examined
inserts are put in desiccator containing anhydrous calcium
formulation is introduced to a vial containing a solution whose
chloride, which ensures dry conditions inside the container.
components simulate a tear fluid and visual technique is
After a suitable time period, inserts are taken out and
employed to assess the sol-gel phase transition [55, 61].
weighed again, and the percentage moisture loss is
calculated from the formula [21, 85]
3.2.2. Examinations for Inserts

Swelling Index. Hydrophilic polymers of different


% Moisture Loss = (Initial Weight −Final weight)×100 .
Initial Weight
structuresexhibit different swelling degree, depending on
(3)
relative resis-tance of matrix network structure to water
particles’ move-ment. Polymer chains exhibiting low ability to For eye inserts assessment, examinations of thickness
form hydrogen bonds may not be able to form strong network [26, 70, 79] and weight uniformity [26, 70], as well as
structure, resistant to fast water penetration. The bigger the mechanical strength tests [70, 79], are also advisable.
strength and number of hydrogen bonds between polymer
chains are, the slower the water particles diffuse into the
hydrated matrix. Swelling of the polymer is vital to activation 3.2.3. Examinations for Multicompartment
of bioadhesive abilities, which activate just after swelling Drug Delivery Systems
begins. With the growth of polymer hydration, the adhesion
Encapsulation Efficiency. A sample for encapsulation ef f i-
grows until the moment when excessive hydration leads to
ciency examination is obtained by centrifuging [62, 64, 65] or
sudden fall of adhesion strength, which is an effect of the
untangling of outer polymer layer. The degree and speed of
centrifugal ultrafiltration [13, 44] of mixture formed af ter
insert hydration, as well as swelling, affect drug release from a preparing the formulation. The obtained supernatant or f
dosage form. Therefore, this parameter is of greatest iltrate is examined for amount of free active substance using a
significance for drug release prediction and bioadhesive matrix spectrophotometric method [44, 62, 64] or HPLC [13, 65].
potential. Swelling examination is performed to measure bulk Encapsulation efficiency is calculated from the formula
hydrophilicity and polymer hydration [21]. In the procedure, a
specified number of inserts are chosen, weighed, and put E.E.(%)= ( total
− free
)× 100 , (4)
total
separately in beakers containing a solution simulating tear
fluid [78], physiological saline buffered with phosphates [21], wheretotal is the total amount of drug in the formulation;free is
or distilled water [85] at fixed temperature, for example, 32∘ C
± 0.5∘ C
the amount of drug in the filtrate/supernatant [13, 62].
[21]. In specified time intervals, inserts are taken out, dried
with filter paper, and weighed once more. The procedure is 3.3. In Vivo Examinations
repeated until the moment when mass growth is not observed 3.3.1. Eye Irritancy Test (Draize Eye Test). There are
anymore [21, 78, 85]. The degree to which the liquid is taken manymodifications of eye toxicity/irritancy test (Draize eye
up, called the swelling index, is calculated from the formula test) performed for dosage forms, that is, solutions, emulsions,
ointments, solids, for example, inserts, and so forth. Exami-
( − 0) nations are usually carried out on rabbits, whose vision organ
Swelling index = [ ]×100, (1)
0 anatomy and physiology are well described in literature.
Moreover, rabbits’ eyes are usually more susceptible to irri-
where 0 is the initial sample weight and is the sample tating compounds than those of humans. For the test, usually
weight at time [21]. from 3 to 6 rabbits are used, which, on one hand, enables
obtaining reliable results, and, on the other hand, is an answer to
Examinations of Moisture Absorption and Loss. These claims for applying toxic substances to as little animals as
exami-nations are performed in order to assess physical possible. T he most of ten used animal subspecies are albino
stability and integrity of inserts’ polymer matrix in dry (e.g., New Zealand) rabbits, which are examined and weighed
conditions and at raised moisture [21, 85].
The Scientific World Journal 11

before the test and then placed in specifically adapted cages, 3.3.3. In Vivo Release Evaluation of Inserts. For in vivo
designed so as to avoid accidental injuries. The examined releaseevaluation, formulations which gave desired results
preparations are introduced to conjunctival sac or applied in in vitro release evaluations are chosen. Inserts are put in
directly on the cornea. At first, about 0.1 mL of analyzed drug conjunctival sacs of healthy rabbits chosen for studies. In
was being applied on the eyeball, but many later examinations specified time intervals, inserts are carefully taken out and
pointed to reducing the amount, for example, to 0.01 mL, which examined for left drug amount using a suitable analytic
more reflects real situations. In the test, one eyeball, usually the technique [24, 26, 81, 84].
lef t one, is used as a control. Af ter introducing a drug form on
the eyeball, the eyelids are usually kept closed for a few 4. Conclusions
seconds, although it is not required. Sometimes sterile solutions
are additionally used for rinsing the eyeball surface. An Despite many achievements in the f ield of ophthalmic dosage
assessment of eyeball condition before and after introducing the forms, still vast majority of active substances for use in ocular
formulation is done by observation of the eyeball in suitable disorders are in the form of eye drops. Some of the more
light, of ten using magnifying glass or a slit lamp, which complex forms appeared on the pharmaceutical market, such as
ensures more precise evaluation. Auxiliary procedures which Ocusert by Alza Corporation, but scientists are still looking for
simplify visualization of changes include dyeing with the perfect ophthalmic system, which would possess desired
fluorescein and taking photos of eyeball. More-over, the properties such as controlled release, minimizing systemic
discomfort level after application may be indicated by the effects, ease of use, and extended retention time at the site of
number of blinkings or rubbings of the eye. The evaluation application. Multicompartment systems appear to be promising
drug forms that can also be combined with other forms, for
takes place usually after 1 h, 24 h, 48 h, and 72 h from
example, polymeric nanoparticles with the active substance
introducing a drug form on the eyeball and, if essential, also
suspended in the in situ gel.
after 7 or 21 days. Duration of examination, as well as its
In connection with the development of new ophthalmic
scheme, is individually adapted to the analyzed formulation.
dosage forms, a problem concerning the analysis of their
Ocular changes are assessed using a scoring system, in which
physicochemical properties and in vitro-in vivo correlation
every change in the area of eyelid, conjunctiva, cornea, and iris appears. This paper is a review of the available literature
is scored. While in literature many scoring systems were which allows planning studies to be conducted on standard
proposed, the modified Friedenwald and Draize methods are and modern ophthalmic drug forms.
still widely employed [21, 48, 54–56, 64, 86].

Conflict of Interests
3.3.2. Transcorneal Permeation Study. For transcorneal per- The authors declare that there is no conflict of interests
meation study, as in the Draize eye test, healthy albino rabbits regarding the publication of this paper.
are chosen in the number which is suitable for obtaining
reliable results. The amount of active substance in aqueous
humor after introducing the formulation to conjunctival sac is References
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