THESIS NASAL GEL

8/12/2019 THESIS NASAL GEL

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PREPARATION AND EVALUATION OF IN SITU

OPTHALMIC GEL OF AN ANTI INFECTIVE DRUG FOR

SUSTAINED OCULAR DELIVERY

yMr. RAJAS N.JB. Pharm.,

Reg. No.09PU330

Dissertation Submitted to theRajiv Gandhi University of Health Sciences, Karnataka, Bangalore

In partial fulfillment of the requirements for the degree of

MASTER OF PHARMACY

IN

PHARMACEUTICS

Under the guidance of

Mrs. K. KAVITHAM. Pharm.,

Department of Pharmaceutics

Bharathi College of Pharmacy

Bharathinagara

2011


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RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,

KARNATAKA, BANGALORE

DECLARATION BY THE CANDIDATE

I hereby declare that the matter embodied in the dissertation entitled

PREPARATION AND EVALUATION OF I N SITU OPTHALM IC GEL OF AN

ANTI INFECTIVE DRUG FOR SUSTAINED OCULAR DELIVERY is a

bonafide and genuine research work carried out by me under the

guidance of Mrs. K Kavitha M.Pharm.,Department of Pharmaceutics,

Bharathi College of Pharmacy, Bharathinagara. The work embodied in

this thesis is original and has not been submitted the basis for the award

of degree, diploma, associate ship (or) fellowship of any other university

(or) institution.

Date:

Place: Bharathinagara Mr. RAJAS N.JB. Pharm.,


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BHARATHI COLLEGE OF PHARMACY

BHARATHINAGARA-571422

CERTIFICATE BY THE GUIDE

This is to certify that the dissertation entitledPREPARATION AND

EVALUATION OF IN SITU OPTHALMI C GEL OF AN ANTI INFECTIVE

DRUG FOR SUSTAINED OCULAR DEL IVERYis abonafide research work

carried out by Mr. Rajas N.J submitted in partial fulfillment for the

award of thedegree ofMaster of Pharmacy in pharmaceutics by the

Rajiv Gandhi University of Health Sciences, Karnataka, Bangalore.

Date: Mrs. K. KAVITHA M Pharm.,Department of Pharmaceutics,

Place: Bharathinagara Bharathi College of Pharmacy,

Bharathinagara571422


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ENDORSEMENT BY THE HEAD OF THE

DEPARTMENT



DRUG FOR SUSTAINED OCULAR DEL IVERYis a bonafide research work

carried out by Mr. Rajas N.J submitted in partial fulfillment for theaward of thedegree ofMaster of Pharmacy in Pharmaceutics by the

Rajiv Gandhi University of Health Sciences, Bangalore, Karnataka. This

work was carried out by him in the library and laboratories of Bharathi

College of Pharmacy, under the guidance ofMrs. K. KavithaM.Pharm.,

Department of Pharmaceutics, Bharathi College of Pharmacy,

Bharathinagara.

Date: Dr. T. SIVAKUMAR M Pharm., Ph. D.Professor and HOD,

Place: Bharathinagara Department of Pharmaceutics,

Bharathi College of Pharmacy,

Bharathinagara571422


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ENDORSEMENT BY THE PRINCIPAL / HEAD OF THE

INSTITUTION



DRUG FOR SUSTAINED OCULAR DEL IVERYis abonafide research work

carried out by Mr. Rajas N.J submitted in partial fulfillment for the

award of thedegree ofMaster of Pharmacy in Pharmaceutics by the

Rajiv Gandhi University of Health Sciences, Karnataka, Bangalore. This

work was carried out by him in the library and laboratories of Bharathi

College ofPharmacy, under the guidance ofMrs. K.Kavitha M.Pharm.,

Department of Pharmaceutics, Bharathi College of Pharmacy,

Bharathinagara.

Date: Dr. T. TAMIZH MANIM.Pharm., Ph.D.Principal,

Place: Bharathinagara Bharathi College of Pharmacy,

Bharathinagara571422.


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OPYRIGHT

DECLARATION BY THE CANDIDATE

I hereby declare that the Rajiv Gandhi University of Health

Sciences, Karnataka shall have the rights to preserve, use and

disseminate this dissertation / thesis in print or electronic format for

academic / research purpose.

Date:

Place: Bharathinagara Mr.RAJAS N.JB.Pharm.,

Rajiv Gandhi University of Health Sciences, Karnataka.


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DEDICATED TO MY

BELOVED FAMILY

AND FRIENDS


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CKNOWLEDGEMENT

Gratitude makes sense of our past, brings peace for today and creates a

vision for tomorrow

Many Thanks toAlmighty Godfor it, who began this work i n me and

carr ied it to completion, who has blessesed me with the people whose names I feel

pri vileged to mention here.

The completion of th is disser tation i s not only ful fi llment of my dreams, but

also the dreams of my Parentswho have taken lots of pain for me in completion of

my higher studies.

I consider th is as an opportuni ty to express my gratitude to all the

dignitaries who have been involved directly or indirectly with the successful

completion of th is dissertati on.

At f i rst, I consider i t as a great pr ivi lege to express my heartfelt grati tude

and sincere thanks to my esteemed guide Mrs. K.Kavitha,Assistant Pr ofessor,

Department of pharmaceutics, Bharathi college of pharmacy, for her valuable

suggestions, encouragement, motivation, guidance and co-operation dur ing my

thesis work.

My deepest appreciation and heartf elt thanks toDr . T. SivakumarHead of

Department of Pharmaceuti cs for h is valuable suggestions and help in making my

study a success. I cher ish h is co-operation throughout my l if e.

I consider myself to be very for tunate to haveDr . Tamizh ManiPrincipal

Bharathi coll ege of pharmacy, Bharathinagara, who with his dynamic approach

boosted my morale, which helped me in the completion of thi s dissertation .


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I take this golden opportun ity to express my heartful grati tude and respect

Dr .K.P Channabasavaraj, M r.Ganesh, and Mr.Rupesh Kumarfor his constant

guidance and encouragement thr oughout my project work.

I take this opportuni ty to express my deep grati tude and heartfelt thanks

Mr. K.M Prasad, ManagerKarnataka Anti biotics and Pharmaceuticals L imi ted,

Bangalore for providing opportunity to do project work in their reputed

organization.

I take this opportuni ty to express my gratefu lness toMs.Parimala, Manager

Formulation Development and Mr.Th imaraya swamyKarnataka Antibiotics and

Pharmaceuti cals L imited, Bangalore for the valuable guidance dur ing the course of

study.

I take this opportun ity to express my gratefu lness toM r.Marigoli, Manager

Quali ty control Department, Ms.Sharmila, Ms.Uma rani, M s.SudhaKAPL for the

valuable guidance dur ing the course of study.

Thanks is a small word to my Father NS Jayaprakash, mother Gayathri

devi, my brothers Raghavendra, Rakesh, my Ati gae Vidyaand all fami ly members

for their encouragement and love that served as a source of i nspiration, strength at

each and every front of my li fe to transport my dreams in to reali ty.

I cordiall y thank to M r.Santhosh, Mr.Sumukhfor their suggestion, support

and encouragement during throughout my studies.

I special ly convey my grati tude to dearest class mates Mr.Mehaboob,

Mr.sandeep, M r.Pavan, Ms. Ashvin i, Mr.Vinay, M r.M angesh, Mr .kiran, M r.M ehul ,

Mr.Dipenfor their timely help, support and memorable company dur ing my course.


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I special ly convey my gratitude to my dearest f r iendsM r.Pavan, M r.Rahul,

Ms.Nisha, Ms.Bharathi, Mr.Vinod, Mr.Bharthesh, Mr.Almesh, Mr.Naveen,

Mr.Mithun, Mr.Bhadri, Mr.Ranjan, Mr.Sunil, Mr.Raghavendra, Mr.Omkar,

Mr.Ravi, Mr.Ajay, Mr.Shashi, Mr.Chiranjivi, Mr.Gowrish, Mr.Dilshad,

Mr.Phaniraj, Ms.Ramya, Ms.Sowmya, Mr.Srikanth, Mr.Vikshith, Mr.Nandhan,

Mr.Manjunath, Mr.Sridhar and all my friends for their continuous support

encouragement th roughout my studies.

I am proud to say it was a fru itf ul and enjoyable exper ience to work with

Mr.Lohith, M r.Ramachandra, Mr .Yusuf , Mr.Guru, M r.Deepak, Mr.Shivakumar

reddy, Mr.Rajufor their continuous support and encouragement throughout my

project in KAPL.

I wish to express my sincere thanks to my seniorsMs.Veena, M r.An il ,

M r.punnet, Mr.Shivaraj , Mr .Panner selvamand all my juniors for their help during

my project work.

I take this golden opportuni ty to express my thanks toManagementof

Bharathi College of pharmacy for providing great environment dur ing my studies.

I t is indeed a dif fi cul t task to acknowledge the services of all those gentl e

people who have extended their valuable suggestion and support directly or

indi rectly whose names have been unable to mention as they are like the coun tless

stars in numerous galaxies.

My Sincere thanks to all .

Date: Rajas N.J B.Pharm

Place: Bharathinagara


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LIST OF ABBREVIATIONS

Department Of Pharmaceutics Bharathi College Of Pharmacy

IP : Indian pharmacopoeia

BP : British pharmacopoeia

CAP : Cellulose acetate phthalate

FTIR : Fourier transform infrared

LEV : Levofloxacin hemihydrate

HPMC : Hydroxy propyl methyl cellulose

r2 : Regression coefficient

SD : Standard deviation

SRDF : Sustained release dosage form

USP : United states pharmacopoeia

HPC : Hydroxy propyl cellulose

Rpm : Revolutions per minute

UV : Ultraviolet

Hrs : Hours

Avg : Average

G : Gram

mg : Milligram

mm : Milli meter

g : Microgram

pH : Negative logarithm of hydrogen ion concentration

min : Minutes

Conc. : Concentration



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Department Of Pharmaceutics Bharathi College Of Pharmacy

NDDS : Novel Drug Delivery system

Cps : Centipoise

STF : Simulated tear fluid.

F1 : in situ gel prepared with gelrite concentration 0.2%






SD : Standard deviation


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ABSTRACT

Department of Pharmaceutics Bharathi College of Pharmacy

OBJECTIVE

Ocular bioavailability is always poor from conventional ophthalmic drops due

to spillage and nasolachrymal drainage. Ocular in situ gels can increase the drug

residence time thus increasing bioavailability. Purpose of current study is to prepare

sustained release in situ ocular gels of Levofloxacin hemihydrate using gelrite as

gelling polymer, which is used in treatment of various bacterial infections.

METHODS

In situ gels of LEV were prepared by applying principle of ion exchange and

temperature dependant gelation techniques using gelrite as the gelling polymer. In the

present study six different formulations were prepared containing varying

concentrations of gelrite and the formulations were evaluated for physical parameters

like clarity, pH, drug content, gelation, sterility test, rheological studies,in vitrodrug

release study and ocular irritancy studies, anti microbial efficiency study and stability

studies.

RESULTS

FTIR spectras revealed that, there was no interaction between LEV and

excipients. The formulated gels were transparent, uniform in consistency and had

spreadability with a pH range of 7.1 to 7.4. Rheological studies revealed that the

formulations were psuedoplastic in nature, drug content of sterile in situ gels was

found to be 92-98%. From the preliminary studies it was observed that as the

concentration of the polymer was increased, the rate of drug release decreased to

produce sustained drug delivery for more than 8 hours with a maximum of 90.2%

ABSTRACT


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ABSTRACT


drug release. Release kinetic study showed that the formulation followed first order

diffusion controlled and non fickian release mechanism, the optimized formulations

was having good antibacterial efficacy, results of ocular irritancy studies showed that

formulations were non irritant and stability studies indicated that formulations were

stable at room temperature as well as at 400C.

CONCLUSION

The present study conclusively demonstrates the feasibility of effectively

formulating Levofloxacin in situ gels using gelrite as release retardant to form a

potentially effective sustained release drug delivery system.

Keywords: Gelrite; Rheological; ocular; gels; Bioavailability; Anti microbial; in vitro


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CONTENTS


CHAPTER

NO

TITLE PAGE

NO

1 Introduction 1

1.1 Ocular anatomy and physiology 2

1.2 Absorption and bioavailability of the drugs from

the eye10

1.3 General bacterial infections of eye and their

medications12

1.4 Ocular drug delivery systems 14

1.5 Methods of preparation of in situ gels 19

2 Aims and objectives 26

2.1 Plan of work 27

3 Review of literature 29

3.1 Drug profile 37

3.1.1 Levofloxacin Hemihydrate 37

3.2 Polymer profiles 40

3.2.1 Gelrite 403.2.2 Propylene glycol 41

3.2.3 Benzalkonium chloride 43

4 Materials and Methods 44

4.1.Materials 44

4.2 Methods 45

4.2.1 Preformulation studies 45

4.2.2 Preparation of in situ gel of LEV. 464.2.3 Evaluation of prepared in situ gelling system 47

5 Results 54

6 Discussion 79

7 Conclusion 82

8 Summary 83

9 Bibliography 85

10 Annexures 94

CONTENTS


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CONTENTS



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LIST OF TABLES


TABLE

NO

TITLE PAGE

NO

1.1 Topical antibacterial agents for ophthalmic use. 14

4.1 List of chemicals used with grade and supplier. 44

4.2 List of instruments used. 45

4.3 Formulation of in situ gels of LEV. 46

5.1 Interpretations of IR spectra. 57

5.2 Preliminary evaluation of visual appearance, clarity, pH, and

drug content.

58

5.3 Evaluation of gelling capacity. 59

5.4 Rheological studies of in situ gels before gelation. 60

5.5 Rheological studies of in situ gels after gelation. 61

5.6 Test of sterility. 62

5.7 Standard calibration of LEV. 63

5.8 In vitrorelease profile of marketed eye drops. 64

5.9 Comparative in vitrorelease of marketed eye drop and prepared

in situ gels.

65

5.10 Comparative Zero order release kinetics data of in situ gels. 66

5.11 Comparative First order release kinetics data of in situ gels. 67

5.12 Comparative Higuchi release kinetics data of in situ gels. 68

LIST OF TABLES


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LIST OF TABLES


5.13 Comparative Peppas release kinetics data of in situ gels. 69

5.14 Regression co-efficient (r2) values of different kinetic models. 72

5.15 Antimicrobial activity of in situ gels. 73

5.16 Stability studies of Formulation F1. 74






5.22 Stability studies of all formulation at room temperature. 77

5.23 Stability studies of all formulations at 40 c. 77


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LIST OF FIGURES


FIG. NOTITLE PAGE NO

1.1 Different Parts of human eye. 3

1.2 Routes of ocular absorption of drugs. 10

1.3 Classification of hydrogels. 18

1.4 PEO-PPO-PEO (Poloxamer). 20

1.5 In situ gel formation due to change in temperature. 21

1.6 Structure of Carbomer. 22

1.7 Schematic representation of pH dependent in situ gels. 23

1.8 Structure of deacetylated gellan gum. 24

5.1 IR spectra of LEV 55

5.2 IR spectra of Gelrite. 55

5.3 IR spectra of physical mixture of LEV and Gelrite. 56

5.4 IR spectra of in situ gel of Levofloxacin hemihydrate. 56

5.5 Rheological studies in situ gels before gelation. 60

5.6 Rheological studies in situ gels after gelation. 61

5.7 Calibration curve of Levofloxacin hemihydrate. 63

5.8 In vitrorelease profile of marketed eye drops. 64

5.9 Comparative in vitrorelease of marketed eye drop and

prepared in situ gels.70

5.10 Comparitive zero order release kinetics of in situ gels. 70

5.11 Comparitive first order release kinetics of in situ gels. 71

5.12 Comparative higuchi release kinetics of in situ gels. 71

5.13 Comparative peppas release kineticsof in situ gels. 72

5.14 Stability studies of in situ gels at room temperature. 78

5.15 Stabilitystudies of in situ gels at 40C. 78

LIST OF FIGURES


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CHAPTER 1 INTRODUCTION

Department of Pharmaceutics Bharathi College of Pharmacy 1

Eye is unique and vital organ. It is considered as window of the soul. We can

enjoy and view the whole world only with this organ. There are many eye ailments

which affect this organ and one even can loss the eye sight. Therefore many

ophthalmic drug delivery systems are available. These are classified as conventional

and newer drug delivery systems. Eye drops that are conventional ophthalmic delivery

systems often result in poor bioavailability and therapeutic response, because high

tear fluid turnover and dynamics cause rapid precorneal elimination of the drug. A

high frequency of eye drop instillation is associated with patient non-compliance.

Inclusion of excess drug in the formulation is an attempt to overcome bioavailability

problem, is potentially dangerous if the drug solution drained from the eye is

systemically absorbed from the nasolacrimal duct. Various ophthalmic vehicles such

as inserts, ointments, suspensions and aqueous gels have been developed in order to

lengthen the residence time of instilled dose and enhance the ophthalmic

bioavailability. These ocular drug delivery systems however have not been used

extensively because of some drawbacks such as blurred vision from ointments or low

patient compliance from inserts.

Several in situ gelling system have been developed to prolong the precorneal

residence time of a drug and improve ocular bioavailability. These systems consist of

polymers that exhibit sol to gel phase tansititions due to change in specific physico

chemical parameter (pH, temperature) in their environment, the cul de sac in this case.

Depending on the method employed to cause sol-to-gel phase transition on the eye

surface the following three types of systems are recognized, pH triggered system,

1. INTRODUCTION


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temperature dependant system and ion activated system. Using these three methods

above in situ gelling ophthalmic delivery system is developed.1

Topical anti infectives are commonly used to treat bacterial conjunctivitis and

infection of cornea caused by susceptible strains of bacteria such as S. aureus, S.

epidermidis, S. pneumoniae, Enterobacter cloacae, H. influenzae, P. mirabilis and P.

aeruginosa. They are also indicated for the treatment of bacterial corneal ulcers

caused by susceptible strains of the following bacteria: S. aureus, S. epidermidis, S.

pneumoniae, P. aeruginosa, S. marcescens and Propionibacterium acnes etc.2

1.1 OCULAR ANATOMY AND PHYSIOLOGY

The human eye is challenging subject for topical administration of the drugs.

The basis of this can be found in the anatomical arrangements of surface tissue and in

permeability of the cornea. The protective operation of eyelids and lacrimal system

are such that there is rapid removal of material instilled into eyes unless the materials

are suitably small in volume and chemically and physiologically compatible with

surface tissues.3

The eye is referred to as a globe, is actually two spheres, one set in the other, as

shown in Fig. 1.1. The front sphere is the smaller of the two and is bordered anteriorly

by the cornea, whereas the larger posterior sphere is an opaque fibrous shell encased

by the sclera. The combined weight of both spheres has been given as 6.77-7.5 g, with

a volume approximately 6.5 ml. The circumference of the eye is about 75 mm. Along

with the rest of the orbital contents; the eye is located within the boney orbital cavity

of the head.4


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Fig.1.1 Different parts of human eye.

A.Structural support

The orbit

The eyes rest in two boney cavities; i.e, the orbits, located on either side of the

nose. The anterior two-third of the orbit is roughly the shape of a quadrilateral

pyramid, where as the posterior one-third of the orbit narrow to the shape of a

triangular pyramid. The globe occupies approximately 20 % of the cavity, lying

slightly nearer the upper and lateral sides but never in contact with the orbital bones. 4

External ocular tissues Eyebrows and Eyelids

The eyebrows separate upper eyelid from the forehead and may move as part of

changing facial expressions or in consult with change in the direction of the visual

axis and position of the eyelids. The eyebrows perform a variety of specialized

functions. They have a major influence on nonverbal communication by way of facial


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expression. In addition, owing to their position and curvature, the eyebrows help

shield the eyes from bright sunlight coming from directly.4

The eyelids cover the external part of the eye. These mobile folds protect the

eye from mechanical or chemical injury by sweeping the external surface of the eye at

periodic intervals and when closed as a first line of defense.4

The eyelids are lubricated and kept fluid-filled by secretion of the lacrimal

glands and specialized cells residing in the bulbar conjunctiva. The anterior chamber

has the shape of a narrow cleft directly over the front of the eyeball, with pocket-like

extensions upward and downward. The pockets are called the superior and inferior

fornices (vaults) and the entire space, the cul-de-sac. The elliptical opening between

the eyelids is called the palpebral fissure.3

ConjunctivaThe conjunctiva is a vascularized mucus membrane that covers the anterior

surface of the globe with the exception of the cornea. Conjuctival epithelium is

continuous with that of the cornea and with the epidermis of the lids and has a surface

that is about five times of the cornea. Mucus producing goblet cells which are

important for wetting and tear film stability are located in the conjuctiva. In addition

to physical protection of the globe, the conjuctiva has great potential for combating

infection for four reasons.4

It is highly vascular tissue. It contains many immune competent cells. The different types of cells are located within the conjuctiva. The anatomy and biochemistry of conjuctival cells.


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Anatomically the conjunctiva is divided into three areas.4

a. The palpebral (lid) conjuctivab.

The conjunctiva fornix

c. The bulbar (globe) conjuctiva Lacrimal system Tears

The conjuctival and corneal surfaces are covered and lubricated by a film of

fluid secreted by the conjuctival and lacrimal glands. The secretion of the lacrimal

glands; the tears are delivered through a number of fine ducts into the conjunctival

fornix. The secretion is a clear, watery fluid containing numerous salts, glucose, other

organic compounds, approximately 0.7 % proteins, enzymes and lysozyme.3

The cul-de-sac normally holds 7-9 l of tears but can retain up to approximately

20-30 l without overflowing if care is taken not to blink. Under baseline conditions,

the normal tear flow rate and tear film thickness are 1 l/min, i.e, approximately

16%/min and 4-9 m, respectively. Additionally, the normal pH of tears is 6.5-7.6.

Since tears are a well buffered system in part, because instilled solutions of lower pH

are quickly returned to physiological conditions.4

Secretary and Drainage apparatusThe lacrimal system consists of a secretary and collection portion. The secretary

portion of the lacrimal system consists of the main and accessory lacrimal glands.

Secreted tears do not normally flow across the cornea but will be assisted by the wiper

like action of the lids. It is assumed that secreted tears mix relatively rapidly and

thoroughly with tears held in the lower cul de sac.4

Tears are drained from the lacrimal lake through two small openings, the superior

and inferior canaliculi, which themselves join at the common canaliculus that leads


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into the upper part of the nasolacrimal duct, the beginning of which is the lacrimal

sac. The act of blinking exerts a suction force pump action in remaining tears from the

lacrimal lake and emptying them into the nasal cavity.

3

Anterior segment Anterior chamber

The anterior chamber is bounded in front by the cornea and a small portion of

the sclera. The anterior chamber is deepest centrally and shallowest at the periphery,

holding a volume of aqueous humor that has been shown to be 250 l in the human,

with a turnover rate of 1%/min. naturally, there is some overlap of tissues between the

anterior and posterior segments.4

CorneaThe cornea or window of the eye is an optically transparent tissue that conveys

images to the back of the eye. The cornea, being an avascular tissue, receives

nutrients and oxygen from the bathing solutions, i.e, the tears and aqueous humor as

well as from the blood vessels that line the junction between the cornea and sclera.

Corneal diameter is about 11.5 mm with a radius of curvature of anterior corneal

surface of 7.8 mm.4

LimbusThe corneosclerallimbus is a transitional zone 1-2 mm wide between the cornea

proper, sclera and conjuctiva. Externally the limbus is covered by peripheral corneal

epithelium and anterior conjuctival epithelium. Internally the limbus includes aqueous

veins, the canal of schlemm and the trabecular meshwork. The limbal blood supply is

an important source of nutrient and defense mechanism.4


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Trabecular meshwork and Schlemms canalThe trabecular meshwork and canal of schlemm form the conventional pathway

of aqueous humor outflow from the anterior segment of the eye. The trabecular

meshwork is divided into uveal and corneoscleral portions, each with there own

distinct anatomy. Aqueous humor leaves the eye at an angle in the anterior chamber,

where it enters the trabecular meshwork.4

Posterior chamberThe posterior chamber is somewhat triangular in appearance. The apex of the

triangle is located, where the edge of the iris rests on the lens. Ciliary processes form

the base, the posterior valve is formed by the lens and zonule and the anterior valve is

the posterior surface of the iris. The posterior chamber is filled with about 50 l of

aqueous humor. Aqueous humor is produced by ciliary processes, at the rate of 2.0 -

2.5l/ min, flows into the posterior chamber through the pupil and from there into the

anterior chamber.4

Iris and PupilThe pupil is circular opening located near the center of the iris. There are two

sets of smooth muscle in the iris that regulate papillary diameter: the sphincter

(parasympathetically innervated) and the dilator muscle (Sympathetically innervated).

The diameter of the normal pupil varies 2-9 mm. papillary dilation increases the

amount of light entering the eye and enhances the ability of the rods to function. In

contrast, papillary constriction increases depth of the focus of the eye and decreases

optical aberrations from the lens periphery.4

Ciliary bodyThe ciliary body is responsible for many functions in the eye. It secretes

aqueous humor that nourishes the lens, provides the muscle power for accommodation


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and may secrete the unique zonular fibers. The anterior is known as the pars plicata

and contains ciliary processes where aqueous humor is secreted. The posterior portion

of the ciliary body is known as the pars plana. Ciliary epithelium consists of two

layers an inner non-pigmented layer (NPE) and an outer pigmented layer (PE).3

Aqueous humorThe aqueous humor is produced both by active and passive secretion from the

ciliary processes. There are two enzymes located in the non-pigmented epithelium of

the ciliary processes that are intimately involved in the active secretion of aqueous

humor. These are the sodium-potassium activated adenosine triphosphate enzyme

(Na+-K+-ATPase) and carbonic anhydrase.4

Posterior segment Lens

The lens is a transparent biconvex structure located behind the iris and in front

of the vitreous. Like all lenses, that of the eye has two surfaces, anterior and posterior

and a border where these surfaces meet i.e, the equator, at its equator the lens has a

diameter of 10 mm. The average radius of curvature of the anterior surface is10 mm,

where as that of the posterior surface is 6 mm.4

ScleraThe outer coat of the eye is fibrous and serves a protective function. The white

opaque sclera constitutes the posterior five-sixths of the globe, where as the

transparent cornea comprises the anterior one-sixth of the globe. The anterior surface

of the sclera is covered by the episclera and the inner surface is covered by the

laminar fusca.4


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ChoroidThe choroid is the vascular coat of the eye and is divided into four layers.6

Supra choroids

Layer of vessels Chorio capillaries Bruchs membrane

VitreousThe vitreous humor comprises 80% of the internal volume of the eye. Vitreous

humor weighs 4g and occupies a volume of almost 4 ml. The vitreous cavity is

surrounded by the retina and optic nerve posteriorly. Anteriorly, it is bounded by the

ciliary body, zonules, and the posterior surface of the lens. In its normal state, the

vitreous is a clear gel composed almost entirely of water (99%). The vitreous is

impotent as a supporting structure and a metabolic pathway for nutrients for the lens

and retina. However of equal importance is its clarity and ability to transmit light to

the retina.4

RetinaThe retina is the inner most coat of the eye. It is a transparent tissue that lines

the posterior two-thirds of the eyeball. The only firm attachments of the retina are at

its anterior termination, the oraserrata, at the margins of the optic nerve.

4

Photoreceptor cells of the retina consist of rods and cones. The cones are concerned

with visual activity and color discrimination, whereas the rods are concerned with

peripheral vision under decreased illumination. The retina has the highest oxygen

consumption per unit weight of any tissue in the body.4


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Optic nerveThe optic nerve is a bundle of myelinated nerve fibers through which the entire

output of the retina travels. It reveals from the retina to the optic chiasm and is

divided into four portions, intra ocular portion, intra orbital portion, intra canalicular

portion, intra cranial portion.4

1.2 ABSORPTION AND BIOAVAILABILITY OF THE DRUGS FROM THE

EYE

The drug solution instilled as eye drops into the ocular cavity may disappear

from the precorneal area of the eye by any or a composite of the following routes. 5

shown in Fig.1.2.

1. Nasolacrimal drainage,

2. Tear turnover,

3. Productive corneal absorption,

4. Nonproductive Conjuctival uptake.

Fig.1.2 Routes of ocular absorption of drugs.


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Drug administered by instillation must penetrate the eyes and do so primarily

through the cornea. Corneal absorption is much more effective than scleral or

conjuctival absorption, in which removal by blood vessels into the general circulation

occurs.3

Many ophthalmic drugs are weak bases and are applied to the eye as aqueous

solution of their salts. The free base and the salts will be in an equilibrium that will

depend on the pH and the individual characteristics of the drug molecule. To aid in

maintaining storage stability and solubility, the medication may be acidic at the

moment of instillation but usually, the neutralizing action of the lacrimal fluid will

convert it rapidly to the physiological pH range (pH 7.4) at which there will be

enough free base present to begin penetration of the corneal epithelium. Once inside

the epithelium (lipid rich) the undissociated free base dissociates immediately to a

degree that the dissociated moiety then will tend to penetrate the stroma because it is

water-soluble. At the junction of the stroma (lipid poor) and endothelium (lipid rich),

the same process that took place at the outer surface of the epithelium must occur

again. Finally, the dissociated drug leaves the endothelium for the aqueous humor.

Here it can readily diffuse to the iris and the ciliary body, the site of its

pharmacological action.3

The topical application of ophthalmically active drugs to the eye is the most

prescribed route of administration for the treatment of various ocular diseases. It is

generally agreed that the intraocular bioavailability of topically applied drugs is

extremely poor.5 Upon instillation of an ophthalmic solution; most of the instilled

volume is eliminated from the pre corneal area. This loss is mainly due to drainage of

the excess fluid by the nasolacrimal duct or elimination of the solution by tear

turnover, which will results in poor ocular bioavailability.


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1.3 GENERAL BACTERIAL INFECTIONS OF EYE AND ITS

MEDICATIONS

Common bacterial infections of eye are listed as below:

7

ConjuctivitisIt is a common superficial eye disorder and may be caused by infection with a

wide range of bacteria, viruses and less frequently fungi. Staphylococci or

Streptococci commonly cause acute bacterial conjunctivitis in adults, and

Haemophilus influenzae and Morexella catarhallis particularly in children. Other

causes of bacterial conjuctivitis include Gonococci and Chlamydia trachomatis.

Uncomplicated bacterial conjunctivitis may be self limiting but empirical treatment

with topical antibacterial is often given.

BlepharitisIt is an infection of the lid margins. It is usually present in a chronic condition

and may require prolonged treatment, typically involving local hygiene to remove

encrustations and topical application of a broad-spectrum antibacterial ointment.

KeratitisIt may be caused by infection of the cornea by bacteria, fungi, viruses or

protozoa usually following trauma to the surface of the eye, including that due to

contact lens wear. Common bacterial pathogens include Staphylococci, Streptococci,

Pseudomonas spp. and Enterobacteriaceae. Bacterial keratitis is potentially sight

threatening and requires prompt aggressive treatment with broad-spectrum

antibacterials. Frequent or continuous topical application of drops or the use of local

drug delivery devices has been used to ensure prolonged elevated drug concentrations.


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EndophthalmitisIt is a devastating ocular disease resulting from infection of the ocular cavity,

usually following penetrating trauma or surgery. Depending on the route of infection,

causative organisms commonly include Staphylococci, Streptococci, Haemophilus

influenzae, bacillus cereus and Propionibacterium acnes. Bacterial endophthalmitis

requires immediate aggressive treatment with antibacterials, usually given

intravitreally.

A considerable amount of effort has been made in ophthalmic drug delivery

since the 1970. A number of approaches to the delivery of drugs for ocular treatment

has been investigated and proposed. These range from simple systems such as

aqueous suspensions where the viscosity and hence the residence time, has been

increased by cellulosic polymer to complex system such as penetration enhancers,

external devices (collagen shields, Preformed gels, iontophoresis and pumps),

ion-exchange resins, liposomes, microspheres and micro particles, polymeric films,

inserts, prodrugs, mucoadhesives and metabolism based drug design.8

The commercially available antibacterial agents for ophthalmic use are as follows:


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Table 1.1 Topical antibacterial agents for ophthalmic use.

Generic name Formulation Indications

Bacitracin Zinc 500 /g ointment Conjunctivitis Blepharitis

Chloramphenicol 0.5% solution 1% ointment Conjunctivitis Keratitis

Chlortetracyclin 1% ointment Conjunctivitis Blepharitis

Pefloxacin 0.3% solution Conjunctivitis Keratitis

Erythromycin 0.5% ointment Conjunctivitis Blepharitis

Gentamicin sulfate 0.3% solution 0.3% ointmentConjunctivitis Keratitis

Blepharitis

Norfloxacin 0.3% solution Conjunctivitis

Sulfactamide sodium10, 15 at solution

10% ointment

Conjunctivitis Keratitis

Blepharitis

Sulfisovarzole 4% solution 4% ointmentConjunctivitis Blepharitis

Lentitis

Polymixin BVarious solution and various

ointmentConjunctivitis Blepharitis

Tetracycline 1% solution Conjunctivitis Blepharitis

Tobramycin sulfate 0.3 % solution 0.3% ointmentConjunctivitis Keratitis,

Blepharitis

1.4 OCULAR DRUG DELIVERY SYSTEMS9

Solutions and suspensionsSolutions are the pharmaceutical dosage forms most widely used to administer

drugs that must be active on the eye surface or in the eye after passage through the

cornea or the conjunctiva. Solutions also have disadvantages:

The solution stays for a short time on eye surface,

It has poor bioavailability,


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The instability of the dissolved drug and the necessity of usingpreservatives.

Sprays

Although not commonly used, some practitioners use Mydriatics or

Cycloplegics alone or in combination in the form of eye spray. These sprays are used

in the eye for dilating the pupil or for Cycloplegics examination.

Contact lensesContact lenses can absorb water soluble drugs when soaked in drug solutions.

These drug saturated contact lenses are placed in the eye for releasing the drug for a

long period of time. The hydrophilic contact lenses can be used to prolong the ocular

residence time of the drugs.

Artificial tear insertsA rod shaped pellet of Hydroxy propyl cellulose, without preservative is

commercially available (Lacrisert). This device is designed as a sustained release

artificial tear for the treatment of Dry eye disorder.

Filter paper stripsSodium fluorescein and bengal dyes are commercially available as drug-

impregnated filter paper strips. These dyes are used diagnostically to disclose corneal

injuries and infections such as herpes simplex and dry eye disorders.

Micro emulsionDue to their intrinsic properties and specific structures, microemulsions are a

promising dosage form for the natural defense of the eye. Indeed, because they are

prepared by inexpensive processes through auto emulsification or supply of energy

and can be easily sterilized, they are stable and have a high capacity of dissolving the

drugs.


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Ocular InsertsOcular inserts, one of the new classes of drug delivery systems, which are

gaining worldwide praise, release drugs at a pre-programmed rate for a longer period

by increasing the precorneal residence time.10 The goal of this delivery system is to

provide a therapeutic amount of drug to the ocular tissues to achieve promptly and

then maintain the desired drug concentration by increasing the contact time between

the preparation and the Conjunctival tissue.11 To achieve this goal particularly for

chronic diseases such as glaucoma, it would be advantageous and more convenient to

maintain a dosing frequency to once, or at most, twice a week regimen.12An

appropriately designed extended release ocular insert can be a major advance in this

direction compared to conventional immediate release dosage forms.

Collagen ShieldCollagen is regarded as one of the most useful biomaterials. The excellent

biocompatibility and safety is due to its biological characteristics, such as

biodegradability and weak antigenicity, these properties made collagen the primary

resource in medical applications. Collasomes show promise among drug delivery

systems to the human eye. They are first fabricated from procine scleral tissue, which

bears a collagen composition similar to that of the human cornea. The shields are

hydrated before they are placed on the eye. Shields are not individually fit for each

patient, as are soft contact lenses and therefore, comfort may be problematic and

expulsion of the shield may occur.

Ocular IontophoresisIontophoresis is the process in which direct current drives ions into cells or

tissues. When iontophoresis is used for drug delivery, the ions of importance are

charged molecule of the drug. Ocular iontophoresis offers a drug delivery system that


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is fast, painless and safe; and in most cases; it results in the delivery of a high

concentration of the drug to a specific site. But the role of iontophoresis in clinical

ophthalmology remains to be identified.

LiposomesLiposomes are phospholipid lipid vesicles for targeting drugs to the specific

sites in the body; provide the controlled and selective drug delivery and improved

bioavailability. Liposomes offer the advantages of being completely biodegradable

and relatively non toxic but are less stable than particulate polymeric drug delivery

systems.

NiosomesIn order to circumvent the limitations of liposomes, such as chemical instability,

oxidative degradation of phospholipids, cost and purity of natural phospholipids,

niosomes have been developed as they are chemically stable compared to liposomes

and can entrap both hydrophilic and hydrophobic drugs. They are non toxic and do

not require special handling techniques.

Mucoadhesive Dosage FormsThe successful development of fewer mucoadhesive dosage forms for ocular

delivery still poses numerable challenges. This approach relies on vehicles containing

polymers, which will attach via noncovalent bonds to conjuctival mucin.

Nanoparticles and MicroparticlesParticulate polymeric drug delivery systems include micro and nanoparticles.

The upper size limit for microparticles for ocular delivery is about 5-10 mm, above

this size; a scratching feeling in the eye can result after ocular application. After

optimal drug binding to microspheres or nanoparticles, the drug absorption in the eye

enhanced significantly in comparison to eye drops.


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HydrogelsThe most common way to improve drug retention on the corneal surface is

undoubtedly by using polymers to increase solution viscosity.

13

Hydrogels are

polymers endowed with anability to swell in water or aqueous solvents and induce a

liquidgel transition. 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. In situ

forming gels are formulations applied as solutions, sols or suspensions that undergo

gelation after instillation due to physicochemical changes inherent to the eye .14

Fig.1.3 Classification of hydrogels.


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In situ gelDistinguishing from preformed hydrogels, in situ forming gels are formulations,

applied as a solution, which undergoes gelation after instillation due to

physicochemical changes inherent to the biological fluids. In this way, the polymers

which show sol-gel phase transition and thus trigger drug release in response to

external stimuli are the most investigated. In situ hydrogels are providing such

sensor properties and can undergo reversible sol-gel phase transitions upon changes

in the environmental condition. These intelligent or smart polymers play

important role in drug delivery since they may dictate not only where a drug is

delivered, but also when and with which interval it is released.15

A polymer used to prepare in situ gels should have following charteristics: 16

It should be biocompatible. It should be capable of adherence to mucus. It should have pseudo plastic behaviour. It should have good tolerance and optical clarity. It should influence the tear behaviour. The polymer should be capable of decreasing the viscosity with increasing

shear rate there by offering lowered viscosity during blinking and stability of

the tear film during fixation.

1.5METHODS FOR PREPARATION OF IN SITU GELSThere are many methods have been employed to cause reversible sol-gel phase

transition in cul de sac and some of these methods are change in temperature18, pH19

and electrolyte composition.20


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Thermo reversible in situ gelsThese hydrogels are liquid at room temperature (20250C) and undergo gelation

when in contact with body fluids (3537

0

C), due to an increase in temperature.

21

Example: Poloxamers, cellulose derivatives and xyloglucan. The Poloxamers consist

of more than 30 different non ionic surface active agents.

These polymers are ABA-type tri block copolymers (Fig.1.4) composed of

polyethylene oxide (PEO) (A) and polypropylene oxide (PPO) units (B). The

Poloxamer series covers a range of liquids, pastes and solids with molecular weights

and ethylene oxidepropylene oxide weight ratios varying from 1100 to 14,000 and

1:9to8:2.22

Fig.1.4PEO-PPO-PEO (Poloxamer).

The gelation mechanism of Poloxamer (Fig.1.5) solutions has been

investigated extensively, but is still being debated. Ultrasonic velocity, light-scattering

and small-angle neutron scattering measurements of aqueous Poloxamer solutions

have clearly indicated a micellar mode of association. Micelle formation occurs at the

critical micellization temperature as a result of PPO block dehydration with increasing

temperature, micellization becomes more important and at a definite point, micelles

come into contact and no longer move. In addition, the formation of highly ordered

structures, such as cubic crystalline phase, has been proposed as the driving force for

gel formation, but this hypothesis has been questioned recently.22Thus, packing of

micelles and micelle entanglements may be possible mechanisms of Poloxamer

solution gelation with increase of temperature.23 Furthermore, it has suggested that

intramolecular hydrogen bonds might promote gelation.The Poloxamers are reported


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to be well tolerated and non-toxic even though large amounts (25-30%) of polymers

are required to obtained a suitable gel.

Fig.1.5 In situ gel formation due to change in temperature.

Thermo reversible gels can be prepared with naturally occurring polymers. Most

natural polymer aqueous solutions form a gel phase when their temperature is

lowered. Classic examples of natural polymers exhibiting a solgel transition include

gelatin and carrageenan. At elevated temperatures these polymers adopt a random coil

conformation in solution. Upon cooling, a continuous network is formed by partial

helix formation.24

Cellulose derivatives also cause gelation Eg: Methylcellulose and hydroxy

propyl methyl cellulose (HPMC) are typical examples of such polymers.

Xyloglucan a polysaccharide derived from tamarind seed, forms thermo

responsive gels in water, under certain conditions. Gelation is only possible when the

galactose removal ratio exceeds 35%.25 The transition temperature is inversely related

to polymer concentration26 and the galactose removal ratio. For Eg: The solgel

transition of xyloglucan was shown to decrease from 40 to 50C when the galactose

removal ratio increased from 35 to 58%. Xyloglucan is approved for use as a food

additive. However, its relatively low transition temperature (22270C) makes

handling at room temperature problematic.27


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pH sensitive in situ gelsGelling of the solution is triggered by a change in the pH.Eg: Cellulose acetate

phthalate (CAP) latex and its derivatives such as carbomer (Fig.1.6) are used.

Cellulose acetate derivatives are the only polymer known to have a buffer capacity

that is low enough to gel effectively in the cul-de-sac of the eye.28 First preliminary

investigations of pH sensitive latexes for ophthalmic administration began in early

1980 and have been extensively studied by Boye.29

Fig.1.6 Structure of Carbomer.

Cellulose acetate phthalate latex is a polymer with potentially useful properties

for sustained drug delivery to the eye because latex is a free running solution at a pH

of 4.4, which undergoes coagulation when the pH is raised by the tear fluid to pH 7.4.

But the low pH of the preparation can elicit discomfort in some patients.30The poly

acrylic acid and its lightly cross-linked commercial forms (Polycarbophil and

Carbopol) exhibit the strongest mucoadhesion,31 Carbomer (Carbopol) a cross-linked

acrylic acid polymer (PAA) also shows pH induced phase transition as the pH is

raised above its pKa of about 5.5.32Different grades of Carbopol are available. The

manufacturer states that Carbopol 934 gel has the lowest cross-linking density, while

Carbopol 981 intermediate and Carbopol 940 have the highest, higher the cross

linking ability more stiff is the gel formed.

All pH-sensitive polymers contain pendant acidic or basic groups that either

accept or release protons in response to changes in environmental pH. The polymers


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with a large number of ionizable groups are known as polyelectrolytes. Swelling of

hydrogel increases as the external pH, increases in the case of weakly acidic (anionic)

groups, but decreases if polymer contains weakly basic (cationic) groups (Fig.1.7).

Some of examples of that are delivered by pH sensitive method includes

Ciprofloxacin,1 Indometacin,35 Gatifloxacin36etc.

Fig.1. 7 Schematic representation of pH dependent in situ gels.

Ion sensitive in situ gelsPolymers may undergo phase transition in presence of various ions. Some of the

polysaccharides fall into the class of ion-sensitive ones.38

Gellan gum (Gelrite) is a linear, (Fig.1.8) anionic hetero polysaccharide

secreted by the microbe Sphingomonas elodea (formerly known as Pseudomonas

elodea). The polysaccharide can be produced by aerobic fermentation and then

isolated from the fermentation broth by alcohol precipitation. The polymer backbone

consists of glucose, glucuronic acid and rhamnose in the molar ratio 2:1:1.39These are

linked together to give a tetra saccharide repeat unit .The native polysaccharide is

partially esterified with L-glycerate and acetate,40but the commercial product Gelrite


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has been completely de-esterified by alkali treatment.41Formulations with the Gelrite

can be administered to ocular mucosa as a low viscosity solution. On contact with

cations in tear fluid the formulation will form a clear gel.

42

This is caused by cross

linking of the negatively charged polysaccharide helices by monovalent and divalent

cations (Na+, K+, Ca+). Several models have been presented to explain gellan gum

gelation.

Fig1.8 Structure of deacetylated gellan gum.

Mechanism involved in sol to gel transistion by gelrite is as follows, in an ion

free aqueous medium, Gelrite forms double helices at room temperature. This solution

has a viscosity close to that of water and the helices are only weakly associated with

each other (by van der waals attraction). When gel-promoting cations are present,

some of the helices associate into cation-mediated aggregates, which cross-link the

polymer. On heating the polysaccharide in an ion free environment, the

polysaccharide becomes a disordered coil. However, on heating the sample with

cations present, the non-aggregated helices melt out first and the aggregated helices

melt out at a higher temperature in a second transition. The divalent ions such as

magnesium or calcium were superior to monovalent cations in promoting the gelation


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of the polysaccharide.43 However the concentration of sodium in tears (2.6 g/l) is

quite sufficient to induce the gelation. Corneal contact time of formulations based on

gellan gum has been investigated using two main methods, which are fluorometry

44

and -scintigraphy.45 Both techniques have demonstrated improved residence times

with Gelrite when compared with saline or various commercial solutions. Gelrite has

also provided corneal residence times superior to those of other hydrogel preparations

based on polymers such as cellulosic derivatives or xanthan gum.

Alginates being a family of unbranched binary copolymers, alginates consist

of (14) linked -D-mannuronic acid (M) and -L-guluronic acid (G) residues of

widely varying composition and sequence. By partial acid hydrolysis, alginate was

separated into three fractions.46 Two of these contained almost homopolymeric

molecules of G and M respectively, while a third fraction consisted of nearly equal

proportions of both monomers and was shown to contain a large number of MG dimer

residues. It was concluded that alginate could be regarded as a true block copolymer

composed of homo polymeric regions of M and G, termed M and G-blocks,

respectively, inter spaced with regions of alternating structure. It was further shown

that alginates have no regular repeating unit and that the distribution of the monomers

along the polymer chain could not be described by bernoullian statistics. Knowledge

of the monomeric composition is hence not sufficient to determine the sequential

structure of alginates.47, 48 Alginate with a high guluronic acid content will improve

the gelling properties and reduce the total polymer to be introduced into the eye.

The aim of this study is to prepare in situ ophthalmic gel of anti-infective drug

Levofloxacin hemihydrate to enhance ocular bioavailability and reduce dose

frequency and there by increasing patient compliance.


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CHAPTER 2 AIMS AND OBJECTIVES

Department of pharmaceutics Bharathi college of pharmacy 26

Most commonly available ophthalmic preparation are eye drops and ointments.

But these preparations when instilled in to the cul-de sac are rapidly drained away

from the ocular cavity due to the tear flow and naso-lachrymal drainage. Only small

amount is available for its therapeutic effect resulting in frequent dosing. When a drug

solution as dropped in to the eye, effective tear drainage and blinking results in

10-fold reduction of drug concentration in 4-20 minutes.

Ocular therapy could be significantly improved if the pre-corneal residence time

of drugs could be increased, several new preparations have been developed for

ophthalmic use not only prolong the contact time of the vehicle at ocular surface, but

also to slow down the elimination of the drugs. This problem can be overcome by

using in situ gel forming ophthalmic drug delivery systems prepared from polymers

that exhibit reversible phase transition and pseudoplastic behavior to minimize

interference with blinking. Such system can be formulated as liquid dosage form

suitable for administration by instillation in to the eye, which upon exposure to the

eye shift to the gel phase; gelation depends upon physiological pH, temperature and

ionic strength.36, 49

The aim of present work is to prepare and evaluate in situ opthalmic gel of an

anti infective drug for sustained ocular delivery which is used for the treatment of

various infective diseases of the eye, to get better patient compliance by increasing

residence time and bioavailability.

2. AIMS AND OBJECTIVES


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The broad objective of the present work is

To develop an in situ ophthalmic gel of an anti infective drug- Levofloxacinhemihydrate for sustained ocular delivery for the treatment of bacterial infections

of the eye.

To evaluate the prepared formulation for its In vitrogelation and rheological studies, Drug content uniformity, In vitrorelease studies, Sterility testing of the optimized in situ gel, Stability studies, Pharmacodynamic studies and Pharmacokinetic release studies.

To provide a formulation with better residence time enhanced bioavailability aftertopical administration and improved patient compliance.

2.1 PLAN OF WORK

1. Literature survey.

2.Experimental work.

Preformulation studies. Interaction studies of polymers and drug before selection of the formulation by

FTIR.

Formulation of in situ gel by incorporating optimum polymer, excipents anddrug.

Optimization of ion exchange triggered in situ gelling system using differentpolymeric concentrations of gelrite.


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Preliminary evaluation of prepared in situ gel formulations by visualappearance, clarity, pH, drug content, and in vitrogelation.

Estimation of drug by spectrophotometric method. Evaluation studies of prepared in situ gel formulation for rheological

properties, in vitrorelease of the drug, antibacterial studies, sterility, stability

studies and pharmacodynamic studies (irritation study).

Comparative evaluation of in vitrodrug release with marketed product.


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CHAPTER 3 REVIEW OF LITERATURE


Eaga Chandra Mohan et al1.,prepared in situ gels based on pH-triggered in situ

gelation, thermo reversible gelation and ion activated system. Poly acrylic acid

(Carbopol 940) was used as the gelling agent in combination of Hydroxy Propyl

methylcellulose, which acted as a viscosity-enhancing agent. (pH-triggered system).

Pluronic F-127 (14%) was used as the thermal reversible gelation in combination of

HPMC (1.5%) incorporation of HPMC was to reduce the concentration of pluronic

required for in situ gelling property, with 25% w/w pluronic F-127 reported to form

good gels. Gellan gum (Gelrite) is an anionic exocellular polysaccharide produced by

the bacterium pseudo monas elodea, having the characteristic property cation-induced

gelation (0.6%). The developed formulation was therapeutically efficacious, stable,

non irritant and provided sustained release of the drug over a 6 hours period, but

Gelrite formulation showing long duration of release followed by combination of

carbopol, HPMC and pluronic F-127 & HPMC. The developed system is thus a viable

alternative to conventional eye drops.

Chrystele Le Bourlais et al6., given recent advances in ophthalmic drug delivery

systems. Eye drops are the conventional dosage forms that accounts for 90% currently

accessible ophthalmic formulations. Despite the excellent acceptance by patients, one

of the major problems encountered is rapid precorneal drug loss. To improve ocular

drug bioavailability, in situ activated gel forming systems are preferred as they can be

delivered as drops, with sustained release properties.

3. REVIEW OF LITERATURE


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Miyazaki S et al26

.,prepared in situ gelling Xyloglucan formulations for sustained

release ocular delivery of Pilocarpine HCL. They found that the degree of

enhancement of mitotic response following sustained release of Pilocarpine from 1.5

% w/w Xyloglucan gel was similar to that from a 25 % w/w Pluronic F127 gel.

Srividya B et al33., prepared sustained ophthalmic delivery of Ofloxacin from a pH

triggered in situ gelling system using Polyacrylic acid (Carbopol 940) as the gelling

agent in combination with HPMC E50LV. They observed that the developed

formulation was therapeutically efficacious, stable, non-irritating and provided

sustained release of the drug over an 8 h period.

Kumar S et al34., prepared a in situ forming gels by a combination of carbopol and

methylcellulose, they found that solution containing 1.5 % Methylcellulose, 0.3 %

carbopol have low viscosity and form a strong gel under simulated physiological

conditions. increase in concentration of either carbopol or methylcellulose results in

an increased in viscosity, shear stress among the compositions. They also studied the

rheological characterization of such a system at two different pH (4 and 7) and

temperatures (25 and 37C).

Thilek kumar M et al35., prepared pH induced in situ gelling system of

Indomethacin for sustained ocular delivery by using carbopol as the gelling agent in

combination with HPMC K15M. They observed that the carbopol solutions which are

acidic and less viscous, transform into stiff gels upon increase in pH by tear fluid of

the eye and produced sustained release of Indomethacin over 8 hour periods, which

made them an excellent candidate for in situ gelling ocular delivery system.

Doijad RC et al36., studied on preparation of in situ ophthalmic gels of Gatifloxacin

for the treatment of bacterial conjunctivitis, using sodium alginate as the gelling agent

with HPMC which acted as viscosity enhancing agent. The developed formulations


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were therapeutically efficacious, stable, and non-irritant and provided sustained

release of drug over an eight hour period.

Rozieret al

41., prepared novel ion activated in situ gels using Gelrite as a polymer

for ophthalmic vehicles and a 0.6 % Gelrite vehicle has been compared to an

equiviscous solution of hydroxy ethyl cellulose using Timolol Maleate as a drug

probe. They were observed that the formation of the gel prolonged precorneal

residence time and increased ocular bioavailability of Timolol.

Katarina Lindell et al49

., showed in vitrorelease of Timolol maleate from an in situ

gelling polymer, cellulose ether [ethyl (hydroxy ethyl) cellulose] system. They

observed that the release of Timolol maleate was about equal for system with 1-2 %

w/w EHEC, implying that the release was controlled by a low concentration in the

gels and not by any drug-polymer interaction.

Kumar S et al50., studied modification of in situ gelling behaviour of carbopol

solutions by HPMC. They were found that in combination both HPMC and carbopol

form low viscosity liquid at pH 4 and transform into stiff gels with plastic rheological

behaviour and comparable viscosities upon increasing the pH up to 7.4 and HPMC-

PAA gels show slow in vitrorelease of incorporated Timolol Maleate.

Smadar Cohen et al51.,developed a novel in situ forming ophthalmic drug delivery

system from alginate undergoing gelation in the eye. They demonstrated that an

aqueous solution of sodium alginate could gel in the eye, without the addition of

external calcium ions or other bivalent/polyvalent cations. Alginate with guluronic

acid contents of more than 65 %, such as Manugel DMB, instantaneously formed gels

upon their addition to simulated lachrymal fluid, while those having low guluronic

acid contents, such as Ketton LV, formed weak gels at a relatively slow rate and

hence it was indicated that the in situ gelling alginate system, based on polymers with


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high guluronic acid contents was an excellent drug carrier for the prolonged delivery

of Pilocarpine.

Shulin Dinget al

52., given recent development in ophthalmic drug delivery. Recent

research efforts in ophthalmic drug delivery have focused on systems in which drug

may be administered in the form of eye drops. As a result of these efforts, significant

advancements have made in the in situ forming gels.

Dimitrova et al53

., developed a model aqueous ophthalmic solution of Indomethacin

using Pluronic F68 and Pluronic F127. They showed that both Pluronics acted very

similarly and were more effective as solublizers, created an appropriate viscosity and

formed reversible gels at higher temperature, ensured the Indomethacin chemical

stability and prolonged in vitro drug diffusion, and showed high physiological

tolerance on rabbit eyes.

Odile Sechoy et al54

., developed a new long acting ophthalmic formulation of

Carteolol containing alginic acid. They observed that the alginic acid vehicle is an

excellent drug carrier, well tolerated and could be used for the development of a long

acting ophthalmic formulation of Carteolol. In vitro studies indicated that Carteolol

was released slowly from alginic acid formulation, suggesting an ionic interaction.

El-Kamel AH et al 55., demonstrated in vitro/in vivo evaluation of Pluronic F127

based ocular delivery system for Timolol Maleate. He observed that the slowest drug

release was obtained from 15 % Pluronic F127 formulation containing 3 % methyl

cellulose. In vivo study showed that the ocular bioavailability of Timolol maleate

increased by 2.5 and 2.4 fold for 25 % Pluronic F127 gel formulation and 15 %

Pluronic F127 containing 3 % Methyl cellulose respectively, compared with 0.5 %

Timolol Maleate aqueous solution.


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Naseem A Charoo et al 56., prepared in situ forming ophthalmic gels ofCiprofloxacin HCL for the treatment of bacterial conjuctivitis, using HPMC K15 M

and Carbopol 934, they concluded that the sol-to-gel system exhibited a zero-order

drug release pattern over 24 hr .

Sultana Y et al57

., evaluated Carbopol-methyl cellulose based sustained release

ocular delivery system for Pefloxacin mesylate using rabbit eye model. It was found

that the optimum concentration of carbopol solution for in situ gel forming delivery

system was 0.3 % w/w and that for methyl cellulose solution was 1.5 % w/w. The

mixture of solutions showed a significant enhancement in gel strength in the

physiological condition. They observed that both in vitroand in vivostudies and these

studies indicated that the carbopol and methyl cellulose solution alone and mixture

can be used as an in situ gelling vehicle to enhance the ocular bioavailability of

Pefloxacin mesylate.

Kulkarni M.C et al58

., prepared the ophthalmic in situ gelling formulation of

Flubiprofen sodium using Gellan gum. The formulation in gel form showed almost

complete release of drug. The formulation when subjected for accelerated stability

studies showed good physical and chemical stability and the in vivo studies of the

formulation in albino rabbits confirmed its in situ gelling capacity, non irritancy to

eyes as well as its non toxic nature.

Kugalur Ganesan Parthiban et al59.,prepared pH dependent in situ ophthalmic

gels of Ketorolac tromethamine using polyacrylic acid (carbopol 940) which is used

as a gelling agent in combination with hydroxy propyl methyl cellulose (HPMC-

K15M, K4M) as a viscosity enhancer. Benzalkonium chloride at suitable

concentration was used as a preservative.The prepared formulations were evaluated

for clarity, pH measurement, gelling capacity, drug content, and in vitro diffusion


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study. Under rheological investigation both solution and gel was found to be having

pseudo plastic behaviour. The selected formulations showed sustained release over a

period of 8hrs with increased residence times. Eye irritation test using the Draize test

protocol with cross over studies were preformed on selected formulations.

Shivanand swamy PH et al60., studied on formulation and evaluation of novel in

situ gum based ophthalmic drug delivery of Linezolid using various gum based

polymers. The results showed sustained release of Linezolid up to 6 hours.

Kaur IP et al61 .,studied on in situ ophthalmic preparation of Acetazolamide using

polymers such as poly vinyl alcohol, HPMC and Ethylene diamine tetra acetic acid as

a penetration enhancer to increase the absorption of the drug. Formulations were

evaluated for their in vitro release pattern. The effect of these formulations on the

intra ocular pressure in rabbits was investigated. These formulations were found to

maximize the therapeutic effects with decreased frequency of administration.

Divyesh HS et al 62., studied on mucoadhesive ophthalmic in situ hydrogel of

Moxifloxacin Hcl using combination of poloxamer 407 and poloxamer 188 with

mucoadhesive polymers like Gelrite, xantum gum and sodium alginate with a view

for enhancing bioavailability of the drug and developed formulations showed

sustained release of 10 hours.

Sindhu Abraham et al63., studied on in situ ion activated gelling system of

Ofloxacin using sodium alginate as gelling agent and Hydroxyl propyl cellulose

(HPC) as viscosity enhancing agent, in vitrorelease showed combination of HPC and

alginate solution better retained the drug than HPC and alginate alone, the prepared

formulations followed first order diffusion controlled release kinetics.


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Sirish Vodithala et al64., prepared in situ ophthalmic gel of Ketorolac tromethamine

using gelrite as a polymer. The formulations were evaluated for clarity, pH, gelling

capacity, drug content, rheological study,in vitro

drug release, ocular irritancy studies

(as per Draize test) and in vivocorneal permeation studies using isolated goats cornea.

The developed formulations showed sustained release of drug up to 6 hrs. The

formulations were found to be nonirritating with no ocular damage.

Johan Carlfors et al65.,studied the rheology of in situ gels prepared by using gelrite

as a polymer, he also performed a complementary in vivo study for determining

precorneal contact times in humans and in rabbits. He concluded that the elastic

moduli of the gels increased with increasing concentration of electrolytes. At

physiological concentration of the electrolytes, the elasticity of the gels was

independent of Gelrite concentration. The human contact times increased up to 20

hours with decreasing osmolality of the formulations. The results indicate that a high

rate of the sol / gel transition results in long contact times.

Coquelet C et al66., evaluated association between benzalkonium chloride and a poly

acrylic acid in gels by microfiltration and membrane dialysis. The interaction between

benzalkonium chloride and the respective polymers, the related availability

benzalkonium chloride in the corresponding solutions, were studied for aqueous

preparation of hydroxy ethyl cellulose, polyvinyl alcohol and cross linked poly acrylic

acid. The study was performed by means of a cross flow filtration process with an

alumina membrane. In the presence of the poly acrylic acid gel, the rejection rate of

benzalkonium chloride is much higher than with hydroxy ethyl cellulose and

polyvinyl alcohol. These results can be explained by the association of the

benzalkonium cation with the negative carbohydrate group of the poly acrylic acid.

This effect, which was confirmed by dialysis experiment, leads to trapping of


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benzalkonium cation inside the polymer network and could be of interest in the

reduction of the harmful effects of benzalkonium chloride observed in the treatment

of eye diseases.

Furrer et al67., given application of in vivo confocal microscopy to the objective

evaluation of ocular irritation induced by surfactants. An ocular irritation test using

confocal laser scanning ophthalmoscopy has been developed in which corneal lesions

subsequent to instillation of surfactants are specifically marked by fluorescein and

assessed by digital image processing. Benzalkonium chloride, a cationic surfactant at

a concentration range of 0.01 to 0.5% was tested. The cornea was evaluated for

in vivo ocular tolerance by confocal microscopy. In both rabbits and mice, the test

revealed following irritation ranking: cationic > anionic > nonionic surfactants. In

both animal models, the ocular damage increased with the concentration of

benzalkonium. The test was sensitive enough to detect ocular micro lesions at

concentration of surfactants as low as 0.01% for benzalkonium.


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3.1.1LEVOFLOXACIN HEMIHYDRATE

68

Levofloxacin hemihydrate is member of the fluoroquinolone class of

antimicrobial drugs. It is active against a wide range of Gram +ve and Gram -ve

organisms.

Structural Formula

Molecular Formula :C18H20FN3O

Generic name :Levofloxacin.

Nomenclature :(-)-(S)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methyl-

1piperazinyl)7-oxo7Hpyrido [1, 2, 3-de]-1,

4benzoxazine-6-carboxylicacid hemihydrate.

Molecular Weight : 361.3675 Daltons.

Melting Range : 226C.

log p value : 2.1.

Half life : 6 to 8 hours.

3.1 DRUG PROFILE


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Description

A synthetic fluoroquinolone (fluoroquinolones) antibacterial agent that inhibits

the super coiling activity of bacterial DNA gyrase, halting DNA replication.

Appearance

Levofloxacin hemihydrate is pale yellow coloured amorphous solid.

Solubility

Levofloxacin hemihydrate is in soluble in water but its predicted water

solubility is 1.44e+00 mg/ml, it is soluble in chloroform and other organic solvents,

solubility of Levofloxacin hemihydrate can be increased by usingco solvents such as

glycerine and propylene glycol.

It is considered to be freely soluble in this pH range, as defined by USP

nomenclature. Above pH 5.8, the solubility increases rapidly to its maximum at pH

6.7 (272 mg/ml) and is considered freely soluble in this range. Above pH 6.7, the

solubility decreases and reaches a minimum value (about 50 mg/mL) at a pH of

approximately 6.9.

Bio pharmaceutics

Mechanism of Action as Antibacterial Agent

Levofloxacin inhibits bacterial type II topoisomerases, topoisomerase IV and

DNA gyrase. Levofloxacin, like other fluoroquinolones, inhibits the A subunits of

DNA gyrase, two subunits encoded by the gyrA gene. This results in strand breakage

on a bacterial chromosome, supercoiling and resealing; DNA replication and

transcription are inhibited.

Pharmacokinetics

Absorption of Levofloxacin after single or multiple doses of 200 to 400 mg is

predictable and the amount of drug absorbed increases proportionately with the dose.


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The peak and trough plasma concentrations attained following multiple once daily

oral 500 mg regimens were approximately 5.7 g/ml and 0.5 g/ml respectively.

There is no clinically significant effect of food on the extent of absorption of

Levofloxacin.

Adverse Effects

The most frequently reported drug-related adverse reaction was transient.

Ocular burning or discomfort. Other reported reactions include stinging, redness,

itching, chemical conjunctivitis / keratitis, blurred vision, dryness, and eye pain.

Precautions and Warnings

If an allergic reaction to Levofloxacin occurs, discontinue the drug. Serious

acute hypersensitivity reactions may require immediate emergency treatment, oxygen

and airway management, including incubation should be administered as clinically

indicated. As with other anti-infective, prolonged use may result in overgrowth of non

susceptible organisms, including fungi. If super infections occur discontinue use and

institute alternative therapy. Levofloxacin should be discontinued at the first

appearance of a skin rash or any other sign of hypersensitivity reaction.

Usage and Administrations

Topically

Levofloxacin as 0.5 % eye drops is used for treatment of Bacterial infections.

Orally

Levofloxacin is also used for the treatment of susceptible infections of skin,

lungs, ears, airways, bones, and joints caused by susceptible bacteria. Levofloxacin

also is frequently used to treaturinary infections, including those resistant to other

antibiotics, as well as prostatitis. Levofloxacin is effective in treating

infectiousdiarrhoea caused by E.coli, Shigella and Campylobacter jejuni.
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Levofloxacin also can be used to treat various obstetric infections, including mastitis

(infection of the breast).

3.2 PROFILE OF EXCIPENTS.

3.2.1 GELRITE69

Non-proprietary Names :Gellan gum.

Functional Category : Gelling agent alternative to agar.

Synonyms : gellan gum.

Chemical composition :Polysaccharide comprising glucuronic acid,Rhamnose and glucose.

Physical state : Dry powder.

Description : Gelrite is aWhite to tan coloured solid.

pH : The pH of 1 % w/v aqueous dispersion is 7.

Solubility : Soluble in water forming viscous solution

becoming a paste at concentration >5% gels

if heated and cooled.

Materials to avoid : Strongacids, strong bases.

Decomposition products : May include CO2and CO.

Hazardous polymerization:None.

Stability and Reactivity

Storage and handling procedures should follow the normal practices

recognized as desirable for naturally derived polymeric gelling agents. The gel

strength of the dry powder is retained, even after prolonged storage at 50 C

(1220F) and will recover from freezing. However, as with any polysaccharide,

such changes in temperature may tend to reduce its stability and should be


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avoided. Gellan gum should be stored tightly closed, in a cool (


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Heat of vaporization :705.4 J/g at B.P.

Melting point : 59C.

Refractive index :1.4324.

Description

Propylene glycol is a clear, colourless, viscous, and practically

odourless Liquid, with a sweet, slightly acrid taste resembling that of

glycerine.

Functional Category

Anti microbial preservative, co solvent, disinfectant, humectant,

plasticizer, stabilizing agent.

Applications in Pharmaceutical Formulation

As humectant in topicals up to 15%.

As preservative in solutions and semisolids up to 1530%.

As solvent or co solvent in aerosol solutions 10

30%, oral solutions

1025%, Parenterals 1060%, topicals 580%.

Stability and Storage Conditions

At cool temperatures, propylene glycol is stable in a well closed

container, but at high temperatures, it tends to oxidize, giving rise to products

such as propionaldehyde, lactic acid, pyruvic acid and acetic acid. Propylene

glycol is chemically stable when mixed with ethanol (95%), glycerine,

aqueous solutions may be sterilized by autoclaving.


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3.2.3 BENZOLKONIUM CHLORIDE70

Non proprietary Name :Benzalkonium Chloride.

Synonyms : Benzalkonii chloridum.

Chemical Name and CAS Registry

Documents

THESIS NASAL GEL