Devices for Selective Uptake of Surfactant from Ocular Drop Solution

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Devices for Selective Uptake of Surfactant from Ocular Drop Solution
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english
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Mittal, Anand
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University of Florida
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Degree:
Master's ( M.S.)
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University of Florida
Degree Disciplines:
Chemical Engineering
Committee Chair:
Chauhan, Anuj
Committee Members:
Jiang, Peng

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devices -- ocular -- surfactant
Chemical Engineering -- Dissertations, Academic -- UF
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Chemical Engineering thesis, M.S.
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Abstract:
Benzalkonium chloride (BAC) is an antimicrobial agent and preservative used in various ocular drops for treatment of Glaucoma. Usually,these preservatives are toxic and biocidal in nature which can cause numerous health related problems due to non-selective action. To minimize their content,the polymeric devices were synthesized which would act by for absorbing surfactant/preservative from ocular drops before it reaches eye. This absorption of surfactant can be maximized by increasing surface/contact area between the polymer and ocular drops. This in turn can be increased either by increasing the porosity of hydrogels or by making bed of micro-sized spherical particles. For this purpose, phase separated p-HEMA gels were made using solution polymerization with salt as diluting agent. Concentration of cross-linker, spacer thickness and amount of water were varied to obtain the gel that suited most for this purpose. Also, micro-sized particles were made via suspension polymerization with Magnesium Oxide (MgO) as dispersing medium. Solubility of HEMA in water decreased because of the salting-out effect of MgO. The gels and micro-particles were successful in selectively extracting BAC from drug solution.To test the uptake of surfactant in short period of time similar to real-life situation, an experimental setup was made by modifying oxygen-diffusion cell. The uptake in very short time intervals was not as large as expected, but the experimental apparatus designed was path-breaking in nature and could be put to several other applications with minor modifications.
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by Anand Mittal.
Thesis:
Thesis (M.S.)--University of Florida, 2013.
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Adviser: Chauhan, Anuj.
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1 DEVICES FOR SELECTIVE UPTAKE OF SURFACTANT FROM OCULAR DROP SOLUTION By ANAND MITTAL A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MAS TER OF SCIENCE UNIVERSITY OF FLORIDA 2013

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2 2013 Anand Mittal

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3 To my family

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4 ACKNOWLEDGMENTS First of all, I would like to thank Dr. Anuj Chauhan for giving me the opportunity to work in his lab and fo r the advice that he has given me not only in research related matters, but also in my future plans. Secondly, thank you to all of the group members who have allowed me to share the office/lab with them and advised me in developing new experimental procedu res especially Hyun Jung Jung and Lokendra kumar Bengani. Finally, I would like to thank my family for bearing with me and funding my education

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5 TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................ ................................ ................................ .. 4 LIST OF TABLES ................................ ................................ ................................ ............ 6 LIST OF FIGURES ................................ ................................ ................................ .......... 7 LIST OF ABBREVIATIONS ................................ ................................ ............................. 8 ABSTRACT ................................ ................................ ................................ ..................... 9 CHAPTER 1 INTRODUCTION ................................ ................................ ................................ .... 11 HEMA based Hydrogels ................................ ................................ ......................... 12 HEMA based Micro Particles ................................ ................................ .................. 14 Devices ................................ ................................ ................................ ................... 15 Mould To Increase Area Of Contact Between Drops And Polymer .................. 15 Experimental Setup T o Study Surfactant Uptake In Short Period Of Time ....... 15 2 EXPERIMENTAL ................................ ................................ ................................ .... 19 Materials ................................ ................................ ................................ ................. 19 Methods ................................ ................................ ................................ .................. 19 Preparation Of Hydrogels ................................ ................................ ................. 19 Preparation Of Particles ................................ ................................ ................... 20 3 CHARACTERI ZATION ................................ ................................ ........................... 22 Characterization of Hydrogels ................................ ................................ ................. 22 Surfactant Uptake With Time ................................ ................................ ............ 22 Surfactant Uptake With Scaled Time Axis ................................ ........................ 23 Partition Coefficient Of BAC In Hydrogel And Drop Solution ............................ 24 Characterization Of Particles ................................ ................................ .................. 25 Size Determination Of The Particles ................................ ................................ 25 Surfactant Uptake With Time ................................ ................................ ............ 25 4 CONCLUSIONS ................................ ................................ ................................ ..... 30 LIST OF REFERENCES ................................ ................................ ............................... 32 BIOGRAPHICAL SKETCH ................................ ................................ ............................ 33

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6 LIST OF TABLES Table page 1 1 BAC concentration (%) in ocular drops ................................ .............................. 11 3 1 Average actual thickness of gels ................................ ................................ ........ 24 3 2 Partition coefficient of hydrogel at 0.1% and 0.05% BAC concentration ............. 25

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7 LIST OF FIGURES Figure page 1 1 Suspe nsion polymerization technique ................................ ................................ 16 1 2 Copolymerization/Crosslink ing reaction of HEMA with EGDMA ......................... 17 1 3 Mould for polymerization ................................ ................................ .................... 17 1 4 Device to study short time interval uptake of BAC. ................................ ............. 18 3 1 Surfactant uptake at 0.1% BAC in solution ................................ ......................... 26 3 2 Surfactant uptake at 0.05% BAC in solution ................................ ....................... 27 3 3 Surfactant uptake at 0.15% BAC solution ................................ ........................... 27 3 4 Scaled Time Surfactant uptake at 0.1% BAC solution ................................ ........ 28 3 5 Scaled Time Surfactant uptake at 0.05% BAC solution ................................ ...... 28 3 6 Surfactant uptake by particles made using different methods at 0.05% BAC in solution ................................ ................................ ................................ ........... 29 3 7 Average uptake of triplicated samples of MgO. ................................ .................. 29

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8 LIST OF ABBREVIATIONS BAC Benzalkonium Chloride EGDMA Ethylene Glycol dimethyl methacrylate HEMA (Hydroxyethyl) methacrylate UV VIS Ultra violet visible

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9 Abstract of Thesis Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for th e Degree of Master of Science DEVICES FOR SELECTIVE UPTAKE OF SURFACTANT FROM OCULAR DR OP SOLUTION By Anand Mittal May 2013 Chair: Anuj Chauhan Major: Chemical Eng ineering Benzalkonium chloride (BAC) is a n antimicrobial agent and preservative used in various ocular drops for treatment of Glaucoma. Usually, these preservatives are toxic and biocidal in nature which can cause numerous health related pro blems due to n on selective action. To minimize their content the polymeric device s wer e synthesized which would act by for absorb ing surfactant /preservative from ocular drops before it reaches eye Thi s absorption of surfactant can be maximized by increasing surface /co ntact area between the polymer and ocular drops This in turn can be increased e ither by increasing the porosity of hydrogels or by making bed of micro sized spherical particles For this purpose p hase separated p HEMA gels were made using solution polyme rization with salt as diluting agent. Concentration of cross linker, spacer thickness and amount of water were varied to obtain the gel that suited most for this purpose Also, micro sized p articles were made via suspension polymerization with Magnesium Ox ide ( MgO ) as dispersing medium Solubility of HEMA in water decreased because of the salting out effect of MgO. The gels and micro particles were successful in selectively extract ing BAC from drug solution T o test the uptake of surfactant in short

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10 period of time similar to real life situation an experimental setup was made by modifying oxygen diffusion cell. The uptake in very short time intervals was n o t as large as expected, b ut the experimental apparatus designed was path breaking in nature and could b e put to several other applications with minor modifications

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11 CHAPTER 1 INTRODUCTION Since long, ocular drops are being administered into human body for curing several eye related problems and ailments T reatment using ocular drops is the prefe rred choice worldwide for treatment of diseases like glaucoma conjunctivitis among others These drops are made up of drug, drug vehicle, preservative and other chemicals from binding inner surface to plastic container. The presence of preservatives is ne cessary for maintaining the sterility of eye drop solution. Among others, B enzalkonium chloride (BAC) is a highly preferred preservative in anti glaucoma medications with varying concentrations, since it is a very effective preservative T able 1 1 depicts the amount of preservative in various drugs manufactured industrially Table 1 1 BAC concentration (%) in ocular drops 1 G eneric Manufacturer Preservative Apraclonidine Alcon 0.01% BAC Brimonidine/Timolol Allergan 0.005% BAC Brinzolamide/Timolol Alcon 0.01% BAC Carteolol Bausch & Lomb 0.005% BAC Dorzolamide MSD 0.0075% BAC Latanoprost Pfizer 0 .02 % BAC Timolol MSD 0.01 % BAC Categorized as quaternary ammonium group, BAC is a cationic surfac tant that can have multiple applications playing th e role of biocide and phase transfer agent. 1 Despite its extensive us ag e for making ocular formulations it has found criticism in

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12 recent years. BAC can cause disruption of intermolecular interaction and dissolution of cellular membrane lipid bilayers whic h essentially constitutes its biocid al nature 1 Also, t his dissolution of cell walls is n on selective in nature and thus can be a threat to living cell Prolonged exposure to ocular drops using BAC as preservative can cause several side effects and may e ven lead to illness. Some of the harmful effects described in literature are: dry eye syndrome and denaturing of corneal pro tein 2 C ontact loss between adjacent epithelial cells edema and dest ruction of cell can be other potential side effects 3 Also, i t can be cause for cause reversible conjunctiva fibrosis 4 Hence, the motivation of this thesis is to make ocular formulations safe to use by preventing the entry of surfactant into human eye The overall aim w ould be to selectively absorb BAC using a polym er just before it reaches eye. In a way, focus was on architecture and structure of the polymers so as to increase the available surface area inside polymers which can improve the overall uptake of BAC M acro porous polymers were chosen as answer for achi eving the aim of this project Macro porous cross linked polymers are very effective materials for separation processes and hence are also popular as starting material for ion exchange resins and specific sorbents. 5 Due to presence of network of macro siz ed pores the overall available area inside the polymers increases In presence of inert diluent, phase separation takes place during the free radical crosslinki ng reaction which leads to formation of these macro porous polymers 5 HEMA based Hydrogels Hydr ogels are categorized as hydrophilic polymer materials and can be defined as m ulticomponent systems made up of water or aqueous solution, fille d 3 D network

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13 of polymer chains 6 T he porosity of the hydro gels can be varied and is managed by controlling the thermodynamic phase behavior between polymer phase and aqueous phase The formation of porous structure is dependent upon a kinetic race between gel point and phase separation during polymerization process If gelation occurs fir st the resulting material is non macro porous but if phase sep aration occurs first they are macro porous in nature. Poly (2 hydroxy methyl acrylate) [ p ( HEMA)] falls under class of hydrophilic cross linked hydrogels that is bio compatible in nature and is easy to make and modify T heir structure and properties can be easily controlled by varying the reactants, their quantity and reaction conditions. Macro porous HEMA hydrogels have found several applications worldwide. Some of them include drug delivery, tissue regeneration, soft tissue replacement etc. Also, it is the preferred starting material for making contact lenses industrially 7 The HEMA material was found to be having greater affinity for BAC molecule as c ompared to Timolol This property of HEMA gels can be exploited f or selective ly separate BAC from ocular drops p (HEMA) hydro gels are usually made by s olution polymerization T h e water content of s olution is usually kept above 80% to make them macro porous 8 This water content if varied would cause change in porosity o f gels. Over period of time numerous other methods and advancements have been published worldwide for the preparation of macro porous hydrogels. The more vivid explanation about formation mechanism, preparation and properties of hydrogels can be found else where in literature 5

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14 Qing Liu et al. 9 used a novel technique to fabricate macro porous hydrogels by enhanced phase separation using an aqueous NaCl solution. Using this method, the gels were synthesized although there was variation in p olymerization meth od and quantity of reactants used HEMA b ased Micro P articles More re cently, cross linked micro particles using HEMA monomer as starting material have found wide rang e of applications like immobilization of enzymes, controlled release of pharmaceutical ag e nts, immunochemical studies in biomedical industry The reason why they are useful is because of their non toxicity, non carcinogenicity and high biocompatibility. Among others, t he property of HEMA that favors selectivity towards BAC would be applicable here too. And so, micro sized particles could be substitute for hydrogels for separatin g BAC out of ocular drops In past, m icro sized particles have be en synthesized usi ng various techniques Su spension polymerization is one of the several methods which c an generate micro particles In this method, t he size and shape of particles is dependent upon reaction conditions, reactants agitation speed and the design of reactor/vessel in which particles are made. But, due to high solubility of HEMA it becomes har d er to control their size and shape. Using the method to decrease their solubility, p articles of favorable shape and size can be prepared. Kiremicti et al. developed one such method by dissolving MgO in water which renders the solution less attractive to H EMA. Thus, it eliminates use of polymeric stabilizers in the aqueous phase Besides method used b y Kiremicti, hydrophilic polymer particles were dev is ed using Vitamin E oil as suspension medium Another based on reducing the solubility of monomer HEMA b y using a large polymer was thought as an alternate So, PVP was

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15 dissolved in water to reduce solubility of HEMA. Though these methods yielded particles, the control over their size and shape of particles w as minimal. Also, the particles obtained were not p orous enough and uptake was not as high Devices Mould T o I ncrease A rea O f C ontact B etween D rops A nd P olymer Since the aim of this project was to maximize the contact area between gel and drops, using mould was perceived as another possibility A simple mo uld made up of pins and thick piece of cardboard was designed to increase the available surface area in side polymer. Several different pins were pierced through the cardboard such that one side of cardboard had all the pin heads a nd the other side had the pointed end of pin s lined up very closely. The pins were placed such that the mould could stand properly on an even surface. The mould wa s placed such that the pin ends were kept dipped inside the monomer solution poured in a petri dish The system was pla ced on UV illuminator in such a way that UV light entered from the bottom of dish. On polymerization t he pins of mould dipped inside the solution got embedded to the polymer. When the mould was pulled out it left created several holes or gaps on the thic k polymer gel sheet These holes may increase interaction between gel and drops as the contact between them would be greater when drops flow via them Exper imental Setup T o Study S urfactant U ptake I n Short P eriod O f T ime Testing the gels for their uptake in very short period of time, was proving to be difficult and there was a need of system which resembled real life situation. Aim of this project hypothesized that polymer sh ould absorb the BAC from a drop just before it reaches the eye surface. And so th e time it stays w ithin the polymer w ould range from nanoseconds to milliseconds depending upon pressure applied. Since time

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16 of interaction between gel and solution is small, the diffusion cell was modified and designed so that liquid solution flows in and out of it quickly One end of the cell was attached to the syringe to introduce the eye drug solution inside the cell. The other end was kept open so as to collect the solution coming out of gel Two gel sheets of equal thickness were placed in between the two plates. The gel sheets were separated by 400 separator sheet placed along the edges of the gel C ell was clamped from all sides s uch that the gel s were packed firmly The solution was poured into the cell and passed via gel by applying pressure using syringe. The flow of the solution via gel was dependent on the applied pr essure. It can be proposed that this flow followed Hele Shaw flow mechanism. This is because the governing equation of Hele Shaw flow is same as that of the flow via porous media. A lthough the apparatus was able to test the flow of solution in short period of time it had some drawbacks and limitations. The major problem faced was leakage from the gap between two plates. Also, due to limited time exposure to gel the surfact ant uptake was not as high as expected. T h e device will not be useful to test the particles too, as it is difficult to hold them between two plates like gel Figure 1 1 Suspension polymerization technique. 10 [adapted from M. T. Gokmen and F. E. D. Prez Progres s in Polymer Science, 2012, 37 365 405]

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17 Figure 1 2 Copolymerization/Crosslinking reaction of HEMA with EGDMA 11,12, [adapted from M. Kiremitci and H. Cukurova ; Polymer International, 1993, 30(3) 357 361] Figure 1 3 Mould for polymerization [Photo courtesy of Anand Mittal]

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18 Fig ure 1 4 Device to study short time i nterval uptake of BAC [Photo courtesy of Anand Mittal] Sy ringe Beaker to collect solution Modified Diffusion Cell

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19 CHAPTER 2 EXPERIMENTAL Materials 2 hydroxymethyl methacrylate (HEMA) monomer Ethylene glycol dimethacrylate (EGDMA) Sigma Aldrich C hemicals (St. Louis, MO). 2 4, 6 Trimethylbenzoyl diphenyl phosphineoxide (Darocur TPO ) was kindly provided by Ciba Specialty Chemicals (Tarrytown, NY). S odium chloride (NaCl) was purchased from Fischer Scientific Acrylic acid and Magnesium Oxide (MgO) w ere purchased from Sigma Aldrich Chemicals (St. Louis, MO). 2 azobis ( 2 methyl propionitrile), the initiator was obtained from Ciba Vision. All the Methods Preparation O f H ydroge ls As mentioned earlier, solution polymerization/crosslinking technique was employed for the preparation of macro porous hydrogels. Aqueous sodium chloride solution of 0.7 M was used as diluen t In all experiments, the monomer to diluent volume ratio was m aintained constant In a typical method, 5.4 mL HEMA monomer, 4 mL of 0.7M NaCl solution and 0.030 mL of EGDMA were mixed in a clean vial to make a monomer solution The solution was stirred for five minutes using magnetic stirrer. The mixture was purged w ith bubbling nitrogen for 15 min to remove dissolved oxygen. Dissolved oxygen acts as radical scavenger which leads to loss of radicals generated from photo initiator. Hence to prevent this loss of radical, degassing is done.

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20 After degassing 0.0070 gm. o f Darocur TPO was added to the solution. The solution was stirred again at 600 rpm for 5 minutes to properly dissolve the photo initiator. In the meantime, apparatus for polymerization was prepared. Using t wo clean, transparent 5 mm thick glass plates sepa rated by a plastic spacer of desired thickness a m ould for polymeriz ation was made S pacers of four different thicknesses like 100m, 200m, 400m and 800m were used to get the gels of diff e rent sizes The plates were clamped using binding clips from 3 s ides to hold the solution between plates. One end was left open to pour the solution between plates. After five minutes of stirring, t he solution was poured in between the two plates using pipette. The glass plate apparatus was irradiated with UV B for ar ound 1 hour utilizing an ultraviolet Trans illuminator UVB 10 (UltraLum, Inc.) The prepared gels were boiled 3 times in boiling water for 1 hr each. To thoroughly leach out the salt and unreacted monomer from polymer water was changed every time. The ge ls were kept hydrated in water for further testing. Preparation O f Particles Recipe developed by Kiremitci & Cukurova was used for making poly (HEMA) particles For an appropriate amount of monomer (HEMA), the cross linker was kept at 0.3 mol/mol and the initiator at 0.0015 mol/mol. The MgO solution contained 0.015 g MgO/ml which made up the aqueous phase. 11 So, 1.2 mL of HE M A and 0.0024 gm. of initiator were mixed to make suspension medium in clean vial By varying the amount of EGDMA in monomer phase, th e size of particle was varied to get optimum size of particles. For 1.2 mL of HEMA, three different quantities of EGDMA (0.2 mL, 0.3 mL and 0.4 mL) were tried out.

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21 1 2 mL of MgO solution in water constituting the aqueous phase was mixed to suspension phas e. Th e mixture was kept at 70 C for 3 h and at around 85 90 C for 1 h using a water bath Solution was continuously stirred using magnetic stirrer Due to continuous stirring of solution, small globules containing HEMA monomer and EGDMA were formed in th e solution which remained suspended in aqueous phase. These globules acted as individual tiny reactors for polymerization reaction which led to formation of micro sized particles. After completion of polymerization the p articles were transferred in to a s eparate beaker to wash away impurities and unreacted monomer The residual MgO stuck to particles was dissolved by adding dilute hydrochloric acid solution. The particles were washed next with water and ethanol Finally, they were dried in a vacuum desicca tor at room temperature. For making particles using oil monomer solution similar to one used for making gels was made but was mixed with equivalent amount of Vitamin E oil. The amount of reactants and diluent was kept same The mixture was stirred at cons tant temperature of 70C for 3 hrs. After polymerization, the particles were washed with ethanol and water to remove impurities. Similarly for making the particles using PVP, powder ed PVP was dissolved in water. The monomer phase containing HEMA, EGDMA and initiator was mixed and stirred at cons tant temperature for 3 hours. Similar washing procedure was repeated for these particles.

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22 CHAPTER 3 C HARACTERI ZATION Characterization of H ydrogels Surfactant Uptake W ith T ime UV Visible spectroscopy method was u sed for determination of BAC uptake Many molecules in solution have the ab ility to absorb UV and visible light. Due to the difference in their size, they show different absorption spectrum from each other. Also, there is change in spectrum with the change in concentration of these molecules Beer Lambert 's Law states that absorbance A = bc where is a constant of proportionality, called the absorptivity b is a path length and c is concentration of solution Hence, u sing absorption spectrum and intensity peaks of the blank sample and solution measured at proper intervals of time, the change in concentration of solution with time can be determined. T he standard solution containing drug and surfactant was made by dissolving them in PBS, pH 7.4 The concentration of drug ( Timolol ) was kept constant at 100 ppm (0.01%) The concentration of the surfactant (BAC ) was varied from 0.05% to 0.1 5% To ensure the authenticity and reproducibility of the results, the samples are usually triplicated. And to do so, 0 .05 gm. of hydrogel sheet was placed in three different vials. 5mL of solution was pou red in each vial. The absorption spectrum was measured at proper intervals of time starting from zero. A control experiment without gels was conducted to ensure that surfactant adsorption to the vials was negligible. As evident from Figure 3 1, the 100m g el attains equilibrium at shortest time. As the thickness of the gel increases, the equilibration time increases. But, the net uptake

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23 in every gel is same for a given concentration of surfactant in solution. At 0.1% BAC concentration, the net uptake is 80 mg per gram of gel. Fig ure 3 2 shows that at 0.05% BAC the graph above shows similar trend but there is one major difference between the two graphs. Theoretically, the net uptake should decrease proportional to the concentration of surfactant in the soluti on. This assumption was proved correct as the net uptake at 0.05% concentration was halved to 40 mg per gram of gel. Similar ly when the BAC conce ntration was increased to 0.15% of solution, t he net uptake i ncreased to 120 mg per gram of the gel (Fig 3 3) Surfactant Uptake W ith Scaled T ime A xis Surfactant uptake in Fig 3.1 3.3 show strong dependency on the thickness of the gel. The uptake profile suggests that transport is controlled by surfactant diffusion into gel. Also since the ratio of volume of surfac tant solution to gel mass was kept fixed for every sample, the equilibrium time should scale as the square of the gel thickness for a purely diffusion controlled process. 13 And so to verify the claim that the uptake is diffusion controlled the data for sur factant uptake was plotted as a function of scaled time. Theoretically, s caled time can be defined as : So, the t ime axis was modified by dividing the time s by squared of the average actual thickness. The factor of H 0 is constant and if neglected will n ot affect the data much. The actual thicknesses of the gels were determined by measuring single gel film at three different locations and taking their average. The thickness of dry gels was determined using Vernier caliper.

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24 Table 3 1 Average actual thick ness of gels Spacer Thickness Reading1 Reading2 Reading3 Av era g e St d. dev iation 100 65 64 64 64.33 0.57 200 152 153 154 153 1 400 428 429 425 427.33 2.08 800 952 952 952 The graphs in Fig ure 3 4 and 3 5 show the uptake by hydrogels when measured v s. scaled time The graphs show that the uptake trend lines do not overlap S imilar, result was obtained at 0.05% with net uptake being hal f that of at 0.1% BAC concentration. So, i t can be concluded that there is non homogeneous diffusion inside the gels Also, there is increased uptake in the gels due to presence of micro sized pores within the gels. When the drop solution enters the gels, it fills up the pores quickly And then there is slow diffusion of BAC inside the gel material. This may be reason w hy there is non homogeneous diffusion inside gels. Partition Coefficient Of BAC I n Hydrogel A nd D rop S olution Partition coefficient of hydrogels indicate the ratio of the concentration of the uptake of surfactant in gel to the concentration of the surfacta nt in original solution. Determination of this coefficient can be used to prove the preference of the compound to one phase as comp ared to the other. Similar method has been used earlier to prove the uptake of lysosomes from solution 1 3 by demonstrating hig h values of the partition coefficient a s evidence T he preference of the BAC to hydrogel over solution can be verified by calculat ing the high values of p artition coefficient from the data available with us.

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25 Table 3 2 Partition coefficient of hydrogel a t 0.1% and 0.05% BAC concentration 0.1% BAC 0.05% BAC Gel Thickness Kavg Stdev Kavg Stdev 100 422.10 28.023 397.41 37.27 200 4 42.92 11.70 302.04 81.02 400 442.92 20.89 361.00 27.78 800 415.39 68.94 300.19 41.93 As evident from Table 3 2, t he partition coefficient s for gel are pretty high and can be attributed to preference of the surfactant to gels over solution. Characterization O f P articles Size D etermination O f T he P articles Since the particles were big ger and could be measured physic ally, their size was determined manually using digital images of the particles It was estimated that the diameter of the particles was around 400 600 Surfactant Uptake W ith T ime UV Vis spectroscopy method was used to determine the uptake by the partic les. Similar to method for determining the uptake by hydrogels, the surfactant solution at 0.1% concentration was prepared by dissolving BAC in PBS solution. Same solution containing drug and surfactant in PBS was prepared. The particles made using the thr ee routes were cleaned, measured and placed in vial. The surfactant solution was poured into vial such that the ratio of the mass of particles to volume of surfactant solution was constant. T he absorption spectrum was dete rmined at proper time intervals a nd control experiment was done to ensure negligible absorption of surfactant by gels.

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26 As evident from the graph in Fig ure 3 6 the uptake by particles made out using MgO suspension method was highest at any point of time. Also, the data showed that drug up take by these particles was very minimal. And so, the particles made out of MgO suspension method were tested further to verify the results by testing the triplicate of samples. From Fig ure 3 7, it can be concluded from the data that the particles can sele ctively absorb the BAC from eye drops. The bed of particles made out of these particles can be used for detoxification of drops. Figure 3 1 Surfactant uptake at 0.1% BAC in solution

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27 Figure 3 2 Surfactant uptake at 0.05% BAC in solution Figure 3 3 Surfactant uptake at 0.15% BAC solution

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28 Figure 3 4 Scaled Time Surfactant uptake at 0.1% BAC solution Figure 3 5 Scaled Time Surfactant uptake at 0.05% BAC solution

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29 Fig ure 3 6 Surfactant uptake by particles made using different methods a t 0.05% BAC in solution Figure 3 7 Average uptake of triplicated samples of MgO

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30 CHAPTER 4 CONCLUSIONS Although many attempts have been made recently for alternative preservatives in ocular drugs, BAC is still preferred worldwide as an antimicrobial pr eservative as it is cheaper to make industrially. To prevent the harmful toxic effects of BAC on human eye, polymeric devices were developed which c an selec tively soak BAC from ocular drop s. The HEMA gels made using enhanced phase separation method were ai med to be macro porous in nature so as to increase the contact between drops and polymer in shortest possible time These gels showed selective uptake of BAC over Timolol effectively over a period of time. Although the uptake v / s time graph was obtained as per expectation, scaled time graphs were not overlapping. Reason why this is happening is that upta ke by these gels is due to non homogeneous diffusion The possible hypothesis for inhomogeneous diffusion can be that after entering the gels, the solution flow in vertical directions (where pores exist) takes place rapidly as they do not face resistance from vacuum After filling the se pores, the flow in horizontal direction begins which is slower comparatively due to the resistance from gel material. Also, the flow of liquid solution across gels was very slow which can derail our objectives of obtaining solution at other end. Porous m icroparticles can be other potential incumbents for selective uptake of BAC as the bed made out of this particles can incre ase the contact between polymer and drops The se particles also show ed selective uptake of BAC over Timolol. Hence, t he bed made out of these particles can be used for extracting toxic contents of drug including BAC. ross the bed.

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31 The experimental setup constructed to study uptake of polymers in short period of time is innovative and can be utilized in other studies to understand the diffusion of liquids into solids. The results of this study could be applied to stud y various other methods to reduce the side effects of using preservatives and other toxic ingredients in ocular dr ops

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32 LIST OF REFERENCES 1 Y Louati and T Shaarawy J Current Glau Prac 2012 6(3) 104 107 2 K. Swan Am J Ophthalmol 1994 27 118. 3 P. Y. Lin, Ophthalmology 2003 10 1096 1101 4 S. E. Moss, Arch Ophthalmology 2000 118 1264 12 68 5 O. Okay, Prog. Polym. Sci., 2000 25 711 779 6 N. Irina Savina, P E. Tomlins, S V. Mikhalovsky and I Y. Galaev Macroporous polymers, 2009, 9 211 235 7 N. A. Peppas and H. J. Moynihan Hyd rogels in medicine and pharmacy, 1987, 2 49 64. 8 K. P. Antonsen J. L. Bohnert Y. Nabeshima, M. S. Sheu X. S. Wu and A. S. Hol man Biomater Artif Cells Immobil Biotechnol 1993 21 (1) 1 22. 9 Q. Liu E L. Hedberg Z L iu, R Bahulekar, R K. Meszlenyi and A G. Mikos, Biomaterials 2000 21 2163 21 69 10 M. T. Gokmen and F E. Du Prez Progress In Polymer Science, 2012, 37 365 405 11 M. Kiremitci and H. Cukurova, Polymer International 1993 30 ( 3 ) 357 361 12 B. Podkol cielna A. Bartnicki and B. Gawdzik eXPRESS Polymer Letters 2012 6(9 ) 759 771 13 L. C. Bengani, J. Leclerc and A Chauhan, J Colloidal Sci., 2012, 3 86 441 450

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33 BIO GRAPHICAL SKETCH A nand Mittal was born in Mumbai C ity, India in the year of 1988. He completed his undergraduate studies at Visvesvaraya National Institute of Technology, Nagpur and obtained Bachelor of Technology (B.Tech) degree He was a graduate student at University of Florida and finish ed in the May of 2013 with Master of Science (MS) degree