DEVELOPMENT OF ENOXAPARIN SODIUM POLYMERIC MICROPARTICLES FOR COLON-SPECIFIC DELIVERY

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1 DOI: /cjmed-442 DEVELOPMENT OF ENOXAPARIN SODIUM POLYMERIC MICROPARTICLES FOR COLON-SPECIFIC DELIVERY DANA HALES 1, MAXIME CASTERAN 2, ANNE SAPIN-MINET 2, IOAN TOMUŢA 1, MARCELA ACHIM 1, LAURIAN VLASE 1, PHILIPPE MAINCENT 2 1 Deprtment of Phrmceuticl Technology nd Biophrmcy, Fculty of Phrmcy, Iuliu Htiegnu University of Medicine nd Phrmcy, Cluj-Npoc, Romni 2 Deprtment of CITHEFOR, EA 3452, Université de Lorrine, Frnce Astrct Bckground nd ims. Recent studies hve shown tht low moleculr weight heprins re effective in the tretment of inflmmtory owel disese. Therefore, there is considerle interest in the development of n orl colonic delivery phrmceuticl system llowing trgeted relese of heprin in the inflmed tissue. The ojective of this study ws to prepre microprticles for the orl dministrtion nd colonic relese of enoxprin nd to evlute the influence of certin formultion fctors on their chrcteristics. Methods. Microprticles were prepred y wter/oil/wter doule emulsion technique followed y solvent evportion. The influence of severl formultion fctors on the chrcteristics of microprticles were evluted. The formultion fctors were lginte concentrtion in the inner queous phse, polymer (Eudrgit FS 30D nd Eudrgit RS PO) concentrtion in the orgnic phse nd rtios etween the two polymers. The microprticles were chrcterized in terms of morphology, size, entrpment efficiency nd enoxprin relese. Results. The results showed tht incresing sodium lginte percentge reduced the encpsultion efficiency of enoxprin nd ccelerted enoxprin relese. Regrding the influence of the two polymers, reducing polymer concentrtion in the orgnic phse led to smller size of microprticles, lower entrpment efficiency nd n importnt retrdtion of enoxprin relese. The formultion prepred with Eudrgit FS 30D limited the relese to mximum of 3% in gstric simulted environment, specific chrcteristic of orl systems for colonic delivery, nd fulfilled our ojective to dely the relese. Conclusions. Microprticles prepred with Eudrgit FS 30D represent suitle nd potentil orl system for the colonic delivery of enoxprin. Keywords: microprticles, colonic delivery, enoxprin, Eudrgit FS 30D, Eudrgit RS PO Bckground nd ims Heprin hs een used for more thn 80 yers s n nticogulnt nd dministered y prenterl route. In the lst decdes, mny groups of reserchers ttempted to Mnuscript received: Accepted: Address for correspondence: tomution@umfcluj.ro improve heprin non-invsive drug delivery systems, in order to enhnce ptient complince nd minimize side effects [1]. Two min strtegies hve een used to llow orl dministrtion of heprin: incresing the intestinl sorption with permetion enhncers [2,3,4,5] nd incresing the stility y the encpsultion of heprin into vrious phrmceuticl systems microprticles (MP) 357

2 Phrmcy [6,7], nnoprticles [8,9] or pellets [10]. A few reserch groups hve studied the development of polymeric nno- or micro-systems for the orl delivery of low moleculr weight heprins (LMWH). Wter/oil/ wter doule emulsion technique followed y solvent evportion ws used for the encpsultion of heprin. This method, developed y Alex nd Bodmeier in 1990 [11], llows the encpsultion of hydrophilic ctive principles in hydrophoic mtrix. A polymer tht gve good results in terms of LMWH s encpsultion efficiency nd drug relese ws Eudrgit RS PO (ERS), methcrylic ctionic copolymer, which llows time controlled relese y the presence of mmonium slts tht mke the polymer permele [12]. Tinzprin-loded nnoprticles which were prepred with ERS hd n nticogulnt effect prolonged up to 8 h [9]. Also, ERS microprticles loded with tinzprin nd ndroprin, hd higher nd stle nti-x/nti-ii rtio compred to the commercil rtio [13]. Recently, these studies hve ecome even more interesting since there is proof of other potentil pplictions of heprins, including cncer nd vrious inflmmtory diseses, such s inflmmtory owel disese [14]. The tretment of these diseses is sed on the immunomodultory nd nti-inflmmtory properties, such s modultion of cytokine production, cytotoxic ctivity of T lymphocytes nd inhiition of dhesion, leukocyte ctivtion nd trnsporttion [15]. Therefore, there is mjor interest in the development of n orl colonic delivery phrmceuticl system llowing trgeted relese of heprin in the inflmed tissue. One of the most common colonic dosge forms re ph-controlled relese systems [16,17,18,19], which use polymers or polymer lends in order to protect the ctive drug from the gstric nd intestinl fluid nd t the sme time enle its relese in specific regions of the gstrointestinl trct [20]. The types of ph controlled relese systems rnge from single-unit tlets or cpsules [16] to multiprticulte formultions, such s pellets, grnules, microprticles nd nnoprticles [21]. Microprticles re one of the phdependent systems tht showed potentil s colonic delivery dosge forms nd the methods used for their preprtion were commonly doule emulsion technique followed either y solvent evportion or y solvent extrction [13,22], ut lso spry-drying [23] or emulsion dehydrtion technique followed y oil-in-oil solvent evportion method [24]. There is lrge vriety of ph-dependent polymers commercilly ville, mong them polyvinyl cette phthlte nd cellulose cette phthlte or copolymers of crylic nd methcrylic cid, known s Eudrgit. Eudrgit P-4125F, ph-dependent polymer used to prepre microprticles for the colonic delivery of enoxprin, prevented LMWH relese t ph<6 nd llowed fst relese t ph 7.4 [22]. Eudrgit FS 30D (EFS) is n nionic methcrylic copolymer which llows trgeted colon delivery y dissolving ove ph 7.0 y slt formtion [25]. This polymer ws previously used for colonic drug delivery y other reserchers, oth in conventionl phrmceuticl dosge forms s tlets nd pellets [26,27,28], or novel phrmceuticl dosge forms s microprticles [29], ut not for the encpsultion of LMWH. The min purpose of this study ws to develop nd evlute the influence of certin formultion vriles type nd rtios of polymers (Eudrgit RS PO, Eudrgit FS 30D) used s controlled relese polymers, concentrtion of sodium lginte (NAlg) introduced in the queous phse on the chrcteristics of enoxprin-loded microprticles: morphology, size, entrpment efficiency nd in vitro relese. Mterils nd methods Mterils Mrketed sodium enoxprin (Clexne UI nti-x/1 ml) ws purchsed from Snofi-Aventis (Frnce). Alginic cid sodium slt from rown lge (medium viscosity, 2000 cp, 2% (25 C)) nd poly(vinyl lcohol) (87-90% hydrolyzed, verge mol wt ) were purchsed from Sigm-Aldrich Chemie GmH (Germny). Eudrgit FS 30D nd Eudrgit RS PO were received y courtesy of Evonik (Germny). All other chemicls were of nlyticl grde. Preprtion of microprticles The microprticles were prepred y wter/oil/ wter doule emulsion technique nd solvent evportion, s previously descried [11]. Tle I shows the composition of enoxprin microprticles. The formultions F1-0 to F were intended to study the influence of the concentrtion of sodium lginte nd the type of polymer on the chrcteristics of microprticles nd the formultions F9-0 to F studied the influence of EFS/ERS rtio. Drugfree microprticles nd enoxprin-loded microprticles were prepred y the sme method in order to study if drug loding hd influence on the chrcteristics of microprticles. Briefly, Eudrgit FS 30D, Eudrgit RS PO or their mixture were dissolved in dichloromethne (DCM). Then, wter or n queous solution of sodium lginte (0.5%, 1% or 1.5%) with or without enoxprin, ws emulsified into the orgnic polymer solution using n ultrsound proe (Vircell, Frnce) for 30 seconds. The resulting w/o-emulsion ws then poured into n queous solution of poly(vinyl lcohol) (1%) nd stirred with mgnetic stirrer for 1.5 minutes t 510 rpm to otin the w/o/wemulsion. This emulsion ws dded to 400 ml wter nd stirred with three-ldes propeller for 1.5 h t 500 rpm t room temperture in order to llow the evportion of the orgnic solvent from the internl phse. During solvent evportion, the polymers precipitted nd the microprticle 358

3 Tle I. Composition of enoxprin microprticles. Code Eudrgit FS (%) Eudrgit RS (%) cores solidified. Microprticles were collected y filtrtion (Porfil, CA, 0.2 µm) nd frozen t -30 C [11]. Sodium lginte (%) Clexne (sodium enoxprin) UI/ ml (ml) DCM (ml) F F F F F ml (5000 UI) 2 F ml (5000 UI) 2 F ml (5000 UI) 2 F ml (5000 UI) 2 F F F F F ml (5000 UI) 2 F ml (5000 UI) 2 F ml (5000 UI) 2 F ml (5000 UI) 2 F F F F F ml (5000 UI) 5 F ml (5000 UI) 5 F ml (5000 UI) 5 F ml (5000 UI) 5 encpsultion efficiency ws expressed s the percentge of enoxprin entrpped in reltion to the theoreticl vlue. Morphology of microprticles The morphology of microprticles ws nlyzed y n opticl microscope equipped with cmer (Nikon Eclipse Ti-S, Frnce). Size of microprticles The men dimeter of microprticles ws evluted y lser light diffrction, using Mstersizer 2000 device (Mlvern Instruments, UK). In this respect, 20 mg of microprticles were re-suspended in 2 ml queous solution of Tween 80 (0.1%) nd dispersed in n ultrsonic th for 5 minutes. Ech smple ws mesured in triplicte. Encpsultion efficiency The encpsultion efficiency (%) of enoxprin ws determined from the externl queous phse y n indirect turidimetric method sed on the quntittive precipittion rection occurring etween sulfte nd croxyl groups of heprin nd the mine groups of cetylpyridinium chloride t ph 6.8 [30]. All experiments were performed in triplicte. Aliquots (500 μl) of ech smple were rected for 1 hour t 37 C with sodium cette uffer (500 μl) nd n queous solution of cetylpyridinium chloride 0.1% in NCl solution (2 ml). The precipittes were ssyed y spectrophotometry (Shimdzu, Jpn) t 500 nm. The In vitro drug relese study The enoxprin relese from the microprticles ws ssessed y dissolution testing using wter th mintined t temperture of 37 C nd plced on mgnetic stirrer djusted t rottion speed of 200 rpm. Microprticles were suspended in 20 ml HCl 0.1M or simulted gstric fluid (ph 1.2) for 2 h, then the medium ws replced y 20 ml phosphte uffered sline (ph 6.8) for nother 3 h, nd finlly, y 20 ml phosphte uffered sline (ph 7.4) until the end of the 24 h in order to simulte the ph vlues in the stomch, the proximl nd middle smll intestine (duodenum nd jejunum), nd the distl smll intestine (ileum) respectively. Smples of 1.5 ml were withdrwn t 0.5, 1, 2, 3, 4, 5, 6, 8 nd 24 h time intervls nd replced with n equl volume of fresh medium. The content of enoxprin sodium in the withdrwn smples ws nlyzed y the spectrophotometric method descried ove, t 500 nm. Sttisticl evlution The results were expressed s men vlues ± S.D. Sttisticl nlysis ws crried out with the Anlysis Toolpk from Excel Anlysis of vrince ws used to nlyze the differences etween groups nd regression nlysis to ssess whether there ws liner connection 359

4 Phrmcy etween independent nd dependent vriles. In ll cses, proility vlue of less thn 0.05 ws considered to e significnt. Results microprticles morphology nd size Microscope imges of enoxprin-loded microprticles re presented in Figure 1 nd the influence of the concentrtion of sodium lginte nd the type of polymer on the size of microprticles is displyed in Tle II. Tle III presents the size of microprticles ccording to the EFS/ERS rtio. encpsultion efficiency Tle IV shows the influence of the concentrtion of sodium lginte nd type of polymers (Eudrgit FS or Eudrgit RS) on the encpsultion efficiency, wheres Tle V presents the encpsultion efficiency vlues otined with different mixtures of the two polymers. Influence of the formultion fctors on in vitro drug relese The relese of enoxprin from polymeric microprticles, depending on the concentrtion of sodium lginte nd type of polymers, is shown in Figure 2. Figure 3 presents the relese profile of enoxprin F (0% NAlg) F (0.5% NAlg) F (1% NAlg) F (1.5% NAlg) F (0% NAlg) F (0.5% NAlg) F (1% NAlg) F (1.5% NAlg) c F (100% EFS) F (75% EFS) F (50% EFS) F (0% EFS) Figure 1. Microprticles nlyzed with opticl microscope (x10). EFS microprticles prepred with different NAlg concentrtions; ERS microprticles prepred with different NAlg concentrtions; c Microprticles prepred with lends of EFS/ERS in different weight rtios. Tle II. Influence of sodium lginte concentrtion nd type of polymer on the size of microprticles (MP). Conc. Eudrgit FS, Eudrgit RS, of Drug-free MP Drug-loded MP Drug-free MP Drug-loded MP NAlg (%) Code Size (µm) d Code Size (µm) Code Size (µm) d Code Size (µm) c 0 F ±23 F ±17 F5-0 68±11 F ± F ±38 F ±119 F ±1 F ±30 1 F ±64 F ±56 F ±2 F ± F ±29 F ±3 F ±66 F ±34 Dt re shown s men ± S.D. (n=3); Sttisticlly different for drug-free formultions compred to drug-loded formultions (p<0.05); c Sttisticlly different depending on the concentrtion of sodium lginte (p<0.05); d Sttisticlly different depending on the type of polymer (p<0.05). from microprticles prepred using different rtios of Eudrgit FS/Eudrgit RS. 360

5 Tle III. Influence of EFS/ERS rtio on the size of microprticles. Rtio of polymers (%) Drug-free MP Drug-loded MP Eudrgit FS Eudrgit RS Code Size (µm),,c Code Size (µm),,c F ±2 F ± F ±12 F ± F ±33 F ± F ±4 F ±12 Dt re shown s men ± S.D. (n=3). No sttisticl difference for drug-free formultions compred to drug-loded formultions (p>0.05); c No sttisticl difference depending on the EFS/ERS rtio (p>0.05). Tle IV. Influence of the concentrtion of sodium lginte nd type of polymer on encpsultion efficiency (EE). Concentrtion of Eudrgit FS,,c Eudrgit RS,,c sodium lginte (%) Code EE (%) Code EE (%) 0 F ±7 F ±1 0.5 F ±11 F ±5 1 F ±10 F ±4 1.5 F ±5 F ±10 Dt re shown s men ± S.D. (n=3); No sttisticl difference relted to the concentrtion of sodium lginte (p>0.05); c Sttisticlly different depending on the type of polymer (p<0.05). Tle V. Influence of EFS/ERS rtio on encpsultion efficiency (EE). Rtio of polymers (%) Eudrgit FS Eudrgit RS Code EE (%), F ± F ± F ±4-100 F ±1 Dt re shown s men ± S.D. (n=3); Sttisticlly different depending on the rtio of EFS/ERS (p<0.05). * Sttisticlly different t ll times depending on the type of polymer (p<0.05); Sttisticlly different t ll times from F (ERS, 1.5% NAlg) (p<0.05). Figure 2. In vitro relese profile of enoxprin from microprticles. Influence of sodium lginte on enoxprin relese in prticles prepred with Eudrgit FS. Dt re presented s men ± S.D. (n=2); Influence of sodium lginte on enoxprin relese in prticles prepred with Eudrgit RS. Dt re presented s men ± S.D. (n=2). 361

6 Phrmcy * Sttisticlly different t ll times depending on the rtio of EFS/ERS (p<0.05). Figure 3. In vitro relese profile of enoxprin from microprticles Influence of Eudrgit FS/Eudrgit RS rtio on enoxprin relese (NAlg=0%) Discussion microprticles morphology The imges nlyzed under the microscope reveled tht the enoxprin-loded microprticles hd firly sphericl shpe nd dense spect. The differences in size etween different types of microprticles could lso e seen, nd the visul oservtions were correlted with the size determintion. The increse of the concentrtion of sodium lginte incresed the size of prticles, while Eudrgit FS determined the formtion of prticles greter in size thn Eudrgit RS (Figure 1-). The microprticles prepred with one type of polymer (F nd F ) hd smller size compred to the ones prepred with the mixture of EFS/ERS (F nd F ) (Figure 1c). Figure 1 lso showed tht the increse of DCM volume, which consequently decresed the viscosity of the polymeric orgnic solution, led to reduction of the size of prticles: F formultion presented smller prticles compred to F formultion nd F formultion showed smller prticles compred to F formultion. microprticles size As shown in Tle II, the results indicted tht drug-free microprticles hd smller dimeter ( µm) compred to enoxprin-loded microprticles ( µm). Other reserchers otined similr results for Eudrgit RS microprticles, unloded (27 µm) nd loded with other low moleculr weight heprins, tinzprin (49 µm) nd ndroprin (53 µm) [13]; therefore, it my e concluded tht encpsultion of enoxprin incresed the size of prticles. These studies suggested tht the surfctnt properties of Eudrgit RS determined the smll dimeter of the unloded prticles [13]. In the cse of heprin-loded prticles, the electrosttic interctions etween positively chrged Eudrgit RS nd negtively chrged enoxprin sodium led to decrese of the surfctnt properties of the polymer nd the formtion of lrger prticles. Another fctor which influenced the size of the microprticles ws sodium lginte: overll, s the concentrtion of sodium lginte ws higher, the microprticles were lrger (Tle II). However, its influence ws sttisticlly significnt only for Eudrgit RS enoxprin-loded microprticles; the explntion is the existence of electrosttic interctions mentioned efore. The comintion of positively chrged Eudrgit RS nd negtively chrged sodium lginte leds to comintion similr to the one of Eudrgit RS nd enoxprin, resulting in reduced surfctnt properties of the polymer nd formtion of incresingly lrger prticles with incresing concentrtions of lginte. Although the results were not significnt in the cse of Eudrgit FS, the size vlues incresed from no lginte to 1.5% lginte; it is hypothesized tht negtively chrged Eudrgit FS nd negtively chrged sodium lginte rejected ech other nd therefore the prticles size incresed. In ddition, microprticles prepred with Eudrgit FS were lrger thn those prepred with Eudrgit RS (Tle II). This might e due to the different chrge of the two polymers: positively chrged Eudrgit RS proly tends to ttrct negtively chrged enoxprin nd lginte, resulting in more compct orgniztion of the prticles nd thus to reduced size, while negtively chrged Eudrgit FS proly rejects negtively chrged enoxprin nd lginte, leding to looser structure, therefore lrger prticle size. Regrding the influence of the mixture of the two polymers (Eudrgit FS nd Eudrgit RS), the results were not sttisticlly significnt (Tle III), ut prticles consisting exclusively of one type of polymer were inferior in size ( µm) to those formulted y mixing the two polymers ( µm). In this cse, the difference in the level of encpsultion could e t the origin of heterogeneity in size of formultions consisting of the two polymers (Tle V). The formultion prepred with 100% Eudrgit FS (F9-5000) presented the lowest encpsultion efficiency (38%) nd dimeter of 108 µm, while the formultion prepred with 100% ERS showed the highest encpsultion efficiency (60%) nd dimeter of 79 µm. Normlly, the expected results would hve een decrese of the size of prticles with the ddition of ERS nd n increse of the encpsultion efficiency, ut the formultions prepred with 75%/25% nd then 50%/50% EFS/ERS rtios led to n increse of oth encpsultion efficiency (46% nd 50%, respectively) nd size (133 µm nd 244 µm, respectively). Finlly, incresing the volume of DCM from 2 ml 362

7 to 5 ml (Tle I) led to smller prticles due to the decrese of the viscosity of the polymeric orgnic solution. For exmple, microprticles prepred with Eudrgit FS 100% nd 2 ml DCM (F1-5000) were greter thn microprticles prepred with Eudrgit FS 100% nd 5 ml DCM (F9-5000). The sme cn e oserved for drug-free Eudrgit FS microprticles (F1-0 nd F9-0) nd for drug-free (F5-0 nd F12-0) nd drug-loded (F nd F ) Eudrgit RS 100% microprticles. A smll volume of orgnic solvent led to more viscous solution tht determined the formtion of lrger drops during emulsifiction, which ecme lrge microprticles, proly hving porous structure. Incresing the volume of orgnic solvent conducted to low viscosity solution, which formed smll drops tht ecme smll microprticles, proly with more compct orgniztion. encpsultion efficiency Heprins, eing compounds which re solule in wter, hve tendency to pss into the queous outer phse prior to the precipittion of the polymer, which lowers entrpment efficiency [13]. For this reson, n excipient, which is cple to retin enoxprin inside the microprticles, is highly desirle. An increse of the concentrtion of sodium lginte decresed the entrpment percentge y pproximtely 10% in the formultions prepred using no sodium lginte to the ones otined with 1.5% sodium lginte (p<0.05) (Tle IV). The intermedite vlues of 0.5% nd 1% hd no significnt influence. The type of polymer hd n importnt effect. Indeed, there ws more efficient encpsultion of the drug for Eudrgit RS (74-88%) thn for Eudrgit FS (39-47%) (Tle IV). These findings cn e explined sed on the structure of the two polymers nd on the dt otined y other reserchers. Hoffrt et l stted tht in the cse of Eudrgit RS, polyctionic quternry mmonium groups of the polymer, minly oriented towrds the continuous queous phse, my interct with nionic sulfte nd croxylte chins of enoxprin, which results in reduction of the migrtion of the emedded sustnce in the externl queous phse prior to the precipittion of the polymer [9]. The sme reserchers elieve tht the encpsultion of enoxprin cn occur y two mechnisms: encpsultion in the internl queous phse nd dsorption on the surfce of prticles through the electrosttic interctions mentioned ove. Similr percentges were otined for Eudrgit RS microprticles loded with other low moleculr weight heprins, tinzprin (72%) nd ndroprin (85%) [13]. On the other hnd, polynionic groups of Eudrgit FS rejected proly the negtively chrged chins of enoxprin, which tended to migrte towrds the externl queous phse prior to precipittion of the polymer, resulting in lower entrpment efficiency. Depending on the EFS/ERS rtio (Tle V), the lowest entrpment percentge of enoxprin ws oserved for microprticles contining 100% Eudrgit FS. An increse of the entrpment percentge of the drug in the presence of Eudrgit RS ws systemticlly oserved, s descried in the literture [9,13]. Regrding the decrese of orgnic solutions viscosity, the entrpment efficiency ws lowered with the reduction of the viscosity of the solution. The results could e correlted to the size of microprticles, so tht smller size led to lower percentge of encpsulted drug. Influence of the formultion fctors on in vitro drug relese The relese profiles of enoxprin were influenced y the mount of sodium lginte ( %) nd y the type nd rtio of the polymer used (Eudrgit RS or Eudrgit FS). Regrding the concentrtion of sodium lginte, the relese ws different depending on the type of polymer. For Eudrgit FS (Figure 2), the influence of lginte on relese ws not significnt, while for Eudrgit RS (Figure 2) it ws found tht incresing the mount of sodium lginte resulted in incresed percentges of enoxprin relesed. The formultion prepred with 1.5% lginte (F8-5000) significntly distnced from the other three, showing the highest percentges of enoxprin relesed (80% vs %). For this lst formultion, the interctions etween the ctive ingredient nd the polymer were proly disrupted y the presence of lrge quntities of sodium lginte, leding to incresed relese rte. Depending on the influence of polymers, it ws found tht enoxprin sodium relese fter 24 h from the microprticles prepred with Eudrgit RS ws incomplete (48-81%), compred with the percentges relesed from the microprticles prepred with Eudrgit FS (93-97%). Other studies hve lso reported n incomplete relese in the cse of Eudrgit RS, mking the ssumption tht certin polyscchride chins intercted strongly with Eudrgit RS, which resulted in incomplete relese [9,13]. As regrds Eudrgit FS, the presumed repulsions etween the polymer nd the drug determined the relese of the entire mount fter out 4-5 h. However, there ws no sttisticlly significnt influence of the type of polymer on the relese (p>0.05) etween formultions prepred with 1.5% sodium lginte, which could men tht high concentrtion of sodium lginte ccelerted the relese of enoxprin, irrespective of the type of polymer. Enoxprin relese profiles were lso influenced y the EFS/ERS rtios introduced in the formultion (Figure 3). First, there ws rpid relese from the formultion prepred with 50%/50% EFS/ERS (F ): 46% of the totl mount of enoxprin ws relesed fter 2 h, in HCl ph 1.2. This urst effect diminished with the increse of the mount of Eudrgit FS in the formultion: 20% fter 2 h for the formultion with 75%/25% EFS/ERS (F ) nd 3% fter 2 h for 100% EFS formultion (F9-5000). A 363

8 Phrmcy colonic drug delivery system must e cple of llowing mximum 10% drug relese in the gstric environment, therefore the formultion tht meets this requirements is the F formultion. Compring F nd F formultions, prepred with 2 ml DCM nd 5 ml DCM respectively, the relese of enoxprin decresed considerly for the F formultion. The microscope imges nd the size vlues lso showed n importnt size decrese for the prticles prepred with higher volume of DCM. The delyed relese for smller prticles ws proly due to more compct orgniztion of the polymer tht prevented fst drug relese, compred to prticles greter in size, which proly presented more porous structure, closer contct with the medium nd fcilitted the drug relese. Conclusions This study presented the development of enoxprin-loded polymeric microprticles using wter/ oil/wter doule emulsion technique followed y solvent evportion, nd the evlution of certin formultion fctors concentrtion of sodium lginte, type of polymers nd rtios of polymers on enoxprin prticles morphology, size, entrpment nd relese. Sodium lginte hd negtive influence on microprticles chrcteristics, i.e. incresingly concentrtions of lginte decresed drug encpsultion nd ccelerted the relese. The polymer type tht ensured higher encpsultion efficiency ws Eudrgit RS. The most importnt chievement ws tht the colon specific polymer, Eudrgit FS (100%), prevented the relese in the gstric simulted fluid. In conclusion, this study proved tht Eudrgit FS 30D is suitle polymer for the preprtion of colon-specific orl delivery systems of enoxprin. Acknowledgements The support of the trining progrms Ersmus LLP nd Ersmus STT is gretly cknowledged. 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