1、COLONIC TARGETING OF A PEPTIDE ANTI-BACTERIAL AGENTProf. Dr. Adel SakrIndustrial Pharmacy Graduate Program College of Pharmacy, University of CincinnatiCincinnati, Ohio, USACONGRESS CHAIR, FIP-INDUSTRIAL PHARMACY SECTIONDradelsakr1Prof. Dr. Adel SakrIntroduction NisinSignificance of colonic delivery

2、 of NisinMethods of colonic targetingEUDRAGIT L30D-55Wurster fluid bed coating2Prof. Dr. Adel SakrIntroductionPeroral drug delivery is the most convenient method of drug delivery.Peroral delivery of proteins and peptides presents a challenge.Targeting drugs to the colon is becoming of great interest

3、 for systemic absorption or local action.3Prof. Dr. Adel SakrNisinPeptide antimicrobial agentBelongs to the class of AntibioticsProduced by Lactococcus lactis Used as a food preservativeConsists of 34 amino acidsSolubility decreases with increasing pHSolubility decreases in the presence of salt4Prof

4、. Dr. Adel SakrNisinActive against many Gram-positive bacteria species.Reported activity against:Mycobacterium ActinomycesPenicillin-resistant Neisseria gonorrhoeaeHelibacter pyloriClostridium difficileVancomycin-resistant enterococci5Prof. Dr. Adel SakrNisin shows activity against both types of org

5、anisms.Nisin will be digested in the small intestine.Protection of Nisin until it reaches the colon is essential.Significance of Colonic Delivery of Nisin7Prof. Dr. Adel SakrMethods of Colonic TargetingChemical/Microbiological ApproachBacterial count in the colon is much higher than that of the smal

6、l intestine.Colonic bacteria is capable of extensive enzymatic activity.Colonic targeting is achieved by compounds selectively degraded by the microbial enzymatic activity (prodrugs or universal systems).Most of the materials are not approved for human use.8Prof. Dr. Adel SakrMethods of Colonic Targ

7、etingTechnological/Physiological ApproachUtilization of gastrointestinal pHIt was believed that gastrointestinal pH constantly rises from the stomach to the colon.Colonic targeting is accomplished by using polymer coatings having pH dependent solubility (soluble above pH 6-7).Good protection against

8、 gastric conditions, but unpredictable release in the intestine due to pH variation.10Prof. Dr. Adel SakrMethods of Colonic Targeting11Prof. Dr. Adel SakrMethods of Colonic Targeting12Prof. Dr. Adel SakrMethods of Colonic TargetingDeveloping a colonic delivery system with potential to succeed should

9、 be based on combining both pH and time dependent release mechanisms.14Prof. Dr. Adel SakrPolymethacrylate PolymersCommercially known as EudragitR.Generally are esters of polymethacrylic acid.Mainly used for coating of peroral dosage forms.Available as solid substance, organic solution, and aqueous

10、dispersion.15Prof. Dr. Adel SakrHypothesisIt is possible to target the delivery of Nisin to the Colon through the application of the appropriate polymers by WURSTER FLUID BED TECHNOLOGY17Prof. Dr. Adel SakrObjectiveTo formulate a TARGETED colonic delivery system for the antimicrobial agent Nisin whi

11、ch disintegrates in or near the proximal colon.18Prof. Dr. Adel SakrSpecific Aims1.To formulate Nisin core tablets having the following properties:a)Disintegrate in a reasonable time in phosphate buffer pH 6.8-7.0.b)Hard enough to withstand fluidization and coating in the Wurster fluidized bed.2.To

12、validate the tablet production process.19Prof. Dr. Adel SakrSpecific Aims3.To coat the core tablets using polymethacrylate polymers to resist gastric conditions for 2 hours.4.To study the effect of polymethacrylate coat thickness on the disintegration of the tablets in phosphate buffer pH 6.8.20Prof

13、. Dr. Adel SakrSpecific Aims5.To establish in-vivo feasibility of colonic delivery of Nisin in human volunteers by radiolabling and monitoring the behavior of the delivery system in the gastrointestinal tract using gamma scintigraphy.6.To study the effect of different storage conditions on the stabi

14、lity of the delivery system.21Prof. Dr. Adel SakrExperimental & ResultsCoating of core tabletsFeasibility of colonic delivery22Prof. Dr. Adel SakrFormula24Prof. Dr. Adel SakrX 3Mixing(Intragranular)Nisin+ Prosolv+ NaCl+ Ac-Di-solCompactionGranulation+TalcMixing(Intragranular)Mixing(Extragranular)+ P

15、rosolv+ NaCl + Ac-Di-Sol+ PVPMixing(Extragranular)+Mg StearateTableting1500 lbs25Prof. Dr. Adel SakrCoating Nisin Core TabletsCoating Formula:Eudragit L30D-5530% aqueous dispersionTriethyl citrate15% w/w of solid polymer contentTalc2% w/w of solid polymer contentWaterto dilute to a 20% aqueous dispe

16、rsion27Prof. Dr. Adel SakrPreparation of the Coating DispersionWater was added to the Eudragit L30D-55 aqueous dispersion.Triethyl citrate was added slowly to the dispersion drop-wise while stirring.The dispersion was allowed to stir for two hours.Talc was added gradually and allowed to completely d

17、isperse for 30 minutes.28Prof. Dr. Adel SakrCoating Process100 g of core tablets were placed in the Wurster fluid bed and preheated to 30oC.The tablets were fluidized.Coating dispersion was sprayed using 1.0 bar atomization pressure.The process was stopped after reaching the desired coat thickness.2

18、9Prof. Dr. Adel SakrWurster Chamber30Prof. Dr. Adel SakrBottom spray fluidized-bedModified from Glatt brochureFluidization airPartition chamberAir distribution plate31Prof. Dr. Adel SakrExperimental & MethodologyTablet EvaluationEvaluation of coated tablets:Coat thickness / coat weight determination

19、Resistance to gastric conditionsCoat thickness and tablet disintegration in pH 6.832Prof. Dr. Adel SakrResultsThe core tablets withstood coating in the Wurster fluidized bed.The coated tablets resisted disintegration and no Nisin release was detected in acidic medium.A linear relationship was observ

20、ed between onset of tablet disintegration and coat thickness/weight in pH 6.833Prof. Dr. Adel Sakr34Prof. Dr. Adel SakrExperimental & MethodologyFeasibility of Colonic DeliverySamarium oxide was incorporated intragranularly in the core tablets.Tablets with three different coat thicknesses (40%, 120%

21、, and 188% w/w) were chosen for in-vivo testing in humans.The coated tablets were neutron activated, to emmit gamma rays.35Prof. Dr. Adel SakrExperimental & MethodologyFeasibility of Colonic DeliveryThe delivery systems were administered to 12 healthy human subjects:Delivery system - B (188% coat):2

22、 subjectsDelivery system - A (120% coat):4 subjectsDelivery system - C (40% coat):6 subjectsThe dosage forms were monitored by gamma Scintigraphy.36Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 002-DSD; Date: June 8, 1997; Treatment: A0.5 hrs0.0 hrs1.5 hrs3.0 hrs3.5 hrs5.0 hrsStom

23、achJejunumIleum37Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 002-DSD; Date: June 8, 1997; Treatment: A12.0 hrs5.5 hrs16.0 hrs38Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 001-TLB; Date: June 8, 1997; Treatment: B0.5 hrs0.0 hrs1.5 hrs3.0 hrs4.5 hrs5.0 hrsStom

24、achJejunumIleum39Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 001-TLB; Date: June 8, 1997; Treatment:B8.0 hrs7.0 hrs9.0 hrs10.0 hrs16.0 hrs24.0 hrsStomachJejunumIleumColon40Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 004-TLC; Date: June 8, 1997; Treatment: B0

25、.5 hrs0.0 hrs1.5 hrs2.5 hrs4.5 hrs5.0 hrsStomachJejunumIleumColon41Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 004-TLC; Date: June 8, 1997; Treatment: B7.0 Hr12.0 Hr42Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 007-R_S; Date: June 14, 1997; Treatment: B2.5 h

26、rs3.0 hrs3.5 hrs1.0 hrs0.0 hrs1.5 hrsStomachJejunumIleum43Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 007-R_S; Date: June 14, 1997; Treatment: B10.0 hrs12.0 hrs6.0 hrs4.0 hrs8.0 hrsStomachColon44Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 010-JBB; Date: June

27、 14, 1997; Treatment: B2.5 hrs3.5 hrs5.0 hrs0.5 hrs0.0 hrs1.5 hrsStomachJejunumIleum45Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 010-JBB; Date: June 14, 1997; Treatment: B10.0 hrs12.0 hrs13.0 hrs7.0 hrs5.5 hrs9.0 hrsStomachColon46Prof. Dr. Adel SakrStomachJejunumIleumDelivery o

28、f Nisin to the Colon2.5 hrs0.0 hr3.5 hrs4 hrs4.5 hrs5 hrsSubject number: 003-EDD; Date: June 8, 1997; Treatment:CStomachJejunumIleum47Prof. Dr. Adel Sakr5.5 hrs6 hrs7 hrsDelivery of Nisin to the ColonSubject number: 003-EDD; Date: June 8, 1997; Treatment: C48Prof. Dr. Adel SakrDelivery of Nisin to t

29、he ColonSubject number: 011-SCK; Date: June 14, 1997; Treatment: C2.0 hrs3.0 hrs3.5 hrs1.0 hrs0.5 hrs1.5 hrsStomachJejunumIleum49Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 011-SCK; Date: June 14, 1997; Treatment: C7.5 hrs11.0 hrs14.0 hrs5.0 hrs4.5 hrs5.5 hrsStomachJejunumIleumC

30、olon50Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 009-SDT; Date: June 14, 1997; Treatment: C2.0 hrs2.5 hrs3.5 hrs0.5 hrs0.0 hrs1.5 hrsStomachJejunumIleum51Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 009-SDT; Date: June 14, 1997; Treatment: C8.0 hrs10.0 hrs12

31、.0 hrs5.0 hrs4.0 hrs7.0 hrsStomachColon52Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 012-LBS; Date: June 14, 1997; Treatment: C3.0 hrs3.5 hrs4.0 hrs1.5 hrs0.5 hrs2.5 hrsStomachJejunumIleum53Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 012-LBS; Date: June 14, 1997; Treatment: C6.5 hrs5.0 hrs4.5 hrs5.5 hrsStomachColonJejunumIleum54Prof. Dr. Adel SakrDelivery of Nisin to the ColonSubject number: 006-SHA; Date: June 8, 1997; Treatment: C1.0 hrs0.5 hrs1.5 hrs2.0 hrs2.5 hrs3.0 hrsStomac