Study Of PLGA Microspheres For Sustained Release Of Triptorelin Acetate

During the past two decades, interest in developing microparticles withthe ability of a continuous drug release over a long period of time as modernparenteral drug delivery systems has obviously grown. Considerable interesthas been focused on the use of biodegradable polymers for specializedapplications such as controlled release of drug formulations. Amongst thedifferent classes of biodegradable polymers, the thermoplastic aliphatic poly(lactide) (PLA), poly (glycolide) (PGA), and especially the copolymer oflactide and glycolide, poly (lactide-co -glycolide) (PLGA) have generatedimmense interest due to their favorable properties such as goodbiocompatibility, biodegradability, and mechanical strength. They are syntheticin origin and are degraded in vivo to produce biocompatible, toxicologicallysafe by-products which are further eliminated by the normal metabolicpathways. Also, they have been approved by the Food and DrugAdministration (FDA) for drug delivery.Recent the encapsulation of highly water soluble compounds includingproteins and peptides have been extensively investigated. The long-term depotmicrospheres parenterals of luteinizing hormone-releasing hormone (LHRH)and its analogs are the most successful products among the controlled releasepreparation. Triptorelin is a decapeptide analog of LHRH and has almost 100times the biological activity of that. Its once-a-month injectable depotformulation is already available on the world market. Its trade name isDecapeptyl. This depot formulation can provide a fairly constant release of thepeptide over one month in animals and humans after s.c. or i.m. injection.Therefore, this preparation shows reliable efficacy for treatment of patientswith advanced prostate cancer, endometriosis, precocious puberty, and otherhormone-dependent diseases through persistent chemical castration withoutany need for repeated daily injection, and thus it has markedly improvedpatient compliance.We wished to prepare a controlled-release dosage form that would releasetriptorelin acetate at a constant rate for one month which is modelled onDecapeptyl. The purpose of this study is looking for a kind of simple andconvenient, high-efficient microsphere preparing technology, exploring eachparameter's influence disciplinarian on the products, put up a technologicalplatform for the microsphere preparation of the polypeptide and proteinmedicine.The purpose of the present study was to find a suitable technique for adosage form for controlled release of triptorelin acetate for one month byrelease of the drug. The microencapsulation method employed must includethe following requirements: (1) The stability and biological activity of the drugshould not be adversely affected during the encapsulation process or in thefinal microsphere product. (2) The yield of the microspheres having therequired size range (upto 250 μm, ideally<125μm) and the drug encapsulationefficiency should be high. (3) The microsphere quality and the drug releaseprofile should be reproducible within specified limits. (4) The microspheresshould be produced as a free flowing powder and should not exhibitaggregation or adherence. Injectable microspheres can be prepared by severaltechniques, with organic phase separation and solvent evaporation methodsbeing used most frequently.Acording to the depot of Decapeptyl SR (3.75mg), the most suitablepolymer was a PLGA with a weight average molecular weight of 43000 and amolar ratio of lactic to glycolic acid of 50/50. The molecular weight wasdetermined by gel-permeation chromatography (GPC), and the lactic/glycolic(LA/GA) copolymer ratio of PLGA was obtained from the integrated signalratio between the methyl moiety of lactic acid and the methylene moiety ofglycolic acid in the NMR spectrum. Acording to the results of the testing andanalyzing we investigate the microsphere preparing technology.We initially selected the phase separation method (coacervation). Thisprocess consists of decreasing the solubility of the encapsulating polymer byaddition of a third component to the polymer solution in an organic solution.At a particular point, the process yields two liquid phases (phase separation):the polymer containing coacervate phase and the supernatant phase depleted inpolymer. The drug which is dispersed / dissolved in the polymer solution iscoated by the coacervate.First we investigate the addition methods of the coacervationreagent(silicone oil). Only when the silicone oil is added droplets could thepolymer be extracted slowly, so that the polymer had sufficient time to depositand coat evenly on the drug particle surface during the coacervation process,and then could we obtain free flowing microspheres. The concentration of boththe drug and the polymer used are important as well. When the otherpreparation parameters are unchangeable the higher concentration of thepolymer, the higher encapsulation efficiency of the drug, and the lower yieldof the drug by reason of the aggregation of the polymer. Acording to ourresearch work, we can prepare microspheres using the phase separationmethods with high encapsulation efficiency of the drug. The microspheres hasa porous and rough surface, which cause a bulk burst of drug. Thecoacervation method tends to produce particles which are agglomerated, thereis difficulty in mass production, the method requires large quantities of organicsolvent, and it is difficult to remove residual solvents from the finalmicrosphere product.Double emulsion process is a water-in-oil-in-water (W/O/W) method andis best suited to encapsulated water-soluble drugs like triptorelin acetate. Abuffered or plain aqueous solution of the drug is added to an organic phaseconsisting of PLGA solution in DCM with vigorous stirring to form the firstmicrofine W/O emulsion. This emulsion is added gently with stirring into alarge-volume water containing an emulsifier like PVA to form the W/O/Wemulsion. The emulsion is then subjected to solvent removal by evaporationprocess. The solid microspheres are then washed and collected bycentrifugation. These are lyophilized to give the final free flowing microsphereproduct.The formation of the microspheres is affected by a number of factors. Westudy the main variables that influence the microencapsulation process and thefinal microsphere product using the orthogonal experimental design software:(a) the volume and the additions (salt and gelatin) of the drug solution; (b) thevolume of the organic solvent; (c) the continuous phase composition andvolume; (d) the ultrasonic power in forming the first emulsion; (e) theagitation speed of the second emulsion process. The experimental resultsshowed that we can abtain free-flowing microsphere with high encapsulationefficiency following those proper conditions below: (1) small drug solution'svolume, without any additions; (2) appropriate volume of the organic solvent;(3) high concentration of the emulsifier used, adding 2.5mg/ml NaCl, withappropriate volume. This method was selected because the preparationprocedure is simple, the materials for parenterals are easily available and feworganic solvents with a lower boiling point are used to overcome the problemsassociated with residual solvent. Because of those advantages we selected thismethod to prepare microsphere of triptorelin in the end.We confirme the preparation parameters of the microsphere's preparationvia the small amount of experimentation above. According to the preparationparameter of the bench scale we enlarge the preparation quantity of the microsphereto five times of that. Finally we prepare microsphere of triptorelin using thedouble emulsion process in mass production. The microsphere is spherical andsmooth, with a high drug encapsulation efficiency of 73.5%, yield of 88.3%,and in the size range of 10～50μm. The in vitro release rate of the drug fromthe microsphere of this product was determined by HPLC after extractionusing DCM-water solution. First it has a rapid release about 20% in 1 hour,followed by a gradually release of triptorelin over 4 weeks, the amount of drugremaining in the mirosphere after 4 weeks was 80% of the peptide for thisexamination. Those results accord with the demands of the controlled-releasepreparation.