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Formulation development and evaluation of colonic drug delivery system of budesonide microspheres by using spray drying technique
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/jopr
Original Article
Formulation development and evaluation of colonic drug delivery system of budesonide microspheres by using spray drying technique Raosaheb S. Shendge a,*, Fahim J. Sayyad b a
Lecturer, Department of Pharmaceutics, Sanjivani College of Pharmaceutical Education and Research, Kopargaon 423603, Maharashtra, India b Department of Pharmaceutics, Government College of Pharmacy, Karad, Satara, India
article info
abstract
Article history:
Aim: Present work mainly deals with the formulation of colon targeting dosage form of
Received 6 April 2013
budesonide, using natural polysaccharides as polymer by utilizing spray drying technique.
Accepted 17 April 2013
Budesonide is locally acting corticosteroid utilized for the treatment of inflammatory bowel
Available online 4 May 2013
disease. Budesonide is having high glucocorticoid and low mineralocorticoid effect. Methods: Chemical modification of chitosan was carried out by using glutaraldehyde which
Keywords:
causes crosslinking between chitosan molecules rendering them water insoluble. Drug,
Budesonide
polymer and crosslinking agent were spray dried to form microparticles. 2 ml of 25%
Chitosan
glutaraldehyde was found to be necessary for required crosslinking of 1% chitosan in acetic
Controlled release pellets
acid.
Glutaraldehyde
Results: Formed microparticles showed 20% drug release in initial 5 h and 80% drug release
Spray drying technique
in rat fecal content media which also suggested that specific degradability of chitosan by colonic bacteria remains after crosslinking also. Formed microparticles were then evaluated for various parameters like morphology, particle size distribution, % yield, % entrapment, % of loading and drug release. Conclusion: Spray drying produces narrow and small size microparticles of budesonide. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
Spray drying as one of the method of drying is highly utilized and acceptable method of drying and gained lot attention in past couple of decades. Spray drying is defined as atomization of solution of one or more solids via nozzle, spinning disc or other device followed by evaporation of solvent to obtain dried
particles. Choosing optimum parameter such as inlet temperature, outlet temperature, feed transfer rate, atomization rate and D-block on and off for spray drying is difficult and most important step in whole operation. Once these parameters are optimized for particular type of product, spray drying becomes easy.1e5 Budesonide is a glucocorticoid steroid for the treatment of Crohn’s disease (inflammatory bowel
* Corresponding author. Tel.: þ91 9890614868; fax: þ91 (0) 2423 222682. E-mail addresses: [email protected], [email protected] (R.S. Shendge). 0974-6943/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jopr.2013.04.016
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j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
disease). Budesonide has a high first-pass metabolism. Budesonide has a lower incidence of systemic manifestations than similar medications.6e8 Targeted drug delivery into the colon is highly desirable for local treatment of a variety of bowel diseases such as ulcerative colitis, Crohn’s disease, amebiasis, colonic cancer, local treatment of colonic pathologies, and systemic delivery of protein and peptide drugs. The colon specific drug delivery system (CDDS) should be capable of protecting the drug. CDDS protects peptide drugs from hydrolysis, and enzymatic degradation in duodenum and jejunum, as pH of jejunum is acidic and Eudragit RLPO is acid resistant and time controlled release and eventually releases the drug into ileum or colon which leads to greater systemic bioavailability. Because the colon has a long residence time which is up to 5 days and is highly responsive to absorption enhancers.9e15
2.
Materials and methods
Budesonide was obtained from Glenmark Pharmaceuticals Ltd., Nasik. Pectin, chitosan and other materials used were of AR Grade and were obtained from Loba Chemie.
2.1.
Table 1 e Parameters for spray drying of chitosan for trial 1 to 5. Parameter
Set value
Inlet temperature Outlet temperature Feed pump rate Aspiration rate Atomization pressure
2.3.2.
Stability of microparticles in media
100 mg of microparticles were kept in 100 ml of 0.1 N HCl at 50 rpm on mechanical shaker and observed for solubilization, if any, of microparticles.
2.3.3.
Entrapment efficiency
100 mg of microparticles were weighed and dispersed into 20 ml of ethanol in a beaker and the beaker was wrapped with aluminum foil. Microparticles were then digested for 24 h in the darkness and then sonicated for 1 h. Sonicated sample was then filtered by using Whatman filter paper. Filtered sample was then analyzed by using UV spectrophotometer after suitable dilution. From the reading, by using following formula % of entrapment was calculated.
Spray drying of chitosan with crosslinker % of entrapment ¼
Various crosslinking agents are utilized for crosslinking purpose like glutaraldehyde, genepin, formaldehyde. Crosslinking occurs in between chitosan molecules retarding their water solubility. 25% Glutaraldehyde is utilized for crosslinking of chitosan while spray drying.16e18
2.2.
Trial 1
1 g of chitosan was dissolved in 100 ml 5% dilute acetic acid solution. In it 25 ml of 25% of glutaraldehyde was added. Allowed to crosslink for 15 min. After 15 min very thick gel was formed such that it can’t be passed through the spray drying system. So it was started with 1 ml of glutaraldehyde.
2.3.
Trial 2
1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 1 ml of 25% glutaraldehyde was added and allowed to crosslink for 15 min while stirring. Above solution was kept for stirring and spray dried at conditions given in Table 1. Obtained product was collected, weighed and evaluated for following parameters.
2.3.1.
% of yield
Obtained product was weighed and % of yield was calculated by using following formula: % of yield ¼
Amount of product obtained 100 Amount of total solid in spray drying solution
150 C 100 C 1 ml per min 40 Nm3/h 1 bar
2.3.4.
Practical drug content 100 Theoretical drug content
% Drug loading
% of drug loading was calculated to find out % of amount of drug present in given weight of microspheres. % of drug loading was calculated by using following formula: % of loading ¼
2.3.5.
Drug content 100 Weight of microspheres
Drug release
Drug release was checked for 5 h by using USP paddle apparatus. 900 ml of 0.1 N HCl was utilized as a media. Microparticles were weighed such that it becomes equivalent to 9 mg of budesonide. Then microparticles were filled into size 4 capsule. Capsule was then placed into media at 50 rpm and 37 0.5 C. 5 ml sample was withdrawn at each 1 h and analyzed by UV. If required suitable dilutions were prepared. Dissolution was carried out for 5 h only to check drug release occurring in critical period.19,20 Graph was plotted as % of drug release versus time.
2.4.
Trial 3
In this trial amount of crosslinker (glutaraldehyde) was increased upto 2 ml to increase crosslinking and ultimately to decrease drug release. 1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2 ml of 25% glutaraldehyde was added and allowed to react for 15 min. Above solution was kept for stirring and spray dried at conditions mentioned in Table 1. Outlet temperature was varied between 100 and 60 C.
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Obtained product was collected and weighed. % Yield was calculated. Microparticles were again evaluated for all the above mentioned parameters.
2.7.2.
2.5.
2.7.3.
Trial 4
In this trial again amount of crosslinker was increased.1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 3 ml of 25% glutaraldehyde was added and allowed to react for 15 min. After 15 min change in gel was observed and a very thick jelly like mass was obtained which was not at all passable through spray drying system.
2.6.
Trial 5
Amount of chitosan is increased and in proportion with chitosan amount of glutaraldehyde was also increased. 1.2 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2.4 ml of 25% glutaraldehyde was added and allowed to react for 15 min. Above solution was kept for stirring and dried at conditions given in Table 1. After starting of spray drying when near about 30 ml feed was remained, it got gelled and was unable to pass through spray drying system. So trial was stopped there.
2.7.
DSC
DSC of the microparticles was carried out to find interaction, if any, in between chitosan, glutaraldehyde and drug. DSC was carried out at Diya Labs, Mumbai. Sample was sealed into aluminum pan with lid pierced. Heating range was 10 K/min. with nitrogen purging at 60 ml/min.
2.7.4.
FTIR
FTIR was recorded on Bruker alpha. Microparticles were mixed with moisture free KBr in 1:100 ratio. Pressed into thin disc by using KBr press and FTIR of the disc was recorded from 500 cm1 to 4000 cm1.
2.7.5.
Particle size analysis
Particle size analysis was performed by using laser diffraction particle size analyzer (Malvern Mastersizer, UK). About 500 mg of microcapsules were weighed and suspended in 500 ml benzene, ultrasonicated for 2 min to form uniform dispersion and analyzed for particle size.
3.
Results and discussion
3.1.
Spray drying of chitosan with crosslinker
Glutaraldehyde was utilized for the crosslinking purpose of chitosan.
Trial 3A
Trial 3 was again conducted to check the effect of outlet temperature on product yield. In previous trial outlet temperature was varying between 100 and 60 C, but this time outlet temperature was varied between 100 and 90 C. Product was collected and weighed and evaluated further for the following parameters.
2.7.1.
Scanning electron microscopy
Scanning electron microscopy (SEM) was carried out at Diya labs, Mumbai.
Drug release for 24 h
Dissolution study was carried out for 24 h in USP type 2 apparatus (Paddle) in triplicate manner. Initial 2 h drug release was checked in simulated gastric fluid, then for next 3 h pH of the media was increased upto 6.8 by adding 1 M NaOH and addition of 10 g of pancreatin was done and after 5 h pH of the media was increased upto 7.4 and addition of rat cecal content was done into simulated colonic environment. Dissolution study was carried out in triplicate manner. Graph was plotted as % of drug release versus time.
3.2.
Trial 1
Initially 1 g of chitosan was dissolved in 100 ml of 5% dilute acetic acid solution. In it 25 ml of 25% of glutaraldehyde was added. Allowed to crosslink for 15 min. After 15 min very thick gel was formed such that it can’t be passed through the spray drying system. This may be happened due to excess of glutaraldehyde. Excess of glutaraldehyde causes dense network formation between the molecules of chitosan which results in formation of very thick gel.
3.3.
Trial 2
In next trial amount of glutaraldehyde was reduced to minimum level. In trial 2, 1 ml of glutaraldehyde was utilized for crosslinking purpose. After 15 min of crosslinking no gel formation occurs and solution remains in a condition such that it
Table 2 e Result of evaluation test of various trials of chitosan with crosslinker. Sr. no
1 2 3
Parameter
% Yield % Entrapment % Drug loading
Result Trial 1
Trial 2
Trial 3
Trial 4
Trial 5
Trial 3A
e e e
14.10% 91.23% 45.5%
16.20% 92.28% 46.14%
e e e
11.12% 84.24% 42.12%
24.56% 92.21% 46.15%
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
120
preceding 4 h only 25% of drug release was occurred as shown in Fig. 1.
% Drug Release
100
3.4.
80
Trial 3
Trial 02
60
Trial 03
40
Trial 05 Trial 3A
20 0
459
0
10
20 Time (Hours)
30
Fig. 1 e Drug release profile of various trials.
can be passed through the spray drying system. Spray drying parameters were chosen by considering water as a solvent. When addition of ethanolic solution of drug was added to chitosan solution, precipitation of drug was occurred in very fine particles. Now in this case polymer is in solubilized state and drug is insoluble. So in this case microparticles may be of microcapsule type, embedding drug molecules inside the polymer coat. After spray drying microparticles were weighed, % yield was calculated and checked for integrity purpose. 100 mg of microparticles were kept in 100 ml of 0.1 N HCl at 50 rpm on mechanical shaker and observed for 24 h. After 24 h of shaking, microparticles were found to be as it is. No solubilization of microparticles was occurred, so microparticles were evaluated for further parameters. Further evaluation was carried out for % of drug entrapment, % of drug loading and drug release and results were as shown in Table 2. In 5 h near about 35% of drug release occurred in acidic media which indicated that crosslinking was occurred but not in the amount which was required. Amount of crosslinker was not enough. So in next trial amount of crosslinker was increased in order to increase crosslinking. In initial 1 h about 10% of drug release was occurred which may be indication of presence of drug on the surface of microparticles which is going into media immediately after addition. After 1 h drug release is in sustain manner. In
In this trial amount of crosslinker was increased upto 2 ml. 1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol, dissolved and added to the chitosan solution. After proper mixing 2 ml of 25% glutaraldehyde was added and allowed to react for 15 min and spray dried. Evaluation of product was carried out as per previous batch. Noticeable change was not observed in drug content which suggested that there is no considerable impact of crosslinking agent on the drug content. Drug release was calculated for 5 h and found to be 19% after 5 h as shown in Fig. 2. Result in decrease in drug release was noticed due to increased amount of crosslinking which is caused by increased amount of glutaraldehyde. There are more number of glutaraldehyde molecules present for interchain crosslinking of amino groups of adjacent chitosan molecules. As the number of bridges between two chitosan chains increased, stiffness of chitosan molecules also increased resulting in uptake of lesser amount of water and less swellability and solubility.
3.5.
Trial 4
In this trial amount of crosslinker was increased upto 3 ml. Preparation of feed was done in same manner as that of previous batches. Crosslinking time was also kept 15 min. But due to increased amount of crosslinker thick gel was obtained after 15 min which was not passable through spray drying system. Gel formation occurred due to excess amount of glutaraldehyde. So instead of increasing crosslinking agent to 3 ml, both chitosan and glutaraldehyde were increased in proportion wise manner by taking into consideration 2 ml of glutaraldehyde for crosslinking of 1 g of chitosan.
3.6.
Trial 5
In this trial amount of chitosan and glutaraldehyde was increased in proportion wise manner. 1.2 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of
Fig. 2 e SEM image results of trial 3A.
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budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2.4 ml of 25% glutaraldehyde was added and allowed to react for 15 min. After 15 min no thick gel formation occurred so spray drying was started. When near about 30 ml of feed was remained thick gel formation occurred which was not able to pass through spray drying system. So spray drying was stopped, product was collected and evaluated. After 5 h 25% of drug release occurred as shown in Fig. 1, which was not desirable. This may be happened due to gelling of remaining 30 ml of feed, failing it to be spray dried. From the above trials it was concluded that 2 ml of 25% of glutaraldehyde is maximum amount which can be utilized for crosslinking purpose of 1 g chitosan having degree of deacetylation 70e90% in 5% acetic acid solution without formation of thick gel which can be passed through nozzle of spray dryer by taking 15 min as a crosslinking time.
3.7.
Trial 3A
Trial 3A was conducted to find out the effect of temperature variation on % of yield. In this trial outlet temperature was varied between 100 and 90 C. In previous trial outlet temperature was varying between 100 and 60 C. % of yield obtained in this trial is more as compared to batch 3. This may be happened due to increase in drying rate due to maintaining temperature in the range of boiling point of the solvent. Evaluation of batch 3A was carried out.
3.7.1.
Dissolution study
Dissolution study was carried out 24 h. Graph of % of drug release versus time was plotted as follows. In initial 5 h 20% of drug release has occurred. In initial 1 h 10% drug release was obtained. Once addition of pancreatin was done, drug release increased slightly. After 5 h 20% of drug release was occurred. But once addition of rat cecal content is carried out drug release increased rapidly. In next 2 h 97% of drug was released. Results were shown in Fig. 1. This indicates that after
Fig. 4 e DSC spectra of budesonide microparticles.
crosslinking of chitosan, it retains its specific biodegradability by colonic micro-organisms.19
3.7.2.
Scanning electron microscopy
Scanning electron microscopy was carried out to find out morphology of microparticles. Results of SEM are as shown in Fig. 2. SEM images indicate morphology of microparticles which was smooth in appearance and spherical in shape and having size less than 5 mm. Small size may be contributed to the microparticles due to apparatuses size of atomizer high atomization pressure during spray drying. Surface of the microparticles is smooth without any grooves which indicate that coating has occurred uniformly.
3.7.3.
DSC
DSC of the microparticles was carried out to check possible interaction in between drug and polymer. DSC graph showed endothermic peak near 160 C which is indication of presence of drug. In DSC graph of pure budesonide endothermic peak was also observed at 160 C as shown in Fig. 4.
3.7.4.
FTIR of microparticles
FTIR spectra of microparticles was recorded by using Bruker alpha. Microparticles showed the presence of particular groups
Fig. 3 e FTIR spectra of budesonide microparticles.
Fig. 5 e Particle size distribution of trial 3A.
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
which are present in FTIR spectra of budesonide as shown in Fig. 3.
3.7.5.
Particle size analysis
Particle size analysis was performed on Malvern Mastersizer. Maximum particle size was found be distributed in the range of 2e5 mm. Results were shown in Fig. 5. Less particle size is obtained which may be contributed to the method of microsphere preparation which is spray drying. Other methods such as solvent evaporation, emulsion method generates particles of higher size.
Conflicts of interest All authors have none to declare.
references
1. O’Connor E, Schwartz J, Felton L. Powders. In: Troy David B, ed. Remington: The Science and Practice of Pharmacy. 21st ed.; 2006:702. 2. C¸elik M, Wendel S. Spray drying and pharmaceutical applications. In: Parikh M, ed. Handbook of Pharmaceutical Granulation Technology. 2nd ed.; 2005:129e155. 3. Bete Spray Dry Manual. BETE Fog Nozzle, Inc. BETE. http://www. bete.com/pdfs/BETE_SprayDryManual.pdf. 2005. Accessed 3.04.12. 4. A Primer on Spray Drying. GEN Nitro. http://www.niro.com/ niro/cmsresources.nsf/filenames/A-primer-on-Spray-DryingChemical-Engineering-Nov09.pdf/$file/A-primer-on-SprayDrying-Chemical-Engineering-Nov09.pdf. November 2009. Accessed 3.04.12. 5. Rankell A, Liberman H, Schiffmann R. Dying. In: Lachman L, Liberman H, eds. The Theory and Practice of Industrial Pharmacy. Bangalore: CBS Publishers And Distributors, Indian Edition; 2009:60e62. 6. Aggarwal Samvedna, Sharma Shalini, Lal Sukhbir, Choudhary Neeraj. Recent trends in colon targeted drug delivery system. RJPBCS. OctobereDecember 2011;2(4):406e411.
461
7. Fedorak RN, Bistritz L. Targeted delivery, safety, and efficacy of oral enteric-coated formulations of budesonide. Adv Drug Deliv Rev. 2005;57(2):303e316. 8. Chourasia MK, Jain SK. Pharmaceutical approaches to colon targeted drug delivery systems. Eur J Pharm Sci. 2003;6(1):32e66. 9. Yang L, Chu JS, Fix JA. Colon-specific drug delivery: new approaches and in vitro/in vivo evaluation. Int J Pharm. 2002;235:1e15. 10. Patel Mayur M, Patel Santnu L, Bhadani Manish N, Shah Tejal J, Amin Avani F. A synchronous colon-specific drug delivery system for orally administered mesalamine. Acta Pharmaceutica Sciencia. 2009;51:251e260. 11. Chickpetty M, Baswaraj Raga, Nanjwade BK. Studies on development of novel combined time and pH dependent solventless compression coated delivery systems for colonic delivery of diclofenac sodium. Asian J Pharm Clin Res. 2010;3(2):410e416. 12. Wasnik Shailendra, Parmar Poonam. The design of colonspecific drug delivery system and different approaches to treat colon disease. Int J Pharm Sci Rev Res. 2011;6(2):031. 13. Vinay Kumar KV, Sivakumar T, Tamizh Mani T. Colon targeting drug delivery system: a review on recent approaches. Int J Pharm Biomed Sci. 2011;2(1):11e19. 14. Krishnamachari Y, Madan P, Lin S. Development of pH- and time-dependent oral microparticles to optimize budesonide delivery to ileum and colon. Int J Pharm. 2007;338(1e2): 238e247. 15. Postma DS, Kerstjens HAM. Are inhaled glucocorticosteroids effective in chronic obstructive pulmonary disease? Am J Respir Care Med. 1999;160:566e571. 16. Hennink WE, Nostrum CF. Novel crosslinking method to design hydrogels. Adv Drug Deliv Rev. 2002;54:13e36. 17. Nunthanid J, Luangtana-anan M, Sriamornsak P, et al. Use of spray dried chitosan acetate and ethyl cellulose as compression coats for colonic drug delivery: effect of swelling on triggering in vitro drug release. Eur J Pharm Biopharm. 2009;71:356e361. 18. Aulton ME. Pharmaceutics e The Science of Dosage Form Design. 2nd ed. Churchil Livingstone, International Ed; 1988:133e137. 19. United States Pharmacopeia20-NF25. vol. III. The National Compendia of Standards; 2009:1774e1775. 286. 20. Indian Pharmacopoeia. vol. 84. Ghaziabad: The Indian Pharmacopoeia Commission; 2010:193. 928e929.
Available online at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/jopr
Original Article
Formulation development and evaluation of colonic drug delivery system of budesonide microspheres by using spray drying technique Raosaheb S. Shendge a,*, Fahim J. Sayyad b a
Lecturer, Department of Pharmaceutics, Sanjivani College of Pharmaceutical Education and Research, Kopargaon 423603, Maharashtra, India b Department of Pharmaceutics, Government College of Pharmacy, Karad, Satara, India
article info
abstract
Article history:
Aim: Present work mainly deals with the formulation of colon targeting dosage form of
Received 6 April 2013
budesonide, using natural polysaccharides as polymer by utilizing spray drying technique.
Accepted 17 April 2013
Budesonide is locally acting corticosteroid utilized for the treatment of inflammatory bowel
Available online 4 May 2013
disease. Budesonide is having high glucocorticoid and low mineralocorticoid effect. Methods: Chemical modification of chitosan was carried out by using glutaraldehyde which
Keywords:
causes crosslinking between chitosan molecules rendering them water insoluble. Drug,
Budesonide
polymer and crosslinking agent were spray dried to form microparticles. 2 ml of 25%
Chitosan
glutaraldehyde was found to be necessary for required crosslinking of 1% chitosan in acetic
Controlled release pellets
acid.
Glutaraldehyde
Results: Formed microparticles showed 20% drug release in initial 5 h and 80% drug release
Spray drying technique
in rat fecal content media which also suggested that specific degradability of chitosan by colonic bacteria remains after crosslinking also. Formed microparticles were then evaluated for various parameters like morphology, particle size distribution, % yield, % entrapment, % of loading and drug release. Conclusion: Spray drying produces narrow and small size microparticles of budesonide. Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
1.
Introduction
Spray drying as one of the method of drying is highly utilized and acceptable method of drying and gained lot attention in past couple of decades. Spray drying is defined as atomization of solution of one or more solids via nozzle, spinning disc or other device followed by evaporation of solvent to obtain dried
particles. Choosing optimum parameter such as inlet temperature, outlet temperature, feed transfer rate, atomization rate and D-block on and off for spray drying is difficult and most important step in whole operation. Once these parameters are optimized for particular type of product, spray drying becomes easy.1e5 Budesonide is a glucocorticoid steroid for the treatment of Crohn’s disease (inflammatory bowel
* Corresponding author. Tel.: þ91 9890614868; fax: þ91 (0) 2423 222682. E-mail addresses: [email protected], [email protected] (R.S. Shendge). 0974-6943/$ e see front matter Copyright ª 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jopr.2013.04.016
457
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
disease). Budesonide has a high first-pass metabolism. Budesonide has a lower incidence of systemic manifestations than similar medications.6e8 Targeted drug delivery into the colon is highly desirable for local treatment of a variety of bowel diseases such as ulcerative colitis, Crohn’s disease, amebiasis, colonic cancer, local treatment of colonic pathologies, and systemic delivery of protein and peptide drugs. The colon specific drug delivery system (CDDS) should be capable of protecting the drug. CDDS protects peptide drugs from hydrolysis, and enzymatic degradation in duodenum and jejunum, as pH of jejunum is acidic and Eudragit RLPO is acid resistant and time controlled release and eventually releases the drug into ileum or colon which leads to greater systemic bioavailability. Because the colon has a long residence time which is up to 5 days and is highly responsive to absorption enhancers.9e15
2.
Materials and methods
Budesonide was obtained from Glenmark Pharmaceuticals Ltd., Nasik. Pectin, chitosan and other materials used were of AR Grade and were obtained from Loba Chemie.
2.1.
Table 1 e Parameters for spray drying of chitosan for trial 1 to 5. Parameter
Set value
Inlet temperature Outlet temperature Feed pump rate Aspiration rate Atomization pressure
2.3.2.
Stability of microparticles in media
100 mg of microparticles were kept in 100 ml of 0.1 N HCl at 50 rpm on mechanical shaker and observed for solubilization, if any, of microparticles.
2.3.3.
Entrapment efficiency
100 mg of microparticles were weighed and dispersed into 20 ml of ethanol in a beaker and the beaker was wrapped with aluminum foil. Microparticles were then digested for 24 h in the darkness and then sonicated for 1 h. Sonicated sample was then filtered by using Whatman filter paper. Filtered sample was then analyzed by using UV spectrophotometer after suitable dilution. From the reading, by using following formula % of entrapment was calculated.
Spray drying of chitosan with crosslinker % of entrapment ¼
Various crosslinking agents are utilized for crosslinking purpose like glutaraldehyde, genepin, formaldehyde. Crosslinking occurs in between chitosan molecules retarding their water solubility. 25% Glutaraldehyde is utilized for crosslinking of chitosan while spray drying.16e18
2.2.
Trial 1
1 g of chitosan was dissolved in 100 ml 5% dilute acetic acid solution. In it 25 ml of 25% of glutaraldehyde was added. Allowed to crosslink for 15 min. After 15 min very thick gel was formed such that it can’t be passed through the spray drying system. So it was started with 1 ml of glutaraldehyde.
2.3.
Trial 2
1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 1 ml of 25% glutaraldehyde was added and allowed to crosslink for 15 min while stirring. Above solution was kept for stirring and spray dried at conditions given in Table 1. Obtained product was collected, weighed and evaluated for following parameters.
2.3.1.
% of yield
Obtained product was weighed and % of yield was calculated by using following formula: % of yield ¼
Amount of product obtained 100 Amount of total solid in spray drying solution
150 C 100 C 1 ml per min 40 Nm3/h 1 bar
2.3.4.
Practical drug content 100 Theoretical drug content
% Drug loading
% of drug loading was calculated to find out % of amount of drug present in given weight of microspheres. % of drug loading was calculated by using following formula: % of loading ¼
2.3.5.
Drug content 100 Weight of microspheres
Drug release
Drug release was checked for 5 h by using USP paddle apparatus. 900 ml of 0.1 N HCl was utilized as a media. Microparticles were weighed such that it becomes equivalent to 9 mg of budesonide. Then microparticles were filled into size 4 capsule. Capsule was then placed into media at 50 rpm and 37 0.5 C. 5 ml sample was withdrawn at each 1 h and analyzed by UV. If required suitable dilutions were prepared. Dissolution was carried out for 5 h only to check drug release occurring in critical period.19,20 Graph was plotted as % of drug release versus time.
2.4.
Trial 3
In this trial amount of crosslinker (glutaraldehyde) was increased upto 2 ml to increase crosslinking and ultimately to decrease drug release. 1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2 ml of 25% glutaraldehyde was added and allowed to react for 15 min. Above solution was kept for stirring and spray dried at conditions mentioned in Table 1. Outlet temperature was varied between 100 and 60 C.
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j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
Obtained product was collected and weighed. % Yield was calculated. Microparticles were again evaluated for all the above mentioned parameters.
2.7.2.
2.5.
2.7.3.
Trial 4
In this trial again amount of crosslinker was increased.1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 3 ml of 25% glutaraldehyde was added and allowed to react for 15 min. After 15 min change in gel was observed and a very thick jelly like mass was obtained which was not at all passable through spray drying system.
2.6.
Trial 5
Amount of chitosan is increased and in proportion with chitosan amount of glutaraldehyde was also increased. 1.2 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2.4 ml of 25% glutaraldehyde was added and allowed to react for 15 min. Above solution was kept for stirring and dried at conditions given in Table 1. After starting of spray drying when near about 30 ml feed was remained, it got gelled and was unable to pass through spray drying system. So trial was stopped there.
2.7.
DSC
DSC of the microparticles was carried out to find interaction, if any, in between chitosan, glutaraldehyde and drug. DSC was carried out at Diya Labs, Mumbai. Sample was sealed into aluminum pan with lid pierced. Heating range was 10 K/min. with nitrogen purging at 60 ml/min.
2.7.4.
FTIR
FTIR was recorded on Bruker alpha. Microparticles were mixed with moisture free KBr in 1:100 ratio. Pressed into thin disc by using KBr press and FTIR of the disc was recorded from 500 cm1 to 4000 cm1.
2.7.5.
Particle size analysis
Particle size analysis was performed by using laser diffraction particle size analyzer (Malvern Mastersizer, UK). About 500 mg of microcapsules were weighed and suspended in 500 ml benzene, ultrasonicated for 2 min to form uniform dispersion and analyzed for particle size.
3.
Results and discussion
3.1.
Spray drying of chitosan with crosslinker
Glutaraldehyde was utilized for the crosslinking purpose of chitosan.
Trial 3A
Trial 3 was again conducted to check the effect of outlet temperature on product yield. In previous trial outlet temperature was varying between 100 and 60 C, but this time outlet temperature was varied between 100 and 90 C. Product was collected and weighed and evaluated further for the following parameters.
2.7.1.
Scanning electron microscopy
Scanning electron microscopy (SEM) was carried out at Diya labs, Mumbai.
Drug release for 24 h
Dissolution study was carried out for 24 h in USP type 2 apparatus (Paddle) in triplicate manner. Initial 2 h drug release was checked in simulated gastric fluid, then for next 3 h pH of the media was increased upto 6.8 by adding 1 M NaOH and addition of 10 g of pancreatin was done and after 5 h pH of the media was increased upto 7.4 and addition of rat cecal content was done into simulated colonic environment. Dissolution study was carried out in triplicate manner. Graph was plotted as % of drug release versus time.
3.2.
Trial 1
Initially 1 g of chitosan was dissolved in 100 ml of 5% dilute acetic acid solution. In it 25 ml of 25% of glutaraldehyde was added. Allowed to crosslink for 15 min. After 15 min very thick gel was formed such that it can’t be passed through the spray drying system. This may be happened due to excess of glutaraldehyde. Excess of glutaraldehyde causes dense network formation between the molecules of chitosan which results in formation of very thick gel.
3.3.
Trial 2
In next trial amount of glutaraldehyde was reduced to minimum level. In trial 2, 1 ml of glutaraldehyde was utilized for crosslinking purpose. After 15 min of crosslinking no gel formation occurs and solution remains in a condition such that it
Table 2 e Result of evaluation test of various trials of chitosan with crosslinker. Sr. no
1 2 3
Parameter
% Yield % Entrapment % Drug loading
Result Trial 1
Trial 2
Trial 3
Trial 4
Trial 5
Trial 3A
e e e
14.10% 91.23% 45.5%
16.20% 92.28% 46.14%
e e e
11.12% 84.24% 42.12%
24.56% 92.21% 46.15%
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
120
preceding 4 h only 25% of drug release was occurred as shown in Fig. 1.
% Drug Release
100
3.4.
80
Trial 3
Trial 02
60
Trial 03
40
Trial 05 Trial 3A
20 0
459
0
10
20 Time (Hours)
30
Fig. 1 e Drug release profile of various trials.
can be passed through the spray drying system. Spray drying parameters were chosen by considering water as a solvent. When addition of ethanolic solution of drug was added to chitosan solution, precipitation of drug was occurred in very fine particles. Now in this case polymer is in solubilized state and drug is insoluble. So in this case microparticles may be of microcapsule type, embedding drug molecules inside the polymer coat. After spray drying microparticles were weighed, % yield was calculated and checked for integrity purpose. 100 mg of microparticles were kept in 100 ml of 0.1 N HCl at 50 rpm on mechanical shaker and observed for 24 h. After 24 h of shaking, microparticles were found to be as it is. No solubilization of microparticles was occurred, so microparticles were evaluated for further parameters. Further evaluation was carried out for % of drug entrapment, % of drug loading and drug release and results were as shown in Table 2. In 5 h near about 35% of drug release occurred in acidic media which indicated that crosslinking was occurred but not in the amount which was required. Amount of crosslinker was not enough. So in next trial amount of crosslinker was increased in order to increase crosslinking. In initial 1 h about 10% of drug release was occurred which may be indication of presence of drug on the surface of microparticles which is going into media immediately after addition. After 1 h drug release is in sustain manner. In
In this trial amount of crosslinker was increased upto 2 ml. 1 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of budesonide was added to 20 ml of ethanol, dissolved and added to the chitosan solution. After proper mixing 2 ml of 25% glutaraldehyde was added and allowed to react for 15 min and spray dried. Evaluation of product was carried out as per previous batch. Noticeable change was not observed in drug content which suggested that there is no considerable impact of crosslinking agent on the drug content. Drug release was calculated for 5 h and found to be 19% after 5 h as shown in Fig. 2. Result in decrease in drug release was noticed due to increased amount of crosslinking which is caused by increased amount of glutaraldehyde. There are more number of glutaraldehyde molecules present for interchain crosslinking of amino groups of adjacent chitosan molecules. As the number of bridges between two chitosan chains increased, stiffness of chitosan molecules also increased resulting in uptake of lesser amount of water and less swellability and solubility.
3.5.
Trial 4
In this trial amount of crosslinker was increased upto 3 ml. Preparation of feed was done in same manner as that of previous batches. Crosslinking time was also kept 15 min. But due to increased amount of crosslinker thick gel was obtained after 15 min which was not passable through spray drying system. Gel formation occurred due to excess amount of glutaraldehyde. So instead of increasing crosslinking agent to 3 ml, both chitosan and glutaraldehyde were increased in proportion wise manner by taking into consideration 2 ml of glutaraldehyde for crosslinking of 1 g of chitosan.
3.6.
Trial 5
In this trial amount of chitosan and glutaraldehyde was increased in proportion wise manner. 1.2 g chitosan was dissolved in 100 ml dilute acetic acid solution (5%). 500 mg of
Fig. 2 e SEM image results of trial 3A.
460
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
budesonide was added to 20 ml of ethanol and added to the chitosan solution. After proper mixing 2.4 ml of 25% glutaraldehyde was added and allowed to react for 15 min. After 15 min no thick gel formation occurred so spray drying was started. When near about 30 ml of feed was remained thick gel formation occurred which was not able to pass through spray drying system. So spray drying was stopped, product was collected and evaluated. After 5 h 25% of drug release occurred as shown in Fig. 1, which was not desirable. This may be happened due to gelling of remaining 30 ml of feed, failing it to be spray dried. From the above trials it was concluded that 2 ml of 25% of glutaraldehyde is maximum amount which can be utilized for crosslinking purpose of 1 g chitosan having degree of deacetylation 70e90% in 5% acetic acid solution without formation of thick gel which can be passed through nozzle of spray dryer by taking 15 min as a crosslinking time.
3.7.
Trial 3A
Trial 3A was conducted to find out the effect of temperature variation on % of yield. In this trial outlet temperature was varied between 100 and 90 C. In previous trial outlet temperature was varying between 100 and 60 C. % of yield obtained in this trial is more as compared to batch 3. This may be happened due to increase in drying rate due to maintaining temperature in the range of boiling point of the solvent. Evaluation of batch 3A was carried out.
3.7.1.
Dissolution study
Dissolution study was carried out 24 h. Graph of % of drug release versus time was plotted as follows. In initial 5 h 20% of drug release has occurred. In initial 1 h 10% drug release was obtained. Once addition of pancreatin was done, drug release increased slightly. After 5 h 20% of drug release was occurred. But once addition of rat cecal content is carried out drug release increased rapidly. In next 2 h 97% of drug was released. Results were shown in Fig. 1. This indicates that after
Fig. 4 e DSC spectra of budesonide microparticles.
crosslinking of chitosan, it retains its specific biodegradability by colonic micro-organisms.19
3.7.2.
Scanning electron microscopy
Scanning electron microscopy was carried out to find out morphology of microparticles. Results of SEM are as shown in Fig. 2. SEM images indicate morphology of microparticles which was smooth in appearance and spherical in shape and having size less than 5 mm. Small size may be contributed to the microparticles due to apparatuses size of atomizer high atomization pressure during spray drying. Surface of the microparticles is smooth without any grooves which indicate that coating has occurred uniformly.
3.7.3.
DSC
DSC of the microparticles was carried out to check possible interaction in between drug and polymer. DSC graph showed endothermic peak near 160 C which is indication of presence of drug. In DSC graph of pure budesonide endothermic peak was also observed at 160 C as shown in Fig. 4.
3.7.4.
FTIR of microparticles
FTIR spectra of microparticles was recorded by using Bruker alpha. Microparticles showed the presence of particular groups
Fig. 3 e FTIR spectra of budesonide microparticles.
Fig. 5 e Particle size distribution of trial 3A.
j o u r n a l o f p h a r m a c y r e s e a r c h 6 ( 2 0 1 3 ) 4 5 6 e4 6 1
which are present in FTIR spectra of budesonide as shown in Fig. 3.
3.7.5.
Particle size analysis
Particle size analysis was performed on Malvern Mastersizer. Maximum particle size was found be distributed in the range of 2e5 mm. Results were shown in Fig. 5. Less particle size is obtained which may be contributed to the method of microsphere preparation which is spray drying. Other methods such as solvent evaporation, emulsion method generates particles of higher size.
Conflicts of interest All authors have none to declare.
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