Preparation and evaluation of oleoyl-carboxymethy-chitosan (OCMCS) nanoparticles as oral protein carriers

  • Ya Liu
  • Xiao Jie Cheng
  • Qi Feng Dang
  • Fang Kui Ma
  • Xi Guang Chen
  • Hyun Jin Park
  • Bum Keun Kim


Oleoyl-carboxymethy chitosan (OCMCS) nanoparticles based on chitosan with different molecular weights (50, 170 and 820 kDa) were prepared by self-assembled method. The nanoparticles had spherical shape, positive surface charges and the mean diameters were 157.4, 274.1 and 396.7 nm, respectively. FITC-labeled OCMCS nanoparticles were internalized via the intestinal mucosa and observed in liver, spleen, intestine and heart following oral deliverance to carps (Cyprinus carpio). Extracellular products (ECPs) of Aeromonas hydrophila as microbial antigen was efficiently loaded to form OCMCS–ECPs nanoparticles and shown to be sustained release in PBS. Significantly higher (P < 0.05) antigen-specific antibodies were detected in serum after orally immunized with OCMCS-ECPs nanoparticles than that immunized with ECPs alone and non-immunized in control group in carps. These results implied that amphiphilic modified chitosan nanoparticles had great potential to be applied as carriers for the oral administration of protein drugs.


Chitosan Zeta Potential Protein Drug Aeromonas Hydrophila Mucoadhesive Property 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



This work was supported by grants from the National Natural Science Foundation of China (NSFC, 81071274), International S & T Cooperation Program of China (ISTCP, S2011GR0092).


  1. 1.
    Radwan MA, Aboul-Enein HY. The effect of oral absorption enhancers on the in vivo performance of insulin-loaded poly (ethylcyanoacrylate) nanospheres in diabetic rats. J Microencapsul. 2002;19:225–35.CrossRefGoogle Scholar
  2. 2.
    Steffansen B, Nielsen CU, Brodin B, Eriksson AH, Anderson R, Frokjaer S. Intestinal solute carriers: an overview of trends and strategies for improving oral drug absorption. Eur J Pharm Sci. 2004;21:3–16.CrossRefGoogle Scholar
  3. 3.
    Avadi MR, Sadeghi AMM, Mohammadpour N, Abedin S, Atyabi F, Dinarvand R, et al. Preparation and characterization of insulin nanoparticles using chitosan and Arabic gum with ionic gelation method. Nanomed-Nanotechnol. 2010;6:58–63.CrossRefGoogle Scholar
  4. 4.
    Longer MA, Cheng HS, Robinson JR. Bioadhesive polymers as platforms for oral controlled drug delivery III: oral delivery of chlorothiazide using a bioadhesive polymer. J Pharm Sci. 1985;74:406–11.CrossRefGoogle Scholar
  5. 5.
    Sakuma S, Hayashi M, Akashi M. Design of nanoparticles composed of graft copolymers. Adv Drug Deliv Rev. 2001;47:21–37.CrossRefGoogle Scholar
  6. 6.
    Yin L, Ding JY, He CB, Cui LM, Tang C, Yin CH. Drug permeability and mucoadhesion properties of thiolated trimethyl chitosan nanoparticles in oral insulin delivery. Biomaterials. 2009;30:5691–700.CrossRefGoogle Scholar
  7. 7.
    Janes KA, Calvo P, Alonso MJ. Polysaccharide colloidal particles as delivery systems for macromolecules. Adv Drug Deliv Rev. 2001;47:83–97.CrossRefGoogle Scholar
  8. 8.
    Hassan EE, Gallo JM. A simple rheological method for the in vitro assessment of mucin–polymer bioadhesive bond strength. Pharm Res. 1990;7:491–5.CrossRefGoogle Scholar
  9. 9.
    Moghaddam FA, Atyabi F, Dinarvand R. Preparation and in vitro evaluation of mucoadhesion and permeation enhancement of thiolated chitosan-pHEMA core-shell nanoparticles. Nanomed-Nanotechnol. 2009;5:208–15.CrossRefGoogle Scholar
  10. 10.
    Li YY, Chen XG, Liu CS, Cha DS, Park HJ, Lee CM. Effect of the molecular mass and degree of substitution of oleoylchitosan on the structure, rheological properties, and formation of nanoparticles. J Agric Food Chem. 2007;55:4842–7.CrossRefGoogle Scholar
  11. 11.
    Li YY, Chen XG, Yu LM, Wang SX, Sun GZ, Zhou HY. Aggregation of hydrophobically modified chitosan in solution and at the air-water interface. J Appl Polym Sci. 2006;102:1968–73.CrossRefGoogle Scholar
  12. 12.
    Zhang J, Chen XG, Li YY, Liu CS. Self-assembled nanoparticles based on hydrophobically modified chitosan as carriers for doxorubicin. Nanomed-Nanotechnol. 2007;3:258–65.CrossRefGoogle Scholar
  13. 13.
    Zhang J, Chen XG, Liu CS, Hark HJ. Investigation of polymeric amphiphilic nanoparticles as antitumor drug carriers. J Mater Sci Mater Med. 2009;20(4):991–9.CrossRefGoogle Scholar
  14. 14.
    Azad IS, Shankar KM, Mohan CV, Kalita B. Biofilm vaccine of Aeromonas hydrophila–standardization of dose and duration for oral vaccination of carps. Fish Shellfish Immunol. 1999;9:519–28.CrossRefGoogle Scholar
  15. 15.
    Vivas J, Riaño J, Carracedo B, Razquin BE, López-Fierro P, Naharro G, et al. The auxotrophic aroA mutant of Aeromonas hydrophila as a live attenuated vaccine against A. salmonicida infections in rainbow trout (Oncorhynchus mykiss). Fish Shellfish Immunol. 2004;16:193–206.CrossRefGoogle Scholar
  16. 16.
    Chen XG, Lee CM, Park HJ. O/W emulsification for the self-aggregation and nanoparticle formation of linoleic acids modified chitosan in the aqueous system. J Agric Food Chem. 2003;51:3135–9.CrossRefGoogle Scholar
  17. 17.
    Shigemasa Y, Matsuura H, Sashiwa H, Saimoto H. Evaluation of different absorbance ratios from infrared spectroscopy for analyzing the degree of deacetylation in chitin. Int J Biol Macromol. 1996;18:237–42.CrossRefGoogle Scholar
  18. 18.
    Wan LQ, Hu FQ, Yuan H. Study of the uptake of chitosan oligosaccharide nanoparticles by A549 cells. Acta Pharmacol Sin. 2004;39:227–31.Google Scholar
  19. 19.
    Rodriguez LA, Ellis AE, Nieto TP. Purification and characterisation of an extracellular metalloprotease, serine protease and haemolysin of Aeromonas hydrophila strain B sub (32): all are lethal for fish. Microb Pathog. 1992;13:17–24.CrossRefGoogle Scholar
  20. 20.
    Gan Q, Wang T. Chitosan nanoparticle as protein delivery carrier—systematic examination of fabrication conditions for efficient loading and release. Colloid Surf B. 2007;59:24–34.CrossRefGoogle Scholar
  21. 21.
    Irie T, Watarai S, Iwasaki T, Kodama H. Protection against experimental Aeromonas salmonicida infection in carp by oral immunisation with bacterial antigen entrapped liposomes. Fish Shellfish Immunol. 2005;18:235–42.CrossRefGoogle Scholar
  22. 22.
    Chen XG, Park HJ. Chemical characteristics of O-carboxymethyl chitosans related to the preparation conditions. Carbohyd Polym. 2003;53:355–9.CrossRefGoogle Scholar
  23. 23.
    Xu Y, Du Y. Effect of molecular structure of chitosan on protein delivery properties of chitosan nanoparticles. Int J Pharm. 2003;250:215–26.CrossRefGoogle Scholar
  24. 24.
    Irie T, Watarai S, Kodama H. Humoral immune response of carp (Cyprinus carpio) induced by oral immunization with liposome-entrapped antigen. Dev Com Immunol. 2003;27:413–21.CrossRefGoogle Scholar
  25. 25.
    Owens DR, Zinman B, Bolli G. Alternative routes of insulin delivery. Diab Med. 2003;20:886–98.CrossRefGoogle Scholar
  26. 26.
    Khin YW, Feng SS. Effects of particle size and surface coating on cellular uptake of polymeric nanoparticles for oral delivery of anticancer drugs. Biomaterials. 2005;26:2713–22.CrossRefGoogle Scholar
  27. 27.
    Florence AT, Hillery AM, Hussain N, Jani PU. Nanoparticles as carriers for oral peptide absorption: studies on particle uptake and fate. J Control Release. 1995;36:39–46.CrossRefGoogle Scholar
  28. 28.
    Robert MS, Karlo P, Fanor B, Michael L, Tarek MF. The use of deoxycholic acid to enhance the oral bioavailability of biodegradable nanoparticles. Biomaterials. 2008;29:703–8.CrossRefGoogle Scholar
  29. 29.
    van der Merwe SM, Verhoef JC, Verheijden JHM, Kotzé AF, Junginger HE. Trimethylated chitosan as polymeric absorption enhancer for improved peroral delivery of peptide drugs. Eur J Pharm Biopharm. 2004;58:225–35.CrossRefGoogle Scholar
  30. 30.
    Calvo P, Remuñán-López C, Vila-Jato JL, Alonso MJ. Novel hydrophilic chitosan-polyethylene oxide nanoparticles as protein carriers. J Appl Polym Sci. 1997;63:125–32.CrossRefGoogle Scholar
  31. 31.
    Son YJ, Jang JS, Cho YW, Chung H, Park RW, Kwon IC, et al. Biodistribution and anti-tumor efficacy of doxorubicin loaded glycol-chitosan nanoaggregates by EPR effect. J Control Release. 2003;91:135–45.CrossRefGoogle Scholar
  32. 32.
    Bravo-Osuna I, Vauthier C, Farabollini A, Palmieri GF, Ponchel G. Mucoadhesion mechanism of chitosan and thiolated chitosan-poly (isobutyl cyanoacrylate) core-shell nanoparticles. Biomaterials. 2007;28:2233–43.CrossRefGoogle Scholar
  33. 33.
    Han HD, Lee A, Song CK, Hwang T, Seong H, Lee CO. In vivo distribution and antitumor activity of heparin-stabilized doxorubicin-loaded liposomes. Int J Pharm. 2006;313:181–8.CrossRefGoogle Scholar
  34. 34.
    Bernkop-Schnürch A, Hornof M, Guggi D. Thiolated chitosans. Eur J Pharm Biopharm. 2004;57:9–17.CrossRefGoogle Scholar
  35. 35.
    Schipper NGM, Olsson S, Hoogstraate JA, de Boer AG, Varum KM, Artursson P. Chitosans as absorption enhancers for poorly absorbable drugs. 2: Mechanism of absorption enhancement. Pharm Res. 1997;14:923–9.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Ya Liu
    • 1
  • Xiao Jie Cheng
    • 1
  • Qi Feng Dang
    • 1
  • Fang Kui Ma
    • 1
  • Xi Guang Chen
    • 1
  • Hyun Jin Park
    • 2
  • Bum Keun Kim
    • 3
  1. 1.College of Marine Life ScienceOcean University of ChinaQingdaoPeople’s Republic of China
  2. 2.College of Life Sciences and BiotechnologyKorea UniversitySeoulKorea
  3. 3.Korea Food Research InstituteSeoulKorea

Personalised recommendations