Skip to main content
Log in

Efficient Encapsulation of a Water-Soluble Molecule into Lipid Vesicles Using W/O/W Multiple Emulsions via Solvent Evaporation

  • Original Paper
  • Published:
Journal of the American Oil Chemists' Society

Abstract

We developed a novel method for preparing lipid vesicles with high entrapment efficiency and controlled size using water-in-oil-in-water (W/O/W) multiple emulsions as vesicle templates. Preparation consists of three steps. First, a water-in-oil (W/O) emulsion containing to-be-entrapped hydrophilic molecules in the water phase and vesicle-forming lipids in the oil phase was formulated by sonication. Second, this W/O emulsion was introduced into a microchannel emulsification device to prepare a W/O/W multiple emulsion. In this step, sodium caseinate was used as the external emulsifier. Finally, organic solvent in the oil phase was removed by simple evaporation under ambient conditions to afford lipid vesicles. The diameter of the prepared vesicles reflected the water droplet size of the primary W/O emulsions, indicating that vesicle size could be controlled by the primary W/O emulsification process. Furthermore, high entrapment yields for hydrophilic molecules (exceeding 80 % for calcein) were obtained. The resulting vesicles had a multilamellar vesicular structure, as confirmed by transmission electron microscopy.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10

Similar content being viewed by others

References

  1. Szoka F Jr, Papahadjopoulos D (1980) Comparative properties and methods of preparation of lipid vesicles (liposomes). Ann Rev Biophys Bioeng 9:467–508

    Article  CAS  Google Scholar 

  2. Oku N, Kendall DA, MacDonald RC (1982) A simple procedure for the determination of the trapped volume of liposomes. Biochim Biophys Acta 691:332–340

    Article  CAS  Google Scholar 

  3. Walde P, Ichikawa S (2001) Enzymes in lipid vesicles: preparation, reactivity and applications. Biomol Eng 18:143–177

    Article  CAS  Google Scholar 

  4. Duzgunes N (ed) (2003) Liposomes. Methods in enzymology, vol 367. Academic, San Diego

    Google Scholar 

  5. Torchilin VP, Weissig V (eds) (2003) Liposomes: a practical approach, 2nd edn. Oxford University Press, Oxford

    Google Scholar 

  6. Szoka F Jr, Papahadjopoulos D (1978) Procedure for preparation of liposomes with large internal aqueous space and high capture by reverse-phase evaporation. Proc Natl Acad Sci USA 75:4194–4198

    Article  CAS  Google Scholar 

  7. Zhang L, Hu J, Lu Z (1997) Preparation of liposomes with a controlled assembly procedure. J Colloid Interfaces Sci 190:76–80

    Article  CAS  Google Scholar 

  8. Pautot S, Frisken BJ, Weitz DA (2003) Production of unilamellar vesicles using an inverted emulsion. Langmuir 19:2870–2879

    Article  CAS  Google Scholar 

  9. Tan YC, Hettiarachchi K, Siu M, Pan YR, Lee AP (2006) Controlled microfluidic encapsulation of cells, proteins, and microbeads in lipid vesicles. J Am Chem Soc 128:5656–5658

    Article  CAS  Google Scholar 

  10. Sugiura S, Kuroiwa T, Kagota T, Nakajima M, Sato S, Mukataka S, Walde P, Ichikawa S (2008) Novel method for obtaining homogeneous giant vesicles from a monodisperse water-in-oil emulsion prepared with a microfluidic device. Langmuir 24:4581–4588

    Article  CAS  Google Scholar 

  11. Kuroiwa T, Kiuchi H, Noda K, Kobayashi I, Nakajima M, Uemura K, Sato S, Mukataka S, Ichikawa S (2009) Controlled preparation of giant vesicles from uniform water droplets obtained by microchannel emulsification with bilayer-forming lipids as emulsifiers. Microfluid Nanofluid 6:811–821

    Article  CAS  Google Scholar 

  12. Kuroiwa T, Fujita R, Kobayashi I, Uemura K, Nakajima M, Sato S, Walde P, Ichikawa S (2012) Efficient preparation of giant vesicles as biomimetic compartment systems with high entrapment yields for biomacromolecules. Chem Biodivers 9:2453–2472

    Article  CAS  Google Scholar 

  13. Nishimura K, Suzuki H, Toyota T, Yomo T (2012) Size control of giant unilamellar vesicles prepared from inverted emulsion droplets. J Colloid Interfaces Sci 376:119–125

    Article  CAS  Google Scholar 

  14. Ishii F, Takamura A, Ogata H (1988) Preparation conditions and evaluation of the stability of lipid vesicles (liposomes) using the microencapsulation technique. J Dispers Sci Technol 9:1–15

    Article  CAS  Google Scholar 

  15. Zheng S, Zheng Y, Beissinger RL, Fresco R (1994) Microencapsulation of hemoglobin in liposomes using double emulsion, film dehydration/rehydration approach. Biochim Biophys Acta 1196:123–130

    Article  Google Scholar 

  16. Nii T, Ishii F (2005) Encapsulation efficiency of water-soluble and insoluble drugs in liposomes prepared by the microencapsulation vesicle method. Int J Pharm 198:198–205

    Article  Google Scholar 

  17. Wang T, Deng Y, Geng Y, Gao Z, Zou J, Wang Z (2006) Preparation of submicron unilamellar liposomes by freeze-dyring double emulsions. Biochim Biophys Acta 1758:222–231

    Article  CAS  Google Scholar 

  18. Shum HC, Lee D, Yoon I, Kodger T, Weitz DA (2008) Double emulsion template monodisperse phospholipid vesicles. Langmuir 24:7651–7653

    Article  CAS  Google Scholar 

  19. Kawakatsu T, Kikuchi Y, Nakajima M (1997) Regular-sized cell creation in microchannel emulsification by visual microprocessing method. J Am Oil Chem Soc 74:317–321

    Article  CAS  Google Scholar 

  20. Sugiura S, Nakajima M, Iwamoto S, Seki M (2001) Interfacial tension driven monodispersed droplet formation from microfabricated channel array. Langmuir 17:5562–5566

    Article  CAS  Google Scholar 

  21. Sugiura S, Nakajima M, Yamamoto K, Iwamoto S, Oda T, Satake M, Seki M (2004) Preparation characteristics of water-in-oil-in-water multiple emulsions using microchannel emulsification. J Colloid Interfaces Sci 270:221–228

    Article  CAS  Google Scholar 

  22. Kobayashi I, Lou X, Mukataka S, Nakajima M (2005) Preparation of monodisperse water-in-oil-in-water emulsions using microfluidization and straight-through microchannel emulsification. J Am Oil Chem Soc 82:65–71

    Article  CAS  Google Scholar 

  23. Souilem S, Kobayashi I, Neves MA, Sayadi S, Ichikawa S, Nakajima M (2014) Preparation of monodisperse food-grade oleuropein-loaded W/O/W emulsions using microchannel emulsification and evaluation of their storage stability. Food Bioprocess Technol 7:2014–2027

  24. Garti N, Aserin A (1996) Double emulsions stabilized by macromolecular surfactants. Adv Colloid Interfaces Sci 65:37–69

    Article  CAS  Google Scholar 

  25. Garti N (1997) Progress in stabilization and transport phenomena of double emulsions in food applications. LWT Food Sci Technol 30:222–235

    Article  CAS  Google Scholar 

  26. Lichtenberg D, Markello T (1984) Structural characteristics of phospholipid multilamellar liposomes. J Pharm Sci 73:122–125

    Article  CAS  Google Scholar 

  27. Florence AT, Whitehill D (1981) Some features of breakdown in water-in-oil-in-water multiple emulsions. J Colloid Interfaces Sci 79:243–256

    Article  CAS  Google Scholar 

  28. Kirby C, Gregoriadis G (1984) Dehydration-rehydration vesicles: a simple method for high yield drug entrapment in liposomes. Bio/Technology 2:979–984

    Article  CAS  Google Scholar 

  29. Sugiura S, Ichikawa S, Sano Y, Nakajima M, Liu XQ, Seki M, Frusaki S (2001) Formation and characterization of reversed micelles composed of phosphoslids and fatty acid. J Colloid Interfaces Sci 240:566–572

    Article  CAS  Google Scholar 

  30. Böckmann RA, Hac A, Heinberg T, Grubmüller H (2003) Effect of sodium chloride on a lipid bilayer. Biophys J 85:1647–1655

    Article  Google Scholar 

  31. van den Bogaart G, Hermans N, Krasnikov V, de Vries AH, Poolman B (2007) On the decrease in lateral mobility of phospholipids by sugars. Biophys J 92:1598–1605

    Article  Google Scholar 

Download references

Acknowledgments

We thank Prof. Peter Walde (ETH-Zürich, Zürich, Switzerland) for stimulating discussions. This work was partially supported by the Adaptable and Seamless Technology Transfer Program through Target-driven R&D (A-STEP) (No. AS232Z02816F) of the Japan Science and Technology Agency, and a Grant-in-Aid for Young Scientists (B) (No. 22760613) from the Japan Society for the Promotion of Science.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Sosaku Ichikawa.

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kuroiwa, T., Horikoshi, K., Suzuki, A. et al. Efficient Encapsulation of a Water-Soluble Molecule into Lipid Vesicles Using W/O/W Multiple Emulsions via Solvent Evaporation. J Am Oil Chem Soc 93, 421–430 (2016). https://doi.org/10.1007/s11746-015-2777-2

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11746-015-2777-2

Keywords

Navigation