Applied Nanoscience

, Volume 8, Issue 6, pp 1483–1491 | Cite as

Injectable nanoemulsions prepared by high pressure homogenization: processing, sterilization, and size evolution

  • Martina Rosi Cappellani
  • Diego Romano Perinelli
  • Laura Pescosolido
  • Aurélie Schoubben
  • Marco Cespi
  • Riccardo Cossi
  • Paolo BlasiEmail author
Original Article


Oil-in-water nanoemulsions are promising colloidal dispersions for the delivery of hydrophobic drugs. The use of polysorbate 80 as stabilizing agent in these formulations can improve the delivery of the drug to the brain, thanks to a tropism for apolipoproteins. The aim of this study was to formulate injectable nanoemulsions stabilized by polysorbate 80, which meet United States Pharmacopoeia particle size requirements for parenteral emulsions. Nanoemulsions were prepared by high pressure homogenization and characterized in terms of mean hydrodynamic diameter, Gaussian distribution width, and volume-weighted percentage of fat droplets greater than 1.79 µm (PFAT1.79) or 5 µm (PFAT5). The effect of autoclaving, filtration and loading with nile red (a lipophilic fluorescent dye) on the nanoemulsions was evaluated. Real-time and accelerated stability tests were also performed. To satisfy Unites States Pharmacopoeia particle size specifications, nanoemulsions required six homogenization cycles. PFAT5 and PFAT1.79 were the particle size parameters more sensitive to discriminate the effect of homogenization, autoclaving, filtration and loading as well as globule size evolution during real-time stability tests. Results from accelerated stability studies correlated with the PFAT5 values measured over time. Overall, the study demonstrates that all nanoemulsions studied (autoclaved, filtered or loaded) satisfies United States Pharmacopoeia particle size requirements up to 90 days, maintaining PFAT5 to values lower than 0.05% v/v.


Parenteral emulsions Drug delivery Filtration Autoclaving Accelerated stability Ultrasound technology Y-shape valve 



  1. Aggarwal P, Hall JB, McLeland CB et al (2009) Nanoparticle interaction with plasma proteins as it relates to particle biodistribution, biocompatibility and therapeutic efficacy. Adv Drug Deliv Rev 61:428–437. CrossRefGoogle Scholar
  2. Allegra JR, Hawley SA (1972) Attenuation of sound in suspensions and emulsions: theory and experiments. J Acoust Soc Am 51:1545–1564. CrossRefGoogle Scholar
  3. Bhattacharjee S (2016) DLS and zeta potential – What they are and what they are not? J Control Release 235:337–351. CrossRefGoogle Scholar
  4. Blasi P, Giovagnoli S, Schoubben A et al (2007) Solid lipid nanoparticles for targeted brain drug delivery. Adv Drug Deliv Rev 59:454–477. CrossRefGoogle Scholar
  5. Blasi P, Giovagnoli S, Schoubben A et al (2011) Lipid nanoparticles for brain targeting I. Formulation optimization. Int J Pharm 419:287–295. CrossRefGoogle Scholar
  6. Bonacucina G, Perinelli DR, Cespi M et al (2016) Acoustic spectroscopy: a powerful analytical method for the pharmaceutical field? Int J Pharm 503:174–195. CrossRefGoogle Scholar
  7. Calder PC, Jensen GL, Koletzko BV et al (2010) Lipid emulsions in parenteral nutrition of intensive care patients: current thinking and future directions. Intensive Care Med 36:735–749. CrossRefGoogle Scholar
  8. Chansiri G, Lyons RT, Patel MV, Hem SL (1999) Effect of surface charge on the stability of oil/water emulsions during steam sterilization. J Pharm Sci 88:454–458. CrossRefGoogle Scholar
  9. Chapter < 729>: globule size distribution in lipid injectable emulsion (2013) In: United States Pharmacopoeia. 36th edGoogle Scholar
  10. Chen H, Jin X, Li Y, Tian J (2016) Investigation into the physical stability of a eugenol nanoemulsion in the presence of a high content of triglyceride. RSC Adv 6:91060–91067. CrossRefGoogle Scholar
  11. Dal Magro R, Albertini B, Beretta S et al (2018) Artificial apolipoprotein corona enables nanoparticle brain targeting. Nanomedicine 14:429–438. CrossRefGoogle Scholar
  12. Delmas T, Piraux H, Couffin A-C et al (2011) How to prepare and stabilize very small nanoemulsions. Langmuir 27:1683–1692. CrossRefGoogle Scholar
  13. Driscoll DF (2002) The significance of particle/globule-sizing measurements in the safe use of intravenous lipid emulsions. J Dispers Sci Technol 23:679–687. CrossRefGoogle Scholar
  14. Driscoll DF (2006) Lipid injectable emulsions: pharmacopeial and safety issues. Pharm Res 23:1959–1969. CrossRefGoogle Scholar
  15. Dukhin AS, Goetz PJ (eds) (2010) Acoustic theory for particulates. In: Characterization of liquids, nano- and microparticulates, and porous bodies using ultrasound studies in interface science, chap 4, vol 24. Elsevier, Amsterdam, pp 127–185. CrossRefGoogle Scholar
  16. Floyd AG (1999) Top ten considerations in the development of parenteral emulsions. Pharm Sci Technolo Today 2:134–143. CrossRefGoogle Scholar
  17. Ganta S, Talekar M, Singh A et al (2014) Nanoemulsions in translational research—opportunities and challenges in targeted cancer therapy. AAPS Pharm Sci Tech 15:694–708. CrossRefGoogle Scholar
  18. Göppert TM, Müller RH (2005) Polysorbate-stabilized solid lipid nanoparticles as colloidal carriers for intravenous targeting of drugs to the brain: comparison of plasma protein adsorption patterns. J Drug Target 13:179–187. CrossRefGoogle Scholar
  19. Gupta A, Eral HB, Hatton TA, Doyle PS (2016) Nanoemulsions: formation, properties and applications. Soft Matter 12:2826–2841. CrossRefGoogle Scholar
  20. Hippalgaonkar K, Majumdar S, Kansara V (2010) Injectable lipid emulsions—advancements, opportunities and challenges. AAPS PharmSciTech 11:1526–1540. CrossRefGoogle Scholar
  21. Hörmann K, Zimmer A (2016) Drug delivery and drug targeting with parenteral lipid nanoemulsions—a review. J Control Release 223:85–98. CrossRefGoogle Scholar
  22. Kandadi P, AfzalSyed M, Goparaboina S, Veerabrahma K (2012) Albumin coupled lipid nanoemulsions of diclofenac for targeted delivery to inflammation. Nanomed Nanotechnol Biol Med 8:1162–1171. CrossRefGoogle Scholar
  23. Khachane PV, Jain AS, Dhawan VV et al (2015) Cationic nanoemulsions as potential carriers for intracellular delivery. Saudi Pharm J 23:188–194. CrossRefGoogle Scholar
  24. Laxmi M, Bhardwaj A, Mehta S, Mehta A (2015) Development and characterization of nanoemulsion as carrier for the enhancement of bioavailability of artemether. Artif Cells Nanomedicine Biotechnol 43:334–344. CrossRefGoogle Scholar
  25. Lu W, Kelly AL, Miao S (2017) Bioaccessibility and cellular uptake of β-carotene encapsulated in model O/W emulsions: influence of initial droplet size and emulsifiers. Nanomater (Basel, Switzerland). Google Scholar
  26. Mao L, O’Kennedy BT, Roos YH et al (2012) Effect of monoglyceride self-assembled structure on emulsion properties and subsequent flavor release. Food Res Int 48:233–240. CrossRefGoogle Scholar
  27. McClements DJ (1995) Advances in the application of ultrasound in food analysis and processing. Trends Food Sci Technol 6:293–299. CrossRefGoogle Scholar
  28. Narsimhan G, Goell P (2001) Drop coalescence during emulsion formation in a high-pressure homogenizer for tetradecane-in-water emulsion stabilized by sodium dodecyl sulfate. J Colloid Interface Sci 238:420–432. CrossRefGoogle Scholar
  29. McClements DJ (2012) Nanoemulsions versus microemulsions: terminology, differences, and similarities. Soft Matter 8:1719–1729CrossRefGoogle Scholar
  30. Nicoli DF, Wu JS, Chang YJ et al (1995) Wide dynamic range particle size: analysis by DLS-SPOS: a combination of two technologies. Am Lab 27:41–49Google Scholar
  31. Nicoli DF, Hasapidis K, O’Hagan P et al (1998) High-resolution particle size analysis of mostly submicrometer dispersions and emulsions by simultaneous combination of dynamic light scattering and single-particle optical sensing. In: Provder T (ed) Particle size distribution III—assessment and characterization, chap 6. American Chemical Society, Washington, DC, pp 52–76CrossRefGoogle Scholar
  32. Niederquell A, Kuentz M (2013) Proposal of stability categories for nano-dispersions obtained from pharmaceutical self-emulsifying formulations. Int J Pharm 446:70–80. CrossRefGoogle Scholar
  33. Peng J, Dong W, Li L et al (2015) Effect of high-pressure homogenization preparation on mean globule size and large-diameter tail of oil-in-water injectable emulsions. J Food Drug Anal 23:828–835. CrossRefGoogle Scholar
  34. Perinelli DR, Cespi M, Pucciarelli S et al (2013) Effect of phosphate buffer on the micellisation process of Poloxamer 407: Microcalorimetry, acoustic spectroscopy and dynamic light scattering (DLS) studies. Colloids Surfaces A Physicochem Eng Asp 436:123–129. CrossRefGoogle Scholar
  35. Perinelli DR, Cespi M, Bonacucina G et al (2017) Heating treatments affect the thermal behaviour of doxorubicin loaded in PEGylated liposomes. Int J Pharm 534:81–88. CrossRefGoogle Scholar
  36. Qian C, McClements DJ (2011) Formation of nanoemulsions stabilized by model food-grade emulsifiers using high-pressure homogenization: Factors affecting particle size. Food Hydrocoll 25:1000–1008. CrossRefGoogle Scholar
  37. Schultz S, Wagner G, Urban K, Ulrich J (2004) High-pressure homogenization as a process for emulsion formation. Chem Eng Technol 27:361–368. CrossRefGoogle Scholar
  38. Shaha RB, Zidan AS, Funck T et al (2007) Quality by design: Characterization of self-nano-emulsified drug delivery systems (SNEDDs) using ultrasonic resonator technology. Int J Pharm 341:189–194. CrossRefGoogle Scholar
  39. Sharma S, Sahni JK, Ali J, Baboota S (2015) Effect of high-pressure homogenization on formulation of TPGS loaded nanoemulsion of rutin—pharmacodynamic and antioxidant studies. Drug Deliv 22:541–551. CrossRefGoogle Scholar
  40. Singh Y, Meher GJ, Raval K et al (2017) Nanoemulsion: Concepts, development and applications in drug delivery. J Control Release 252:28–49. CrossRefGoogle Scholar
  41. Stang M, Schuchmann H, Schubert H (2001) Emulsification in high-pressure homogenizers. Eng Life Sci 1:151–157.;2-D CrossRefGoogle Scholar
  42. Stillhart C, Kuentz M (2012) Comparison of high-resolution ultrasonic resonator technology and Raman spectroscopy as novel process analytical tools for drug quantification in self-emulsifying drug delivery systems. J Pharm Biomed Anal 59:29–37. CrossRefGoogle Scholar
  43. Stillhart C, Cavegn M, Kuentz M (2013) Study of drug supersaturation for rational early formulation screening of surfactant/co-solvent drug delivery systems. J Pharm Pharmacol 65:181–192. CrossRefGoogle Scholar
  44. Sun W, Xie C, Wang H, Hu Y (2004) Specific role of polysorbate 80 coating on the targeting of nanoparticles to the brain. Biomaterials 25:3065–3071. CrossRefGoogle Scholar
  45. Taylor TM, Davidson PM, Bruce BD, Weiss J (2005) Ultrasonic spectroscopy and differential scanning calorimetry of liposomal-encapsulated nisin. J Agric Food Chem 53:8722–8728. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Pharmaceutical SciencesUniversity of PerugiaPerugiaItaly
  2. 2.Janssen-Cilag s.p.a.LatinaItaly
  3. 3.School of PharmacyUniversity of CamerinoCamerinoItaly
  4. 4.QI s.r.l.RomeItaly

Personalised recommendations