Microscopy belongs to the group of tests, used in pharmaceutical technology, that despite the lapse of time and the development of new analytical methods, still remain irreplaceable for the characterization of dispersed drug dosage forms (e.g., suspensions and emulsions). To obtain complete description of a specific drug formulation, such as parenteral colloidal products, a combination of different microscopic techniques is sometimes required. Electron microscopy methods are the most useful ones; however, even such basic methods as optical microscopy may be helpful for determination of some properties of a sample. The publication explicates the most popular microscopical techniques used nowadays for characterization of the morphology of nanoparticles suspended in pharmaceutical formulations; ad vantages and disadvantages of these methods are also discussed. Parenteral submicron formulations containing lecithin or a particular phospholipid were chosen as examples.
1. P. Couvreur, Nanoparticles in drug delivery: past, present and future, Adv. Drug Deliv. Rev. 65 (2013) 21–23; DOI: 10.1016/j.addr.2012.04.010.
2. I. F. Uchegbu and A. Siew, Nanomedicines and nanodiagnostics come of age, J. Pharm. Sci. 102 (2013) 305–310; DOI: 10.1002/jps.23377.
3. T. M. Allen and P. R. Cullis, Liposomal drug delivery systems: from concept to clinical applications, Adv. Drug Deliv. Rev. 65 (2013) 36–48; DOI: 10.1016/j.addr.2012.09.037.
4. E. Fattal and Ch. Vauthier, Drug Delivery: Nanoparticles, in Encyclopedia of Pharmaceutical Technology (Ed. J. Swarbrick), Informa Healthcare, New York 2007, pp. 1183–1200.
5. N. Anton and T. F. Vandamme, Nano-emulsions and micro-emulsions: clarification of the critical differences, Pharm. Res. 28 (2011) 978–985; DOI: 10.1007/s11095-010-0309-1.
6. C. R. Rowe, P. J. Sheskey and S. C. Owen, Handbook of Pharmaceutical Excipients, 5th ed., Pharmaceutical Press, London 2006, pp. 409–411.
7. A. Michajlik and E. Bartnikowska, Lipidy i lipoproteiny osocza, PZWL, Warsaw 1999.
8. W. Bernhard, S. Hoffmann, H. Dombrovsky, G. A. Rau, A. Kamlage, M. Kappler, J. J. Haitsma, J. Freihorst, H. von der Hardt and C. F. Poets, Phosphatidylcholine molecular species in lung surfactant: composition in relation to respiratory rate and lung development, Am. J. Respir. Cell Mol. Biol. 25 (2001) 725–731; DOI: 10.1165/ajrcmb.25.6.4616.
9. S. Tamilvanan, Oil-in-water emulsions: implications for parenteral and ocular delivering systems, Prog. Lipid Res. 43 (2004) 489–533; DOI: 10.1016/j.plipres.2004.09.001.
10. N. A. Mazer, G. B. Benedek and M. C. Carey, Quasi elastic light-scattering studies of aqueous biliary lipid systems. Mixed micelle formation in bile salt-lecithin solutions, Biochemistry19 (1980) 601–615; DOI: 10.1021/bi00545a001.
11. W. Mehnert and K. Mäder, Solid lipid nanoparticles: production, characterization and applications, Adv. Drug Deliv. Rev. 64 (2012) 83–101; DOI: 10.1016/j.addr.2012.09.021.
12. S. A. Wissing, O. Kayser and R. H. Müller, Solid lipid nanoparticles for parenteral drug delivery, Adv. Drug Deliv. Rev. 56 (2004) 1257–1272; DOI: 10.1016/j.addr.2003.12.002.
13. M. Brandl, Vesicular phospholipid gels: a technology platform, J. Liposome Res. 17 (2007) 15–26; DOI: 10.1080/08982100601186490.
14. M. Brandl, M. Drechsler, D. Bachmann, C. Tardi, M. Schmidtgen and K. H. Bauer, Preparation and characterization of semi-solid phospholipid dispersions and dilutions thereof, Int. J. Pharm. 170 (1998) 187–199; DOI: 10.1016/S0378-5173(98)00146-X.
15. C. Tardi, M. Drechsler, K. H. Bauer and M. Brandl, Steam sterilization of vesicular phospholipid gels, Int. J. Pharm. 217 (2001) 161–172; DOI: 10.1016/S0378-5173(01)00605-6.
16. M. Sznitowska, E. A. Dąbrowska and S. Janicki, Solubilizing potential of submicron emulsions and aqueous dispersions of lecithin, Int. J. Pharm. 246 (2002) 203–206; DOI: 10.1016/S0378-5173(02)00395-2.
17. M. Sznitowska, M. Klunder and M. Płaczek, Paclitaxel solubility in aqueous dispersions and mixed micellar solutions of lecithin, Chem. Pharm. Bull. 56 (2008) 70–74; DOI: 10.1248/cpb.56.70.
18. M. Sznitowska, M. Bodnar, J. Petrusewicz, H. Janik and E. A. Dąbrowska, Preliminary in vivo studies of a new lecithin-based formulation of paclitaxel, J. Microencapsul. 26 (2009) 588–592; DOI: 10.3109/02652040802586068.
19. R. J. Haskell, Characterization of submicron systems via optical methods, J. Pharm. Sci. 87 (1998) 125–129; DOI: 10.1021/js970331i.
20. V. Klang, N. B. Matsko, C. Valenta and F. Hofer, Electron microscopy of nanoemulsions: an essential tool for characterisation and stability assessment, Micron43 (2012) 85–103; DOI: 10.1016/j.micron.2011.07.014.
21. J. Kuntsche, J. C. Horst and H. Bunjes, Cryogenic transmission electron microscopy (cryo-TEM) for studying the morphology of colloidal drug delivery systems, Int. J. Pharm.417 (2011) 120–137; DOI: 10.1016/j.ijpharm.2011.02.001.
22. S. Bibi, R. Kaur, M. Henriksen-Lacey, S. E. McNeil, J. Wilkhu, E. Lattmann, D. Christensen, A. R. Mohammed and Y. Perrie, Microscopy imaging of liposomes: from coverslips to environmental SEM, Int. J. Pharm. 417 (2011) 138–150; DOI: 10.1016/j.ijpharm.2010.12.021.
23. J. A. Litwin and M. Gajda, Podstawy technik mikroskopowych, Jagiellonian University Press, Cracow 2011.
24. P. E. West, Introduction to Atomic Force Microscopy, Pacific Nanotechnology, Santa Clara 2006, pp. 1–16.
25. P. C. Schmidt, Secondary Electron Microscopy in Pharmaceutical Technology, in Encyclopedia of Pharmaceutical Technology (Ed. J. Swarbrick), Informa Healthcare, New York 2007, pp. 3217–3256.
26. K. Akashi, H. Miyata, H. Itoh and K. Kinosita, Formation of giant liposomes promoted by divalent cations: critical role of electrostatic repulsion. Biophys. J. 74 (1998) 2973–2982; DOI: 10.1016/S0006-3495(98)78004-X.
27. R. M. Fernandez, K. A. Riske, L. Q. Amaral, R. Itri and M. T. Lamy, Influence of salt on the structure of DMPG studied by SAXS and optical microscopy, Biochim. Biophys. Acta. 1778 (2008) 907–916; DOI: 10.1016/j.bbamem.2007.12.005.
28. C. C. Müller-Goymann, Physicochemical characterization of colloidal drug delivery systems such as reverse micelles, vesicles, liquid crystals and nanoparticles for topical administration, Eur. J. Pharm. Biopharm. 58 (2004) 343–356; DOI: 10.1016/j.ejpb.2004.03.028.
29. C. C. Müller-Goymann, Drug Delivery: Liquid Crystals, in Encyclopedia of Pharmaceutical Technology (Ed. J. Swarbrick), Informa Healthcare, New York 2007, pp. 1115–1131.
30. A. Graf, E. Ablinger, S. Peters, A. Zimmer, S. Hook and T. Rades, Microemulsions containing lecithin and sugar-based surfactants: nanoparticle templates for delivery of proteins and peptides, Int. J. Pharm. 350 (2008) 351–360; DOI: 10.1016/j.ijpharm.2007.08.053.
31. N. Rodriguez, F. Pincet and S. Cribier, Giant vesicles formed by gentle hydration and electroformation: a comparison by fluorescence microscopy, Colloids Surf. B. Biointerfaces42 (2005) 125–130; DOI: 10.1016/j.colsurfb.2005.01.010.
32. P. Arunothayanun, M. S. Bernard, D. Q. M. Craig, I. F. Uchegbu and A. T. Florence, The effect of processing variables on the physical characteristics of non-ionic surfactant vesicles (niosomes) formed from a hexadecyl diglycerol ether, Int. J. Pharm. 201 (2000) 7–14; DOI: 10.1016/S0378-5173(00)00362-8.
33. S. R. Pygall, J. Whetstone, P. Timmins and C. D. Melia, Pharmaceutical applications of confocal laser scanning microscopy: the physical characterization of pharmaceutical systems, Adv. Drug Deliv. Rev. 59 (2007) 1434–1452; DOI: 10.1016/j.addr.2007.06.018.
34. J. Ch. Colas, W. Shi, V. S. Rao, A. Omri, M. R. Mozafari and H. Singh, Microscopical investigations of nisin-loaded nanoliposomes prepared by Mozafari method and their bacterial targeting, Micron38 (2007) 841–847; DOI: 10.1016/j.micron.2007.06.013.
35. B. Ruozi, D. Belletti, A. Tombesi, G. Tosi, L. Bondioli, F. Forni and M. A. Vandelli, AFM, ESEM, TEM, and CLSM in liposomal characterization: a comparative study, Int. J. Nanomedicine6 (2011) 557–563; DOI: 10.2147/IJN.S14615.
36. M. A. Schubert and C. C. Müller-Goymann, Characterisation of surface-modified solid lipid nanoparticles (SLN): influence of lecithin and nonionic emulsifier, Eur. J. Pharm. Biopharm. 61 (2005) 77–86; DOI: 10.1016/j.ejpb.2005.03.006.
37. H. Zhou, Y. Yue, G. Liu, Y. Li, J. Zhang, Q. Gong, Z. Yan and M. Duan, Preparation and characterization of a lecithin nanoemulsion as a topical delivery system, Nanoscale Res. Lett. 5 (2010) 224–230; DOI: 10.1007/s11671-009-9469-5.
38. S. A. Abraham, K. Edwards, G. Karlsson, S. MacIntosh, L. D. Mayer, C. McKenzie and M. B. Bally, Formation of transition metal-doxorubicin complexes inside liposomes, Biochim. Biophys. Acta1565 (2002) 41–54; DOI: 10.1016/S0005-2736(02)00507-2.
39. M. Ciobanu, B. Heurtault, P. Schultz, C. Ruhlmann, C. D. Muller and B. Frisch, Layersome: development and optimization of stable liposomes as drug delivery system, Int. J. Pharm. 344 (2007) 54–57; DOI: 10.1016/j.ijpharm.2007.05.037.
40. H. Teixeira, C. Dubernet, V. Rosilio, S. Benita, J. Lepault, I. Erk and P. Couvreur, New bicompartmental structures are observed when stearylamine is mixed with triglyceride emulsions, Pharm. Res. 17 (2000) 1329–1332; DOI: 10.1023/A:1026416208482.
41. K. Jores, W. Mehnert, M. Drechsler, H. Bunjes, C. Johann and K. Mäder, Investigations on the structure of solid lipid nanoparticles (SLN) and oil-loaded solid lipid nanoparticles by photon correlation spectroscopy, field-flow fractionation and transmission electron microscopy, J. Control. Release95 (2004) 217–227; DOI: 10.1016/j.jconrel.2003.11.012
42. A. Graf, E. Ablinger, S. Peters, A. Zimmer, S. Hook and T. Rades, Microemulsions containing lecithin and sugar-based surfactants: nanoparticle templates for delivery of proteins and peptides, Int. J. Pharm. 350 (2008) 351–360; DOI: 10.1016/j.ijpharm.2007.08.053.
43. G. De Rosa, M. De Stefano, F. Ungaro and M. I. La Rotonda, Cold field emission gun-scanning electron microscopy: a new tool for morphological and ultrastructural analysis of liposomes, Int. J. Pharm. 362 (2008) 189–192; DOI: 10.1016/j.ijpharm.2008.06.003.
44. A. Saupe, K. C. Gordon and T. Rades, Structural investigations on nanoemulsions, solid lipid nanoparticles and nanostructured lipid carriers by cryo-field emission scanning electron microscopy and Raman spectroscopy, Int. J. Pharm. 314 (2006) 56–62; DOI: 10.1016/j.ijpharm.2006.01.022.
45. N. G. Eskandar, S. Simovic and C. A. Prestidge, Nanoparticle coated submicron emulsions: sustained in-vitro release and improved dermal delivery of all-trans-retinol, Pharm. Res. 26 (2009) 1764–1775; DOI: 10.1007/s11095-009-9888-0.
46. Y. Perrie, A. U. Mohammed, A. Vangala and S. E. McNeil, Environmental scanning electron microscopy offers real-time morphological analysis of liposomes and niosomes, J. Liposome Res. 17 (2007) 27–37; DOI: 10.1080/08982100601186508.
47. A. Bogner, G. Thollet, D. Basset, P. H. Jouneau and C. Gauthier, Wet STEM: a new development in environmental SEM for imaging nano-objects included in a liquid phase, Ultramicroscopy104 (2005) 290–301; DOI: 10.1016/j.ultramic.2005.05.005.
48. J. Sitterberg, A. Özcetin, C. Ehrhardt and U. Bakowsky, Utilising atomic force microscopy for the characterisation of nanoscale drug delivery systems, Eur. J. Pharm. Biopharm.74 (2010) 2–13; DOI: 10.1016/j.ejpb.2009.09.005.
49. B. Ruozi, G. Tosi, E. Leo and M. A. Vandelli, Application of atomic force microscopy to characterize liposomes as drug and gene carriers, Talanta73 (2007) 12–22; DOI: 10.1016/j.talanta.2007.03.031.
50. X. Liang, G. Mao and K. Y. Ng, Mechanical properties and stability measurement of cholesterol-containing liposome on mica by atomic force microscopy, J. Colloid Interface Sci. 278 (2004) 53–62; DOI: 10.1016/j.jcis.2004.05.042.
51. C. Preetz, A. Hauser, G. Hause, A. Kramer and K. Mäder, Application of atomic force microscopy and ultrasonic resonator technology on nanoscale: distinction of nanoemulsions from nanocapsules, Eur. J. Pharm. Sci. 39 (2010) 141–151; DOI: 10.1016/j.ejps.2009.11.009.
52. T. Tran, T. C. Kupiec and L. A. Trissel, Quality-control analytical methods: particulate matter in injections: what is it and what are the concerns?, Int. J. Pharm. Compd.10 (2006) 202–204.
53. S. E. Langille, Particulate matter in injectable drug products, PDA J. Pharm. Sci. Technol.67 (2013) 186–200; DOI: 10.5731/pdajpst.2013.00922.
54. Ch. M. Hoo, N. Starostin, P. West and M. L. Mecartney, A comparison of atomic force microscopy (AFM) and dynamic light scattering (DLS) methods to characterize nanoparticle size distributions, J. Nanopart. Res.10 (2008) 89–96; DOI: 10.1007/s11051-008-9435-7.
55. H. Kato, A. Nakamura and N. Noda, Determination of size distribution of silica nanoparticles: a comparison of scanning electron microscopy, dynamic light scattering, and flow-field-flow fractionation with multiangle light scattering methods, Mater. Express4 (2014) 144–152; DOI: 10.1166/mex.2014.1150.
56. V. Klang, C. Valenta and N. B. Matsko, Electron microscopy of pharmaceutical systems, Micron44 (2013) 45–74; DOI: 10.1016/j.micron.2012.07.008.