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Morphology and Ultrastructure

Along with particle size, dispersity, and composition, the morphology and ultrastructure are important properties of nanoparticles and their formulations which can control properties such as encapsulation efficiency. Morphology generally relates to the exterior of the particle and may be characterized by shape and surface structure whilst ultrastructure generally relates to the interior of the particle and can relate to internal partitioning through, for example, a core-shell structure.

The ultrastructure of systems should be given due to consideration in predicting the organization of encapsulated drug components within the dispersions. The performance of lipid nanoparticles, drug incorporation, release, and stability are always influenced by particle size, shape, and structure. So it is necessary to investigate the morphology of lipid nanoparticles.

  • Transmission Electron Microscopy (TEM)

TEM is often used to evaluate the morphology and ultrastructure of colloidal carrier systems. Procedures such as negative staining, freeze-fracture, and vitrification by plunge freezing are required for the preparation of samples to be evaluated by TEM. Negative staining employing solutions of heavy metal salts is the most commonly used method to demonstrate the presence of colloidal lipid particles in dispersions by TEM.

Staining techniques are quick and easy to use. Sample preparation does not require any special equipment. However, staining and drying of samples may lead to artifacts that are difficult to distinguish from real colloidal structures.

Fig.1 Transmission electron micrographs of nanostructured lipid carriers (a) and solid lipid nanoparticles (b) after staining with 1 % uranyl acetate solution. (Shah, 2015)

The freeze-fracture technique provides additional information on the ultrastructure of lipid nanoparticle dispersions. This technique has been used to distinguish different colloidal structures such as solid lipid particles, emulsions, and liposomes. The influence of polymorphic transitions on particle structure and morphology has also been shown with this technique.

Freeze-fracture TEM of nanostructured lipid carriers prepared by high-pressure homogenization. Fig.2 Freeze-fracture TEM of nanostructured lipid carriers prepared by high-pressure homogenization. (Shah, 2015)

In contrast to the negative staining and freeze-fracture techniques, cryo-TEM allows direct visualization of vitrified, frozen-hydrated samples without additional preparation. Cryo-TEM is suitable for the investigation of the coexistence of different colloidal carriers such as lipid nanoparticles, vesicles, and anisometric particles in the dispersion. Cryo-TEM was a useful tool for understanding the ultrastructure of nanoemulsions, SLNs, and NLCs.

Cryo-TEM images of nanoemulsions (a), solid lipid nanoparticles (b), and nanostructured lipid carriers (c and d). Fig.3 Cryo-TEM images of nanoemulsions (a), solid lipid nanoparticles (b), and nanostructured lipid carriers (c and d). (Shah, 2015)

  • Scanning Electron Microscopy (SEM)

SEM is the next most commonly used technique for imaging of small particles, however, it is very rarely used to measure the particle size of SLNs. It is more commonly used to investigate particle morphology. SEM imaging allows observation of SLNs in the absence of aggregation.

A major drawback to the technique is that the sample preparation procedures, specifically the application of vacuum, may affect the nature of the particles, leading to uncertainty in the experimental observations.

SEM images of unloaded (a) and praziquantel-loaded (b) solid lipid nanoparticles. Fig.4 SEM images of unloaded (a) and praziquantel-loaded (b) solid lipid nanoparticles. (Shah, 2015)

Cryo-field emission SEM (cryo-FESEM) is another imaging technique used to investigate the ultrastructure of solid dispersions. Solid particles can be viewed in their natural state (dispersed in water) by cryo-FESEM.

FESEM image of lipid nanoparticles. Fig.5 FESEM image of lipid nanoparticles. (Shah, 2015)

  • Atomic Force Microscopy (AFM)

AFM has been commonly used to investigate the morphology of lipid nanoparticles. AFM provides a high-resolution image of the particle surface and is an important characterization tool for particulate or biological samples as it allows imaging under hydrated conditions. The major advantage of this particle characterizing technique is that it requires no sophisticated sample preparation.

Creative Biolabs offers well-established and innovative one-stop-shop solutions for the evaluation of morphology and ultrastructure. Our professional staff will find a way to meet your unique needs. Please contact us for more information and a detailed quote.

Reference

  1. Shah, R.; et al. Lipid nanoparticles: Production, characterization and stability. New York, NY, USA:: Springer International Publishing. 2015.
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