Quality Control Of Liposomes 1

As a drug carrier, liposome not only improves the in vivo behavior of the drug, improves the therapeutic index of the drug, reduces the toxicity of the drug, and reduces the adverse reactions of the drug, and has a certain slow-release and controlled-release effect, reduces the dose or frequency of drug administration, and improves patient compliance . According to the characteristics of liposome formulations, the stability of liposomes has always been a concern of researchers. Liposomes are easy to aggregate, drugs are easy to leak (decrease in encapsulation rate) and oxidative deterioration will directly affect the safety and effectiveness of liposome drugs, so the quality of liposome drugs should be studied and controlled in detail.

Liposome Morphology Control
The structure of the liposome is that the two hydrophobic chains of the phospholipid molecule point to the inside one by one, and the polar heads are on the inner and outer surfaces of the membrane. The phospholipid bilayer constitutes an enclosed compartment, which contains an aqueous solution, and the phospholipid vesicles are separated by an aqueous medium. The liposome may be a single-layer closed bilayer structure or a multi-layer closed bilayer structure. Under the electron microscope, in addition to the common spherical and ellipsoid shapes, liposomes also have long tubular structures with diameters ranging from tens of nanometers to several microns.

Scanning or transmission electron microscopy is the main observation tool for nanoparticles. However, scanning electron microscopy (Scanning electron micro-scope, SEM) needs to dry the object, which will destroy the liposome structure, so it is generally not used for the observation of liposome morphology. Transmission electron microscope (TEM) can be divided into negative dye transmission electron microscope, cryo-electron microscope (CEM) and freeze fracture electron micro-scope (FFEM). The electron microscope uses high-energy electrons to penetrate the object to obtain the image, and the electron flow cannot penetrate the heavy metal elements, so it has a dark background and is called “negative staining”. The negative staining method uses a heavy metal ion solution (such as sodium phosphotungstate and uranium acetate) to negatively stain the liposome surface, so that the surrounding environment is a dark background and the object (liposome) is a bright background. In general, different types of heavy metal ion solutions are selected according to the electrical properties of liposomes. For example, negatively charged liposomes are stained with sodium phosphotungstate, because phosphotungstate is negative ions, and it generates electrostatic repulsion with liposomes, resulting in a negative staining effect. Conversely, if the liposome is positively charged, uranium acetate can be selected for negative staining. If it is stained, the entire area may be a dark background, making it difficult to distinguish the object from the background. Cryogenic electron microscopy requires observation by immersing liposomes in liquid ethane, and the membrane structure can be seen more clearly. Freeze etching is to freeze and fix the biological material under high vacuum and low temperature conditions, then evaporate a very thin layer of platinum at a certain angle on the fracture surface, vertically evaporate a layer of carbon to form a platinum carbon complex, and finally clean the replica and observe under the electron microscope. The cryoetch microscope can clearly see the structure and size of liposomes. Laser confocal microscopy can also be used to observe larger vesicles.

Liposome Size

From a security perspective. The particle size has a significant effect on the drug efficacy of the product, and the particle size requirements of specific products need to be controlled according to the needs of the drug to function. The size and uniformity of liposome size are closely related to their encapsulation efficiency and stability, which directly affects the liposome behavior in vivo.

The methods for measuring particle size mainly include microscope method, Coulter counting method, laser scattering method, centrifugal sedimentation method and microporous filter-absorbance method, and the currently common accepted method is dynamic light scattering method (also known as photon correlation spectroscopy). Because laser has the characteristics of good monochromaticity and non-divergence, laser is generally used as the light source, which is called laser dynamic light scattering method. The dynamic light scattering (Dynamic Light Scattering, DLS) technology obtains the particle size of the particles mixed in it by studying the phenomenon that the scattered light intensity fluctuates with time at a fixed spatial position. This technology has the advantages of short detection time and low cost.