NEWS

Liposomes have been used as an effective means of delivery allowing for more efficient absorption and delivery of both hydrophilic and lipophilic substances and greater protection against oxidation and degradation. Liposomes are lipid vesicles made of phospholipids strung together to form a double membrane. A structure arranged in this way is also present in the cell membranes of the human body. Regardless of size or structure, liposomes can enclose different substances including botanical extracts and nutraceutical ingredients.
The liposomal shield protects the transported substances from the body’s own enzymes in the digestive tract and also from gastric acid. When taking conventional pills and powders, the absorption of the active ingredient is often impaired which results in low bioavailability of the supplement.
Liposomes act like a protective shield that wraps around the botanical ingredient and or nutraceutical ingredient. Thus, the active ingredient does not come into contact with gastric acid and consequently its absorption is not restricted. The active ingredient is protected in a double membrane made of phospholipids. Liposomes can fuse directly with human cell membranes, as they are structurally identical.
Liposomes brought a very interesting proposition into lipid deliveries: Instead of a surfactant stabilizing a drop of oil in water, the phospholipids arranged into spherical cell membrane–like lipid bilayers with their water-loving “heads” toward the aqueous medium and their fatty “tails” tucked toward each other. These structures created pockets of entrapped water along with water soluble compounds and the potential to protect them from hostile digestive conditions and potentially facilitating gastrointestinal (GI) uptake. At the same time, the hydrophobic fatty acid core of the bilayers could host hydrophobic compounds, creating a small spherical package that could carry both hydrophilic and hydrophobic compounds. Interactions between the liposome membrane and cell membranes further support the deal and offered a promise of enhanced cellular uptake through endosomal mechanisms.

Advantages of Liposomal Delivery

· High bioavailability and absorption compared with other oral forms of supplements.
· Micronized encapsulation protects against the harsh environment of the GI tract and increases transmucosal (oral) uptake and absorption.
· Increased intracellular delivery.
· Can hold both hydrophilic and hydrophobic compounds.
· Allow for adjustable and incremental dosing for children and adults.
· Cost effective by being able to take a lower dose for the same effect.

Imaging of Liposomes by Transmission Electron Microscopy (TEM)

TEM is a high-resolution imaging technique to evaluate morphology and architecture of liposomes. TEM is an important method for the characterization of size and shape of nanoparticles as it can directly visualize single particles and even their inner architecture. The best method to visualize liposomes close to their native structure is cryo-electron microscopy, where thin films of suspensions are plunge frozen to create vitrified ice films that can be imaged directly in the electron microscope under liquid nitrogen temperature. Although subject to artifacts, negative staining TEM can also be a useful method to image liposomes, as it is faster and simpler than cryo-EM, and requires less advanced equipment.



Few examples of botanical ingredients and nutraceuticals with liposomal delivery

· Liposomal Glutathione

Glutathione is the master antioxidant in the human body. Structurally, glutathione is a tripeptide made up of cysteine, glycine and glutamic acids. There are two forms of glutathione as oxidized and reduced forms. Reduced form is the active form and potent antioxidant. The problem with reduced glutathione is it gets depleted by stomach acids. Hence, its absorption is pretty low. Therefore, alternative forms such as liposomal glutathione is the best solution.

Liposomal glutathione is an active form of glutathione. It exists encapsulated inside a lipid molecule in order to enhance absorption. The encapsulation process protects the glutathione molecule from stomach acids. It also increases the absorption by up to 80%.

· Liposomal Boswellia

Boswellic acids (BAs) are isolated from oleo gum of Boswellia serrata and are mainly used as potential anti-inflammatory, hypolipidemic and immunomodulatory. Pharmacokinetic investigations of BAs uncover its poor bioavailability through digestive system thus creates a need for improved therapeutic responses which can possibly be achieved by developing formulations through novel delivery system. liposomes have been found wide application in ameliorating BAs bioavailability and efficacy; indeed, various modifications of liposomal BAs have been developed such as polymeric conjugation on the liposome surface to acquire better clinical outcomes. BAs liposome formulations are known to improve bioavailability and efficacy.

· Multivitamin/mineral (MVM) supplements

Multivitamin/mineral (MVM) supplements have been consumed since the early 1940s and remain popular today. National Health and Nutrition Examination Survey (NHANES) data indicate that MVM products are consumed more frequently than any other type of dietary supplement, and that the proportion of the population consuming MVM products increases with age. A particularly notable development influencing the physical form of products and potentially altering the bioavailability of nutrients within MVMs is the introduction of liposomal delivery mechanisms, in which nutrients are packaged in liposomes to promote enhanced absorption and bioavailability. Several investigations support the ability of liposomal packaging to improve vitamin absorption. However, few studies have examined the influence of liposomal packaging on mineral absorption, particularly within the context of an MVM product.

Future Use in Nutraceutical Industry

Companies continue to innovate beyond ingredients by making their offerings more bioavailable and increasing utilization of nutrients by the body. There is no doubt that nutritional industry use of liposomes will grow rapidly in the next 5 to 10 years. Most potential negatives revolve around industry adoption of reproducible, high-quality production techniques and testing methods for characterizing size, stability, and efficacy. As this hurdle is overcome, we will see exciting high-quality products with more complex mixtures of pure compounds and complex botanical mixtures.

In recent years, several events have been organized by various researchers / industry stakeholders in order to consolidate their presence in this field and enhance their existing capabilities to meet the growing demand for these novel delivery systems. It is worth mentioning that majority of the events were focused on discussing the analytical method development, manufacturing and process development steps for liposomes.

The manufacturing of highly potent liposome-based botanicals and nutraceuticals requires an adequate working environment, stringent manufacturing protocols (to comply with the established regulatory standards) and a trained workforce (to satisfactorily handle highly potent materials).

Liposomes are a form of nanocarriers that have been widely investigated for drug-delivery purposes. Liposomes have become widely used as pharmaceutical delivery carriers in numerous clinical applications. The increase in their applications has facilitated the development of analytical and technological approaches to determine their characteristics. They are composed of phospholipid bilayers which enclose a distinct aqueous space, thereby allowing encapsulation of both hydrophilic and hydrophobic compounds. The most routinely investigated parameters in liposome characterization include vesicle size and size distribution (or polydispersity), surface charge (or Zeta potential), shape and morphology, lamellarity, encapsulation efficiency, phase behaviour (or polymorphism) and in vitro release profile. A single analytical technique cannot be used to investigate the above parameters and the choice always relies on what exactly one is aiming at. Visualization analysis is an essential and powerful way for morphology studies, which allows intuitively imaging biological complex system at the molecular level. Through visualization analysis, a variety of useful data will be collected, including size distribution, bilayer organization, vesicles shape and number of lamellae. Liposome visualization data provided by microscopic techniques becomes prerequisite parameters guide the utilization of liposome for drug encapsulation.



Transmission Electron Microscopy (TEM) is an important method for the characterization of size and shape of nanoparticles as it can directly visualize single particles and even their inner architecture. The best method to visualize liposomes close to their native structure is cryo-electron microscopy, where thin films of suspensions are plunge frozen to create vitrified ice films that can be imaged directly in the electron microscope under liquid nitrogen temperature. Although subject to artifacts, negative staining TEM can also be a useful method to image liposomes, as it is faster and simpler than cryo-EM, and requires less advanced equipment. When it comes to TEM, we can understand the particle size and shape, where with cryo-TEM along with the above lamellarity can also be evaluated.

The most frequently used TEM sample preparation techniques for liposomes were drying and staining.

Drying: Drying including freeze drying is the most readily accessible and widely applied method to vesicles. Typically, a drop of 2–5 μl sample is applied onto a carbon or polymer coated grid and dried for a few minutes upto several hours before imaging.

Although the most widely applied, drying is also the riskiest technique for soft materials as sometimes it aggregates upon drying and cause drying patterns, crystals and other high contrast structures to be visible. After drying, it is no longer possible to conclude whether the aggregates in the image are the result of aggregation behaviour of the sample in solution or of the sample upon drying. Even though samples exist that look the same in solution as after drying, the method of drying does not allow this distinction between what structure resulted from the sample and what structure is the result of drying.

Negative staining: Staining is better suitable for soft matter than drying. In order to preserve a sample and enhance contrast, one can use heavy metals to stain the sample. A positive stain, like iodine, ruthenium and osmium tetra oxide is a strong scattering agent that adheres to particular areas of the sample. A positive stain can change dimensions such as membrane thickness. A negative stain does not penetrate the object, but coats the surface and surroundings, obscuring the object itself and all internal structural details, and giving a foot-print like appearance. The most popular stains in soft matter are uranyl acetate (UAc) and phosphotungstic acid (PTA). Negative stain is applied to the grid straight after the sample was blotted off to prevent dehydration and interactions between the sample and the support grid. Even though the structure is much better preserved upon staining than with drying, upon staining, the sample is also dehydrated, which can cause deformation of solvent dependent structures such as lipid vesicles.

Read More...