(5by) Vesicle Transformations: Unilamellar to Multilamellar Structures

Vesicle Transformations:  The Whys and Hows of Multilamellar Structures

Vesicles are an extremely important class of materials, being the foundation for a wide variety of existing industrial products, as well as serving as vehicles for drug delivery, imaging and separations.  However, in spite of their ubiquitous applications and appearance in many products, much of the underlying product development has been empirical, and the basic principles that are necessary for improved product performance (e.g. longer shelf life), and for the development of whole new classes of materials and applications, are largely unexplored.   

Typically, for most vesicle-based products, there are at least two desirable properties:

(a)           There is a target pour viscosity for the product

(b)           The vesicles should not gravitationally phase separate from the suspending fluid over the shelf life of the product. 

These two properties are very intimately connected to the size, shape and lamellarity distributions of the vesicles.  It is recognized in the research community that these distributions depend on the following factors:

(i)                   The constituent lipids of the bilayer, and the weight fraction of the lipid in the solution

(ii)                 Any additives (such as salts, short chain alcohols etc.)

(iii)                The exact procedure by which the lipids are mixed with aqueous phase, including when the additives are added.   

Unfortunately, systematic investigations and reliable models connecting the above variables to the final size and lamellarity distributions are lacking, and the consequences of different surfactant combinations on vesicle formation and properties are understood only at a broad overview level.   In addition, there is no guarantee that once vesicles are prepared, their size and lamellarity distributions remain fixed.  Even if products are prepared with their initial distributions tailored to satisfy conditions (a) and (b), they may eventually evolve towards distributions that may compromise these requirements.  Therefore, the task of systematically achieving favorable vesicle suspensions can be broken down into understanding two major questions:

(A)          How do factors (i) to (iii) control whether a certain formulation and preparation procedure will yield unilamellar or multilamellar structures of certain sizes and shapes?

(B)          How and over what time scales do these structures evolve from being, say, unilamellar to multilamellar after formation?

The questions posed above represent an extremely large number of investigations.   We have chosen to begin with a small subset, namely the exploration of question (B) with the class of vesicles composed completely of positively charged bilayers.  Even though such surfactants are frequently used in fabric softeners, shampoos and conditioners, the number of studies on the vesicles constituted from such surfactants is surprisingly limited as compared to vesicles composed of neutral lipids or even negatively charged lipids.  Our recent observations show that these vesicle suspensions can undergo a transition over a several day period from an initial state of mainly unilamellar vesicles to a state of smaller bilamellar and multilamellar vesicles, without any added salt or other forcing, such as changes of temperature. Our hypothesis is that this transition could be driven by repulsive forces between excessively ?crowded? charged vesicles.  We also propose a mechanism driving this transformation ? unilamellar vesicles deflate, curl up and fuse at the tips to form bilamellar vesicles (see figure 1 below).   We were able to verify this mechanism by fabricating vesicles with a mixture of single and double tailed cationic lipids, which did not exhibit these transformations, presumably because the vesicles prepared

We are currently attempting to delineate the mechanism for the formation of multilamellar vesicles from unilamellar ones.  Since a mechanism involving curling and folding of the vesicle [and in fact, answering both questions (A) and (B)] would invariably depend on the properties of the bilayer membrane, one of the key components of our studies is the measurement of the membrane properties for vesicles made from lipid mixtures, which we are currently performing using micropipette aspiration of giant vesicles prepared from these mixtures.  We have recently performed measurements on giant vesicles prepared from a C18-double tailed cationic lipid, which is, to the best of our knowledge, the first measurement performed for vesicles composed fully of charged lipids.  We also intend to develop a theoretical description of vesicle folding and other processes, both from a molecular and a continuum perspective.

The research outlined here is extremely interesting from a fundamental point of view. For example, the transformation observed in our and prior experiments is reminiscent of the process of endocytosis, whereby a cell ingests some volume of extracellular fluid by initiating an invagination on the cell wall, followed by pinch-off. Further, we anticipate that this work will enable a number of important technological applications:

(i)                   The phenomenon may be utilized as a procedure to encapsulate drugs or other materials within a bilamellar (or multilamellar) vesicle.  Bilamellar vesicles are expected to have longer lifetimes in vivo than unilamellar vesicles, and this is a key property for drug delivery formulations.

(ii)                 Evaporation of the suspending fluid (or other ways of increasing the volume fraction of vesicles) may lead to vesicle transformations, from unilamellar to multilamellar, resulting in a reduction in the average size and vesicle volume fraction, which may be unintended and may have unfavorable consequences.

(iii)                Alternatively, this phenomenon may be utilized as a way of manipulating the vesicle structure without the need to introduce salt or other ?additive? sources of osmotic pressure that can reduce the volume of vesicles and potentially drive similar transitions.

It should be emphasized that the changes in density and the volumetric concentration of vesicles caused by a transition from unilamellar to multilamellar structures will also play an important role in the development of gravitationally-induced phase separation (sedimentation or creaming), which is a critical issue for many of the suspension-based commercial products.

The results from our experiments and future directions will be discussed in greater detail in the poster.

 SHAPE  \* MERGEFORMAT

Figure 1.               Proposed mechanism for bilamellar vesicle formation.