Characteristics of amorphous matrices composed of different types of sugars in encapsulating emulsion oil droplets during freeze-drying

Abstract

The encapsulation of emulsion oil droplets by amorphous sugar matrices, formed by freeze-drying, was investigated, with a focus on the influence of the type of sugar. An oil-in-water emulsion, comprised of linoleic acid methyl ester (LME) and sucrose monolaurate (SML) as an oil phase and surfactant, respectively, were freeze-dried in the presence of different types of sugars. LME-droplet encapsulation during and after freeze-drying were evaluated by FTIR analysis. The loss of LME largely occurred in the early stage of freeze-drying. The size distribution of the encapsulated LME droplets remained unchanged before and after freeze-drying in most cases. The encapsulated fractions of LME droplets could be correlated with the glass transition temperature of the sugars in the fully hydrated state (T_g*), and the existence of an optimum T_g* value for the sugar matrix was predicted. The encapsulation ability of an amorphous sugar matrix was maximized when mono- and polysaccharide were combined so as to give a value for T_g* of approximately − 50 °C, although, individually, mono- and polysaccharides were quite poor for oil droplet encapsulation. These findings suggest that the structural flexibility of the amorphous sugar matrix is a major determinant in oil droplet encapsulation by an amorphous sugar matrix during freeze-drying.

Introduction

When an aqueous solution containing a sugar is dehydrated under appropriate conditions, the sugar molecules are solidified in the amorphous state. Amorphous sugar solids are frequently used as stabilizers and/or encapsulation agents for various substances and particulates. The stabilization of a protein by an amorphous sugar matrix during dehydration and storage has been extensively investigated (Manning et al., 1989, Pikal, 1994). Carpenter, Prestrelski, Anchodorquy, and Arakawa (1994) studied the mechanism by which a protein is stabilized by an amorphous sugar matrix, and concluded that hydrogen bonding between the protein and the surrounding sugar molecules played a major role: hydroxyl groups of sugar molecules form hydrogen bonds with protein molecules that are embedded in the dried sugar matrix, which substitute for water molecules that had served to maintain the intact conformations of a protein (Carpenter and Crowe, 1988, Carpenter and Crowe, 1989). It has also been proposed that the high viscosity of the glassy state of an amorphous sugar matrix aids in maintaining proteins in the folded state, i.e., it prevents unfolding (Franks et al., 1991, Slade and Levine, 1991). Furthermore, various types of amorphous sugars have been compared for their protein-stabilizing effects (Imamura et al., 2003, Izutsu et al., 1991, Prestrelski et al., 1995). It is well known that the smaller sized sugars tend to exert greater protein-stabilizing effects but form a physically more unstable amorphous matrix (Allison et al., 2000, Imamura et al., 2003, Prestrelski et al., 1995).

An amorphous sugar matrix is effective in embedding and preserving larger sized particulates such as cells (Crowe et al., 1987, Crowe et al., 1984). When disaccharides and certain types of polysaccharides are present during the dehydration of a cell suspension, cell-viability is increased (Kurosawa et al., 1997, Wessman et al., 2011). A virus also can be dehydrated in conjunction with a sucrose-based matrix, without the loss of their infectivity and transduction ability (Chen, Liu, Kost, & Chao, 2012).

Lipids also can be embedded in an amorphous sugar matrix when they are emulsified and dehydrated in the presence of a sugar (Desai and Park, 2005, Jonsdottir et al., 2005). This enables the long-term preservation of the dispersion state of oil droplets, since oil droplets are generally susceptible to fusion and aggregation during storage in the emulsion form (Drusch et al., 2006, Shimada et al., 1991). The characteristics of an amorphous sugar matrix in encapsulating emulsion droplets, especially during spray-drying, have been extensively investigated (Gharsallaoui et al., 2007, Hogan et al., 2003). It was found that droplet size is closely related to the encapsulation efficiency of amorphous sugar matrices (Minemoto et al., 2002, Soottitantawat et al., 2003, Soottitantawat et al., 2005). The method used for drying, including spray-drying, spray granulation, and freeze-drying were reported to strongly affect droplet-encapsulation efficiency as well as the storage stability of the material (Minemoto, Adachi, & Matsuno, 1997). It was also proposed that the optimum emulsification conditions could vary depending on the drying method used (Anwar & Kunz, 2011). However, a systematic comparison of the characteristics of amorphous matrices comprised of different types of sugar in the encapsulation of emulsion oil droplets has not been reported. Consequently information concerning how to optimize the encapsulation of oil-droplets by an amorphous sugar matrix upon freeze-drying is currently limited.

In this study, the characteristics of freeze-dried amorphous sugar matrices in encapsulating emulsion oil droplets were investigated for different types of sugars. Linoleic monomethyl ester (LME) and sucrose monolaurate (SML) were used as the oil phase and surfactant, respectively. Freeze-drying was used to dry the oil-in-water emulsion with the objective of comparing oil-droplet encapsulation characteristics for different types of sugar with known data regarding protein-stabilizing characteristics and because of the advantages of using freeze-drying (Anwar and Kunz, 2011, Longmore, 1971). The remaining amounts of LME in the freeze-dried sample matrix were first measured for different sugar/SML/LME compositions and at different stages of drying. The pathway by which LME is lost from the sample during freeze-drying was also investigated as well as the size distribution of the LME droplets before and after freeze-drying. The LME encapsulation efficiencies of amorphous matrices of different types of sugars were then compared to better understand the factors that determine the encapsulation efficiency. Furthermore, based on the obtained findings, methodology for improving oil-droplet encapsulation freeze-drying with a sugar present was investigated.