ABSTRACT
Introduction ........................................................................................................ 697
Experimental Procedures.................................................................................. 698
Reagents....................................................................................................... 698
Sugar Crystallinity ..................................................................................... 698
X-ray Diffraction......................................................................................... 698
Glassy Preservation ................................................................................... 699
Enzyme Assay ............................................................................................ 699
Results and Discussion ..................................................................................... 700
Crystallinity Changes as a Function of RH ........................................... 700
LDH Preservation ...................................................................................... 701
Conclusion .......................................................................................................... 701
References ........................................................................................................... 701
Freeze-drying is the conventional way to stabilize labile biological products,
such as proteins and enzymes (Franks et al., 1991). Freeze-dried products
usually contain sugars, polyols, or amino acids as excipients. For example,
trehalose and other glass-forming solutes have been used as excipients for
enzyme EcoRI stabilization (Rossi et al., 1992). Excipients with the ability to
turn into the glassy state during the freeze-drying process help preserve the
physical and chemical integrity of biological products (Fox, 1995). Solute
crystallization from amorphous mixtures is considered to be of importance,
usually undesirable, for many applications of concentrated aqueous
carbohydrate systems; examples include food processing and preservation
of biological activity in freeze-dried pharmaceutical preparations. Recently,
however, it has been proposed that sugar crystallization may, in the case of
the formation of a stable crystalline hydrate, be of benefit in the enhancement
of chemical and physical stability in humid environments. Thus, if a sugar
can crystallize in the form of a hydrate in real time, then depending on the
hydrate stoichiometry, its crystallization from the amorphous state will
prevent the dilution of the remaining amorphous solution phase. If the
residual (noncrystalline) solution phase then becomes sufficiently concen-
trated, its glass-transition temperature will be raised (Kajiwara and Franks,
1997). In this study, raffinose and trehalose, which at ambient conditions
exist as a crystalline pentahydrate and dehydrate, respectively, were used as
amorphous excipients and its ability to stabilize LDH in various humid
conditions was tested.