Morphological investigations of disaccharide molecules for growth inhibition of ice crystals

Abstract

Freezing of solutions including disaccharides (trehalose, sucrose, and maltose) has been investigated by microscopic observations of freeze-fractured replicas using FE-TEM. Three typical features were observed: the smooth surface considered as the ice crystal, fine particles as the precipitated disaccharide molecules, and remaining part as the glass state of the solution. The expanded observations of fine-particle and its distribution investigations suggested that it was larger than 10 nm in size and averaged approximately 20-30 nm in diameter. The smallest particle was estimated to include several hundred disaccharide molecules.Based on systematic observations of trehalose solutions regarding concentrations and freezing rates, we concluded that ice crystal growth was inhibited by trehalose molecules. Since the ice crystal size reduced exponentially with increase in trehalose concentration, we could control ice crystal size formed in the frozen material. The growth inhibition of ice crystals with trehalose resulted both from a reduction in the free water in the solution due to a significant hydration effect and from an enhancement of nucleation of the ice crystals.It was confirmed that trehalose was more effective than the other disaccharide solutions examined for inhibiting the growth of ice crystals.

Generation of micro- and nano-bubbles in water by dissociation of gas hydrates

“…In this condition, the freezing rate exceeds 10 3 K min -1 . For the subsequent freeze-fracture replica formation, we followed the procedure in [10,26], and give only a brief description here. To reduce the formation of artifacts during the replica film formation, the frozen droplet was fractured under vacuum (10 -4 to 10 -5 Pa) and low temperature (about 100 K).…”

Freezing Properties of Disaccharide Solutions: Inhibition of Hexagonal Ice Crystal Growth and Formation of Cubic Ice

“…Since these investigations allowed us to discuss the inhibition process of disaccharide molecules for the ice crystal growth in the extracellular space, we discuss the role of disaccharide molecules serving as the cryoprotectant of living cells in the final section. Uchida et al (2007) aimed to observe the solute-solution interaction in the liquid phase using a FEG-TEM along with the freeze-fractured replica technique. This technique has usually been applied for biological investigations.…”

Powder X-ray diffraction observations of ice crystals formed from disaccharide solutions

“…15 By the rapid freezing of disaccharide solutions (concentration ranged from 20 wt% to 50 wt%), the sample included both hexagonal and cubic ice crystals. This is the first report of cubic ice formation from disaccharide solutions, which was consistent with the results obtained in the glycerol, 20-21 glucose, 22-23 ionic salts 24 and aqueous aerosol.…”

“…15 Research-grade trehalose dihydrate crystals were donated by Hayashibara Biochemical Laboratories, Inc. To compare the properties observed in the trehalose solutions, we also conducted investigations using sucrose (of the highest quality; Sigma-Aldrich Japan, Inc.) and maltose (research grade, Hayashibara Biochemical Laboratories, Inc.).…”

“…15 The FEG-TEM observations showed three typical textures on the trehalose solution replicas (see Fig. 1): (S) smooth surface, (P) fine particles (average diameter was 20 -30 nm), and (R) remaining part of the boundary between the smooth-surface area co-existing with the fine particles.…”