J. Phys. Chem. B 2002, 106, 3365-3370
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Vitrification and Polymorphism of Trehalose Induced by Dehydration of Trehalose Dihydrate J. F. Willart,*,† A. De Gusseme,† S. Hemon,† M. Descamps,† F. Leveiller,‡ and A. Rameau‡ Laboratoire de Dynamique et Structure des Mate´ riaux Mole´ culaires, ESA CNRS 8024, UniVersite´ de Lille 1, Baˆ t. P5, 59655 VilleneuVe d’Ascq, France and Laboratoire d'Analyse Physique - AVentis Pharma - Vitry-AlforVille ReceiVed: January 31, 2001; In Final Form: January 10, 2002
Calorimetric and X-ray diffraction investigations of anhydrous R trehalose obtained by dehydration of its dihydrate form have been performed. The results show that the structural and thermodynamic properties of the dehydrated state strongly depend on the kinetics of water loss. A fast dehydration promotes the formation of a glassy amorphous phase and thus provides an unusual example of direct transformation from crystal to glass. On the other hand, a slow dehydration promotes the formation of a polymorphic crystalline phase (R) of anhydrous trehalose. We also show how this duality crystalline state/glassy amorphous state has led, in previous studies, to a confused pattern of thermodynamic properties of the dehydrated state of trehalose dihydrate. Our results show, in particular, that the properties of the glassy amorphous trehalose obtained by dehydration are very close to that of the quenched liquid trehalose.
1. Introduction Amorphous solids are generally obtained by the rapid undercooling of a melted crystal.1 However, there exist less usual routes for solid-state amorphization which do not require the melting of the crystalline state.2-4 They are, for instance, milling,3,5,6 dehydration,7-9 irradiation,4 and pressure.10,11 When these so-called “nonthermal routes of amorphization” are used below the glass transition temperature (Tg) of the corresponding liquid, the formation of a glassy amorphous state can be expected. They can thus potentially offer unusual examples of direct transformation from crystal to glass and could contribute to the general understanding of the glass transition itself. Many fundamental questions, still unresolved, arise from the nonthermal routes of amorphization and vitrification. One of them is to decide whether the nature and the properties (lifetime, fragility,12 and local structure) of the amorphous states obtained by nonthermal routes are similar to those of the amorphous state produced by the usual thermal route (quench of the liquid). We can wonder, in particular, if these nonthermal routes can produce new situations of polyamorphism similar to those suspected, for instance, in TPP,13,14 in selenium,15 and in ice.16 Another question concerns the understanding of the physical mechanisms themselves which drive the solid-state amorphizations. This understanding remains strongly elusive, especially for organic molecular materials which have been much less studied, from that point of view, than alloys,17 inter-metallic,18,19 and inorganic compounds.20 The above questions are expected to be enlightened by the study of crystalline samples which can be amorphized and vitrified by different routes independent of one another. Such a very unusual possibility is encountered in trehalose (R-Dglucopyranosyl, R-D-glucopyranoside) which can reach a glassy amorphous state by the usual thermal quench of its liquid * To whom correspondence should be addressed. † Universite ´ de Lille. ‡ Aventis Pharma.
phase,21 by milling of its crystalline anhydrous form β,22 and by dehydration of its dihydrate form T2H2O.23 The possibility to produce glassy amorphous anhydrous trehalose by dehydration of its dihydrate crystalline form was first proposed in 199623 and has been extensively studied since then.24-27 However, the nature and the properties of the resulting dehydrated state are still very unclear. We notice in particular the following: (i) The glassy character itself of the dehydrated T2H2O remains strongly controversial. Sussich et al.,26 for instance, have argued recently (1998) that the dehydration of T2H2O “directly produces an amorphous phase, and not the glassy form, which can be prepared only if the undercooled liquid is quenched at temperature below the glass transition”. (ii) The glass transition temperatures reported in the literature are widely dispersed (79 °C21 < Tg < 115 °C23,28). (iii) Several interpretations have been proposed to explain the unusually low enthalpy which has been found to characterize the glass obtained by dehydration.23 (iv) There is considerable confusion concerning its underlying polymorphism (see references 26 and 24 for a review). In particular, in addition to the well-characterized crystalline phases β29 and T2H2O30,31 of anhydrous and dihydrate trehalose, a polymorphic phase (R) of anhydrous trehalose has been found to develop for some specific dehydration conditions (4 h aging of T2H2O at 85 °C under vacuum26,32). The reason this phase R develops instead of the amorphous phase is not yet understood. Another polymorph (named γ)26 formed by heating T2H2O at 5-20 K/min was also announced by Sussich et al., but it was recently identified by these authors as being, in fact, a mixture of T2H2O and β.25 This shows the difficulties in identifying the polymorphic forms of trehalose and in managing the dehydration procedures by which they are produced. In this paper, we show that the kinetics of water loss acts as a non-equilibrium parameter which can drive the crystalline T2H2O either toward a glassy amorphous state or toward a polymorphic phase of anhydrous trehalose. It is also underlined
10.1021/jp012836+ CCC: $22.00 © 2002 American Chemical Society Published on Web 03/09/2002
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how this behavior can explain some diverging and puzzling results previously reported in the literature. This study could be done by the careful control of suitable dehydration protocols in the course of differential scanning calorimetry (DSC), real time X-ray diffraction (XRD), and thermo-gravimetric analysis (TGA) experiments. 2. Experimental Section R-R Trehalose dihydrate was purchased from Fluka. It was more than 99% pure and was used without further purification. The TGA experiments were performed with the TGA 2950 of TA Instruments. During all the measurements, the sample was placed in an open platinium sample pan and flushed with a highly pure nitrogen gas. The temperature reading was calibrated using the Curie points of alumel and nickel, while the mass reading was calibrated using balance tare weights provided by TA Instruments. The powder X-ray diffraction (XRD) experiments were performed with (i) an Inel CPS 120 diffractometer (λCuKR1 ) 1.5405 Å) equipped with a 120° curved position sensitive detector coupled to a 4096 channel analyzer. This diffratometer was used for the structural analysis of the equilibrium crystalline phases. The samples were placed into Lindemann capillaries (Ø ) 0.7 mm). (ii) a Siemens D5000 diffractometer (λCoKR ) 1.791 Å) equipped with a 15° linear detector coupled to a 512 channel analyzer. This diffractometer was used for rapid time-resolved structural investigations during the dehydration of T2H2O. The samples were spread out on horizontal aluminum plates. The DSC experiments were performed with the DSC 2920 microcalorimeter of TA Instruments. During all the measurements, the calorimeter head was flushed with highly pure helium gas. Temperature and enthalpy readings were calibrated using pure indium at the same scan rates used in the experiments. A small amount (≈4 mg) of T2H2O was used to improve both the resolution and the thermal conductivity. The dehydration effects will be discussed with respect to the conventional glass transition of the undercooled liquid which is shown in run 3 (Figure 1) to occur at Tg ) 120 °C for a heating rate of 5 K/min. The objective of this paper is to show the influence of the dehydration protocol on the dehydrated state of T2H2O. Special attention has thus been paid to the encapsulation of the sample which strongly affects the dehydration characteristics during heating experiments. This clearly appears in Figure 1 which shows two DSC scans (run 1 and 2) obtained with the same heating rate (T˙ ) 5 K/min) but for two different sample encapsulations. (i) In run 1, the sample was placed in an open cell (DSC container with no cover). The dehydration process then appears through a broad endotherm which coincides perfectly with the water loss revealed by a TGA experiment (run 4) obtained with the same operating conditions (T˙ ) 5 K/minsopen cell). This dehydration starts at 50 °C and is completed at 100 °C so that it does not overlap the glass transition range (∼120 °C) of the undercooled liquid. A small endotherm can then be clearly seen between 115 °C and 130 °C. Its nature and its origin are the main points of this paper and will be discussed in more detail in the next section. (ii) In run 2, the sample was placed in a pierced cell (DSC container with a pierced cover). The overall dehydration process then occurs over a larger temperature range indicating some hindrance in the water removal. It always starts at 50 °C but peaks at 100 °C and ends at 130 °C. In these conditions, the endotherm of dehydration merges with the endotherm in the Tg
Figure 1. Run 4: TGA scan of T2H2O placed in an open cell; run 1: DSC scan of T2H2O placed in an open cell; run 3: DSC scan of quenched liquid trehalose placed in an open cell; run 2: DSC scan of T2H2O placed in a pierced cell. Each scan was obtained upon heating at T˙ ) 5 K/min.
range which is then hardly seen. Moreover, the enthalpy of this latter endotherm is clearly smaller than that observed in run 1. Such a difference already shows the influence of the dehydration protocol on the dehydrated state of T2H2O which is the main point of the next section. In our DSC experiments, we have thus systematically used open cells which provide (i) reproducible dehydration experiments. (ii) dehydration conditions fully similar to those of TGA and XRD. (iii) better conditions for the study of the glass transition temperature range. 3. Results 3.1 Calorimetry. Figure 2a shows thermograms obtained upon heating T2H2O with different scan rates ranging from 1 K/min to 50 K/min. A close inspection of the thermogram corresponding to the scanning rate of 5 K/min shows that the endotherm occurring in the Tg range is more complex than a mere glass transition. As shown in the inset, it seems to be structured with a slight shouldering on the left-hand side of the endotherm. For a slower heating rate (1 K/min) this complexity is not perceptible to the eye, while for heating rates higher than 20 K/min this complexity is screened by the shift of the dehydration peak toward the high temperatures. To get rid of this effect and to study in more detail the glass transition range, we have used a more adequate dehydration protocol, sketched in the inset of Figure 2b. In the experiments of Figure 2b, the T2H2O samples have been first dehydrated by heating from 10 °C to 110 °C with heating rates varying from 1 K/min to 50 K/min, kept at this temperature for 5 min, cooled to 30 °C, and then heated again above the melting point at the rate of 5 K/min. The thermograms shown in Figure 2b correspond to this final heating stage. All of them were thus recorded at the rate of 5 K/min, and they only differ from each other by the heating rate with which they were previously dehydrated. In these conditions, there is no more dehydration peak which overlaps the glass transition range, and
Vitrification and Polymorphism of Trehalose
Figure 2. (a) DSC thermograms of T2H2O obtained for different heating rates (1, 5, 10, 20, and 50 K/min). For clarity, only the decreasing part of the dehydration endotherm has been reported for 5, 10, 20, and 50 K/min. The insert shows a close-up view of the glass transition range corresponding to the thermogram obtained at 5 K/min. (b) DSC heating curves (T˙ ) 5 K/min) obtained from different samples of T2H2O which have been previously heated at 110 °C at different rates q (ranging from 1 to 50 K/min) to be dehydrated, as indicated in the insert.
the complexity of the thermogram in the Tg range, suspected in Figure 2a for a heating rate of 5 K/min, is now clearly seen for any dehydration rate. In particular, it appears that the endotherm in the Tg range can be divided in two successive and close enthalpic events: (i) A Cp jump at 120 °C, immediately followed by the endothermic overshoot characteristic of annealed glasses. These features coincide perfectly with those of the quenched liquid whatever the dehydration rate. (ii) A weak endothermic peak at 125 °C whose enthalpy strongly depends on the dehydration rate. This peak is nearly nonexistent for a rapid dehydration (50 K/min), while for a slow dehydration rate (1 K/min) this peak is so developed that it masks the glass transition. The latter is, however, still perceptible through a small shouldering located on the left-hand side of the endotherm. Compared to that of the liquid glass, the DSC traces of the dehydrated T2H2O samples (Figure 2b) are unexpectedly flat between 50 °C and 100 °C which suggests smaller Cp variations in this temperature range. In fact, it has been checked that the dehydrated T2H2O is highly hygroscopic so that a small amount of water ( 50 K/min) dehydration of crystalline dihydrate trehalose (T2H2O) at 110 °C, (b) amorphous trehalose obtained by quench of the liquid state, and (c) crystalline anhydrous trehalose and crystalline dihydrate trehalose.
3.2 X-ray Diffraction. The structural state of the sample after dehydration of T2H2O has been investigated by X-ray diffraction. Figure 3a shows two diffractograms recorded at RT after a fast (T˙ > 50 K/min) and a slow (T˙ ) 1 K/min) dehydration of T2H2O at 110 °C. They correspond to the two extreme situations studied in Figure 2b. After a fast dehydration, the X-ray diffraction pattern of the sample does not present any Bragg peak and is thus characteristic of an X-ray amorphous solid. This diffractogram is fully similar to that of the quenched liquid shown in Figure 3b. On the other hand, after a slow dehydration, the X-ray diffraction pattern clearly shows Bragg peaks which indicate that the sample is not in an amorphous state. Direct confrontation with the diffractograms of crystalline trehalose dihydrate and crystalline anhydrous trehalose shown in Figure 3c indicates that these Bragg peaks are neither characteristic of phase T2H2O nor characteristic of phase β. They thus correspond to a polymorphic phase of anhydrous trehalose. Moreover, the diffraction lines are broad which indicates that this polymorphic phase is probably highly strained or has a small coherent spatial extent. Upon heating, these diffraction lines have been found to collapse at 125 °C indicating the transition of this polymorphic phase toward an amorphous state. The Bragg peaks collapse is thus the structural signature corresponding to the endotherm observed at the same temperature (just above Tg) in the thermograms of Figure 2b. Both its X-ray diffraction pattern and its melting temperature indicate that the polymorphic phase formed during a slow dehydration (Figure 2b) is likely to be the phase R detected by Perlin32 and Sussich26 after a 4 h aging of a T2H2O sample at 85 °C in a vacuum. 3.3 Time-Resolved X-ray Diffraction. The kinetics of the transformation T2H2O f R induced by water removal cannot be studied in detail by DSC since it superimposes unavoidably to the highly endothermic dehydration process itself. On the other hand, this masking effect does not occur in X-ray diffraction experiments. We could thus follow the transformation T2H2O f R by the rapid measurements of the X-ray diffraction pattern during a slow dehydration of T2H2O. This real-time experiment was performed during an isothermal dehydration at 50 °C for
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Figure 5. DSC heating curves (T˙ ) 5 K/min) obtained for a T2H2O sample previously dehydrated by a 3 h annealing at 50 °C and for the quenched liquid trehalose.
Figure 4. Temporal evolution of the X-ray diffraction pattern during a slow dehydration of T2H2O obtained by a 2 h annealing at 50 °C. The temporal evolutions of the fraction of phase T2H2O and the fraction of phase R derived from the X-ray diffraction patterns, as well as the dehydrated fraction, measured by TGA during the same dehydration process, are shown together in the inset.
which the time scale of the conversion (2 h) is long in regard to the time scale of the measurement (30 s/scan). The results are reported in Figure 4. They show the progressive disappearance of the sharp Bragg peaks characteristic of T2H2O concomitant with the development of the broader Bragg peaks characteristic of phase R. The corresponding temporal evolution of the fraction of each phase during the 2 h dehydration is shown in the inset of Figure 4, together with the water loss of the sample, measured by TGA in the same conditions (open cell) and during the same thermal treatment (2 h aging at 50 °C). The results indicate that after 2 h the sample is fully dehydrated and no more structural evolution could be detected when further annealing the sample. Moreover, the dehydration kinetics appears to be perfectly parallel to the interconversion T2H2O f R. This indicates that, for a slow dehydration, the initial structure of T2H2O which has lost its two water molecules is unstable and transforms, without any delay, toward phase R. The enthalpy corresponding to the melting of phase R increases with the slowness of the dehydration (Figure 2). This is likely due to the variable proportion of phase R generated during the dehydration process. A good estimation of the enthalpy of melting of phase R thus requires a slow dehydration procedure. Figure 5 shows the thermogram obtained upon heating (T˙ ) 5 K/min), after a 3 h isothermal dehydration of T2H2O at 50 °C. In these conditions, the endothermic overshoot ending the Cp jump at Tg is no longer perceptible so that one may think that most of the sample has transformed toward phase R. The enthalpy of melting (∆HR ) 11.6 kJ/mol) is at least twice bigger than that previously published26 and must be considered as a lowest limit of the true value corresponding to the melting of the pure phase R. 4. Discussion The calorimetric and X-ray diffraction results shown in Figures 2 and 3 reveal clearly that the structural and thermodynamic properties of the dehydrated T2H2O strongly depend on the dehydration rate. A fast dehydration (T˙ > 50 K/min) mainly produces amorphous anhydrous trehalose with a clear glassy character. On the other hand, a slow dehydration (T˙ < 1 K/min)
mainly produces a polymorphic crystalline phase (R) of anhydrous trehalose. For intermediate dehydration rate, a mixed state made of amorphous trehalose and polymorphic phase R is formed in proportion fixed by the dehydration rate. (i) Dehydration-Induced Glassy Amorphous Trehalose. The glassy character of the amorphous trehalose obtained by dehydration of T2H2O was never clearly detected in previous studies of trehalose. Recently (1998), Sussich et al.26 have even argued that the amorphous state obtained by dehydration of T2H2O was not glassy. This conclusion is likely to be due to the specific experimental procedure which was used for the dehydration. As shown in section 2, the combined use of a simple temperature ramp and a pierced cell in heating DSC experiments of T2H2O provides a huge endothermic dehydration peak in the Tg range which masks the much weaker glass transition signature. Earlier (1996), the possible vitrification of trehalose by water loss from its crystalline dihydrate had, however, been put forward by Ding et al. in ref 23. In this paper, the dehydration was performed by keeping a T2H2O sample under vacuum at 70 °C during 30 h. The subsequent heating of this dehydrated sample has revealed above 125 °C a clear endotherm which was attributed to the glass transition of the amorphous dehydrated sample. Striking features of the glass obtained by this route, compared to that obtained by the quench of the liquid, were its abnormally large endothermic overshoot accompanying the glass transition and the slightly higher glass transition temperature. Two interpretations were then proposed to tentatively explain this unexpected glass transition signature: Either the glass formed by dehydration is effectively of unusually low enthalpy, or the Cp overshoot includes a heat absorption component corresponding to the melting of residual pockets of the initial T2H2O phase which were not amorphized. Moreover, if the structural water released by T2H2O upon dehydration remains trapped inside the DSC cell, it can have a plasticizing effect which can strongly depress the glass transition temperature of the sample. In light of our results, the endotherm observed above 125 °C in the work of Ding et al. corresponds to the endotherm observed, in our paper, for a dehydration rate of 1 K/min (Figure 2). These authors have thus seen the melting of the polymorphic crystalline phase R generated by their specific dehydration protocol and not the glass transition of an amorphous state produced by the dehydration of T2H2O. Moreover, this melting cannot be that of residual pocket of T2H2O23 since, as shown in Figure 4, the phase which melts at 125 °C does not vanish but develops during a slow dehydration.
Vitrification and Polymorphism of Trehalose Contrary to the previous results published by Ding et al., the glass transition reported in our paper (Figure 2) and detected after a fast dehydration of T2H2O (T˙ ) 5 K/min) is very similar to that of the quenched liquid. In particular, it occurs exactly at the same temperature and shows nearly the same enthalpic reequilibration signature. Only weak differences in the endothermic overshoot accompanying the Cp jump have been detected which must clearly be attributed to the effective different aging process undergone by the amorphous part of the sample during the thermal treatment use for the dehydration. Figure 2 shows that amorphous trehalose obtained by dehydration of T2H2O or melting of phase R undergoes systematically, upon heating, a cold crystallization toward the most stable anhydrous phase β. On the other hand, this behavior is never observed in the undercooled liquid. This may indicate a delicate difference between the structures of the two amorphous states which has strong repercussions in the nucleation rate of phase β. One may think, in particular, that the amorphous state obtained directly from a crystalline phase is less “disordered” than that obtained from the quench of a liquid. This could be due to a slightly different local order which is reminiscent of the initial structure of T2H2O and which can promote the nucleation of phase β. In trehalose, this effect may be enhanced by the fact that the starting phase (T2H2O) and the coldcrystallized phase (β) are structurally very close to each other.29,30 Other examples of non-equivalence between amorphous states obtained by melt cooling and by nonthermal routes (like ball milling) have already been reported for both metallic compounds (selenium15) and molecular crystals (cephalotin sodium33). (ii) Dehydration-Induced Polymorphism of Trehalose. The very broad Bragg peaks which characterize the X-ray diffraction pattern of phase R suggest that this phase is highly strained and has a small coherent spatial extent which makes its structural analysis difficult. However, the weak enthalpy characterizing the melting of the polymorphic phase R, as well as its nonbirefringent character reported by Perlin et al.,32 indicate that this phase is very likely of high symmetry. Moreover, contrary to amorphous trehalose and to the anhydrous phase β, the return to the most stable dihydrate form is nearly instantaneous at RT, even for very weak relative humidity. The structure of phase R is thus expected to be close to that of T2H2O. The parallelism between the kinetics of water loss and that of transformation toward phase R (inset of Figure 4) indicates that phase R develops inside the anhydrous zones as and when they appear. The morphology of phase R is thus expected to mimic that of the dehydration pattern which is not governed, a priori, by the usual laws of phase transformations (e.g., nucleation and growth) but by hydrodynamic laws. Such a morphology is probably much more complex than mere macroscopic compact zones. Moreover, the growth rate of phase R is obviously limited by that of the dehydrated zones so that the development of this phase nearly occurs in confined geometry. Such a mechanism is likely to provide small coherent domains of phase R and thus contribute to explain the broad Bragg peaks which characterize its X-ray diffraction pattern. Another striking feature of phase R concerns its melting temperature which is unusually located just above Tg. This suggests that, even below Tg, phase R is unstable in regard to the amorphous trehalose but keeps living because the sub-Tg dynamics are far too slow to drive effectively the material toward an amorphous state. Such a transformation can only occur above Tg. Another interpretation of this close to Tg melting can, however, be put forward which is similar to the non-
J. Phys. Chem. B, Vol. 106, No. 13, 2002 3369 equilibrium melting of polymers.34 In this latter case, phase transition temperatures are sometimes very depressed when the “size” of the phase is weak. This so-called Gibbs Thomson effect35 could also apply to the polymorphic phase R of trehalose whose small coherent spatial extent is strongly suggested by the structural analysis. It cannot thus be excluded that the endotherm observed at 125 °C corresponds to the nonequilibrium melting of this phase R which is strongly depressed in regard to the equilibrium melting temperature of the bulk crystalline phase. References 26 and 24 provide thermograms obtained for thermal treatments “similar” to those of Figure 2. However, these thermograms reveal either nearly no sign of phase R or much more complex behaviors. In reference 26, these differences are most probably due to the use of pierced cells instead of open cells. This strongly modifies the kinetics of water removal and also induces spurious effects in the DSC scans as clearly described in reference 24. In this latter work, open cells were used but most of the dehydration procedures were performed under vacuum, which obviously increases the rate of water removal. This protocol promotes the formation of amorphous trehalose to the detriment of phase R which is then nearly not detected. These differences between the results of references 24 and 26 and ours reveal the very high sensitivity of the dehydrated state of T2H2O to the dehydration protocol. A perfect management of the structural and thermodynamic evolution of the sample upon dehydration thus requires to control and report all the parameters which govern the kinetics of water loss such as the temperature, pressure, and atmosphere histories undergone by the sample. 5. Conclusion This work shows that the structural and thermodynamic properties of anhydrous trehalose obtained by dehydration of T2H2O is very sensitive to the dehydration protocol itself. A fast dehydration of T2H2O produces amorphous anhydrous trehalose which has clear glassy properties very similar to those of conventional glass-forming liquids. On the other hand, a slow dehydration produces a polymorphic phase (R) of anhydrous trehalose which is characterized by a small coherent spatial extent. For intermediate dehydration rate, a mixed state made of glassy amorphous phase and polymorphic phase (R) is formed in proportion fixed by the dehydration rate. The glass transition of the amorphous state and the melting of phase R occur in a very narrow temperature range and exhibit enthalpic events with comparable amplitudes so that they can be easily mistaken one for the other. This possible confusion is most probably at the origin of some of the controversial results previously published about the dehydrated state of T2H2O.23,26 For rapid-enough dehydration, T2H2O thus provides an unusual example of direct transformation from ordered crystal to glass. In such a transformation, the glassy amorphous state is interestingly reached from a highly ordered state (crystalline) instead of from a highly disordered state (liquid) for conventional glasses. From that point of view, further investigations of this unusual route to the glassy state are expected to improve our understanding of the general glass transition process itself. Trehalose is currently of much interest in biology for its very efficient biopreservation properties of proteins and lyposomes against water stresses induced by dehydration or freezing. The origin of these interesting biological properties are not yet clearly understood,36-40 but the ability of trehalose to form a dihydrate which amorphizes upon dehydration is generally considered to
3370 J. Phys. Chem. B, Vol. 106, No. 13, 2002 be essential. It is thus a challenge to control and predict the structure, the microstructure and the degree of stability of the different crystalline and amorphous phases of trehalose against dehydration and any other constraints (e.g., temperature, milling, pressure) undergone during industrial processing and inside living organisms. Acknowledgment. This work was performed in the framework of an Interreg II grant between Nord-Pas de Calais and Kent. References and Notes (1) Ediger, M. D.; Angell, C. A.; Nagel, S. R. J. Phys. Chem. 1996, 100, 13200. (2) Angell, C. A. Science 1995, 267, 1924. (3) Suga, H. J. Therm. Anal. Calorim. 2000, 60, 957. (4) Martin, G.; Bellon, P. Solid State Physics, New York 1997, 50, 189. (5) Font, J.; Muntasell, J.; Cesari, E. Mater. Res. Bull. 1995, 267, 1924. (6) Chen, Y.; Bibole, M.; Hazif, R. L.; Martin, G. Phys. ReV B 1993, 48, 14. (7) Onodera, N.; Suga, H.; Seki, S. Bull. Chem. Soc. Jpn. 1968, 41, 2222. (8) Petit, S.; Coquerel, G. Chem. Mater. 1996, 8, 2247. (9) Galwey, A. K. Thermochim. Acta 2000, 355, 181. (10) Kruger, M. B.; Jeabloz, R. Science 1990, 249, 647. (11) Sharma, S. M.; Sikka, S. K. Prog. Mater. Sci. 1996, 40, 1. (12) Angell, C. A. J. Phys. Chem. Solids 1988, 49, 363. (13) Ha, A.; Cohen, I.; Iee, M.; Kievielson, D. J. Phys. Chem. Solids 1996, 100, 1. (14) Hedoux, A.; Guinet, Y.; Descamps, M. Phys. ReV. B 1998, 58, 1. (15) Lu, K.; Guo, F. Q.; Zhao, Y. H.; Jin, Z. H. J. Metastable Nanocrystalline Mater. 1999, 2-6, 43. (16) Mishima, O. Nature 1996, 384, 546. (17) Johnson, W. L.; Li, M.; Krill, C. E. J. Non-Cryst. Solids 1993, 156/158, 481. (18) Richet, P.; Gillet, P. Eur. J. Mineral. 1997, 9, 907.
Willart et al. (19) Fan, G. J.; Guo, F. Q.; Hu, Z. Q.; Quan, M.; Lu, K. Phys. ReV. B 1997, 55, 11010. (20) Wentzcovitch, R.; Silva, C. D.; Chelikowsky, J. Phys. ReV. Lett. 1998, 80, 2149. (21) Green, J. L.; Angell, C. A. J. Phys. Chem. 1989, 93, 2880. (22) Willart, J. F.; Gusseme, A. D.; Hemon, S.; Odou, G.; Danede, F.; Descamps, M. Solid State Commun. 2001, 119, 501. (23) Ding, S.; Fan, J.; Green, L.; Lu, Q.; Angell, C. A. J. Therm. Anal. 1996, 47, 1391. (24) Macdonald, C.; Johari, G. P. J. Mol. Struct. 2000, 523, 119. (25) Sussich, F.; Princivalle, F.; Cesaro, A. Carbohydr. Res. 1999, 322, 113. (26) Sussich, F.; Urbani, R.; Princivalle, F.; Cesaro, A. J. Am. Chem. Soc. 1998, 120, 7893. (27) Sussich, F.; Urbani, R.; Cesaro, A.; Princivalle, F.; Bruckner, S. Carbohydr. Lett. 1997, 2, 403. (28) Roos, Y. H. Carbohydr. Res. 1993, 238, 39. (29) Jeffrey, G. A.; Nanni, E. Carbohydr. Res. 1985, 137, 21. CCSD refcode: DEKYEX (30) Taga, T.; Senma, M.; Osaki, K. Acta Crystallogr., Sect. B 1972, B28, 3258. CCSD refcode: TREHAL03. (31) Brown, G. M.; Rohrer, D. C.; Berking, B.; Beevers, C. A.; Gould, R. O.; Simpson, R. Acta Crystallogr., Sect. B 1972, B28, 3145. CCSD refcode: TREHAL01/02. (32) Reisener, H. J.; Goldschmid, H. R.; Ledingham, G. A.; Perlin, A. S. Can. J. Biochem. Phys. 1962, 40, 1248. (33) Otsuka, M.; Kaneniwa, N. Int. J. Pharm. 1990, 124, 307. (34) Moyniham, C. T.; Macedo, P. B.; Montrose, C. J.; Gupta, P. K.; Debolt, M. A.; Dill, J. F.; Dom, B. E.; Drake, P. W.; Easteal, A. J.; Elterman, P. B.; Moeller, R. P.; Sasabe, K.; Wilder, J. A. Ann. N.Y. Acad. Sci. 1976, 279, 15. (35) Keller, A.; Hikosaka, M.; Rastagi, S.; Toda, A.; Barham, P. J.; Goldbeck-Wood, G. J. Mater. Sci. 1994, 29, 2579. (36) Roser, B. BioPharm 1991, 4, 47. (37) Crowe, L. M.; Reid, D. S.; Crowe, J. H. Biophys. J. 1996, 71, 2087. (38) Carpenter, J. F.; Crowe, J. H. Biochemistry 1992, 28, 3916. (39) Levine, H.; Slade, L. BioPharm. 1992, 5, 36. (40) Aldous, B. J.; Auffret, A. D. Cyo-Lett. 1995, 16, 181.
