Pharmaceutical Research

, Volume 20, Issue 4, pp 660–667

Effective Inhibition of Mannitol Crystallization in Frozen Solutions by Sodium Chloride



Purpose. The purpose of this work was to study the possibility of preventing mannitol crystallization in frozen solutions by using pharmaceutically acceptable additives.

Methods. Differential scanning calorimetry (DSC) and low-temperature X-ray diffractometry (LTXRD) were used to characterize the effect of additives on mannitol crystallization.

Results. DSC screening revealed that salts (sodium chloride, sodium citrate, and sodium acetate) inhibited mannitol crystallization in frozen solutions more effectively than selected surfactants, α-cyclodextrin, polymers, and alditols. This finding prompted further studies of the crystallization in the mannitol-NaCl-water system. Isothermal DSC results indicated that mannitol crystallization in frozen solutions was significantly retarded in the presence of NaCl and that NaCl did not crystallize until mannitol crystallization completed. Low-temperature X-ray diffractometry data showed that when a 10% w/v mannitol solution without additive was cooled at 1°C/min, the crystalline phases emerging after ice crystallization were those of a mannitol hydrate as well as the anhydrous polymorphs. In the presence of NaCl (5% w/v), mannitol crystallization was suppressed during both cooling and warming and occurred only after annealing and rewarming. In the latter case however, mannitol did not crystallize as the hydrate, but as the anhydrous δ polymorph. At a lower NaCl concentration of 1% w/v, the inhibitory effect of NaCl on mannitol crystallization was evident even during annealing at temperatures close to the Tg′ (−40°C). A preliminary lyophilization cycle with polyvinyl pyrrolidone and NaCl as additives rendered mannitol amorphous.

Conclusion. The effectiveness of additives in inhibiting mannitol crystallization in frozen solutions follows the general order: salts > alditols > polyvinyl pyrrolidone > α-cyclodextrin > polysorbate 80 ∼ polyethylene glycol ∼ poloxamer. The judicious use of additives can retain mannitol amorphous during all the stages of the freeze-drying cycle.

Mannitol sodium chloride crystallization DSC low temperature XRD 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    K. Izutsu, S. Yoshioka, and T. Terao. Decreased protein-stabilizing effects of cryoprotectants due to crystallization. Pharm. Res. 10:1232-1237 (1993).Google Scholar
  2. 2.
    K. Izutsu, S. Yoshioka, and T. Terao. Effect of mannitol crystallinity on the stabilization of enzymes during freeze-drying. Chem. Pharm. Bull. 42:5-8 (1994).Google Scholar
  3. 3.
    K. Izutsu and S. Kojima. Excipient crystallinity and its protein-structure-stabilizing effect during freeze-drying. J. Pharm. Pharmacol. 54:1033-1039 (2002).Google Scholar
  4. 4.
    M. J. Pikal, K. M. Dellerman, M. I. Roy, and R. M. Riggin. The effects of formulation variables on the stability of freeze-dried human growth hormone. Pharm. Res. 8:427-436 (1991).Google Scholar
  5. 5.
    K. Tanaka, T. Takeda, and K. Miyajima. Cryoprotective effect of saccharides on denaturation of catalase by freeze-drying. Chem. Pharm. Bull. 39:1091-1094 (1991).Google Scholar
  6. 6.
    L. Yu, D. S. Mishra, and D. R. Rigsbee. Determination of the glass properties of D-mannitol using sorbitol as an impurity. J. Pharm. Sci. 87:774-777 (1998).Google Scholar
  7. 7.
    A. I. Kim, M. J. Akers, and S. L. Nail. The physical state of mannitol after freeze-drying: effects of mannitol concentration, freezing rate, and a noncrystallizing cosolute. J. Pharm. Sci. 87:931-935 (1998).Google Scholar
  8. 8.
    T. R. Kovalcik and J. K. Guillory. The stability of cyclophosphamide in lyophilized cakes. Part I. Mannitol, lactose, and sodium bicarbonate as excipients. J. Parenter. Sci. Technol. 42:29-37 (1988).Google Scholar
  9. 9.
    M. G. Fakes, M. V. Dali, T. A. Haby, K. R. Morris, S. A. Varia, and A. T. M. Serajuddin. Moisture sorption behavior of selected bulking agents used in lyophilized products. PDA J. Pharm. Sci. Technol. 54:144-149 (2000).Google Scholar
  10. 10.
    S. D. Allison, B. Chang, T. W. Randolph, and J. F. Carpenter. Hydrogen bonding between sugar and protein is responsible for inhibition of dehydration-induced protein unfolding. Arch. Biochem. Biophys. 365:289-298 (1999).Google Scholar
  11. 11.
    M. J. Pikal. Mechanisms of protein stabilization during freeze-drying and storage: the relative importance of thermodynamic stabilization and glassy state relaxation dynamics. Drugs Pharmaceut. Sci. 96:161-198 (1999).Google Scholar
  12. 12.
    R. K. Cavatur, N. M. Vemuri, A. Pyne, Z. Chrzan, D. Toledo-Velasquez, and R. Suryanarayanan. Crystallization behavior of mannitol in frozen aqueous solutions. Pharm. Res. 19:894-900 (2002).Google Scholar
  13. 13.
    M. Yoshioka, B. C. Hancock, and G. Zografi. Inhibition of indomethacin crystallization in poly(vinylpyrrolidone) coprecipitates. J. Pharm. Sci. 84:983-986 (1995).Google Scholar
  14. 14.
    R. Haikala, R. Eerola, V. P. Tanninen, and J. Yliruusi. Polymorphic changes of mannitol during freeze-drying: effect of surface-active agents. PDA J. Pharm. Sci. Technol. 51:96-101 (1997).Google Scholar
  15. 15.
    N. A. Williams and T. Dean. Vial breakage by frozen mannitol solutions: correlation with thermal characteristics and effect of stereoisomerism, additives, and vial configuration. J. Parenter. Sci. Technol. 45:94-100 (1991).Google Scholar
  16. 16.
    L. Yu, N. Milton, E. G. Groleau, D. S. Mishra, and R. E. Vansickle. Existence of a mannitol hydrate during freeze-drying and practical implications. J. Pharm. Sci. 88:196-198 (1999).Google Scholar
  17. 17.
    E. Y. Shalaev, F. Franks, and P. Echlin. Crystalline and Amorphous Phases in the Ternary System Water-Sucrose-Sodium Chloride. J. Phys. Chem. 100:1144-1152 (1996).Google Scholar
  18. 18.
    K. Ito. Freeze drying of pharmaceuticals. Eutectic temperature and collapse temperature of solute matrix upon freeze drying of three-component systems. Chem. Pharm. Bull. 19:1095-1102 (1971).Google Scholar
  19. 19.
    A. Martini, K. Silvia, M. Crivellente, and R. Artico. Use of subambient differential scanning calorimetry to monitor the frozen state behavior of blends of excipients for freeze-drying. PDA J. Pharm. Sci. Technol. 51:62-67 (1997).Google Scholar
  20. 20.
    P. Meredith, A. M. Donald, and R. S. Payne. Freeze-drying: in situ observations using cryoenvironmental scanning electron microscopy and differential scanning calorimetry. J. Pharm. Sci. 85:631-637 (1996).Google Scholar
  21. 21.
    R. K. Cavatur and R. Suryanarayanan. Characterization of phase transitions during freeze-drying by in situ X-ray powder diffractometry. Pharm. Dev. Technol. 3:579-586 (1998).Google Scholar
  22. 22.
    K. Izutsu, S. Yoshioka, and S. Kojima. Effect of cryoprotectants on the eutectic crystallization of NaCl in frozen solutions studied by differential scanning calorimetry (DSC) and broad-line pulsed NMR. Chem. Pharm. Bull. 43:1804-1806 (1995).Google Scholar
  23. 23.
    B. S. Chang and C. S. Randall. Use of subambient thermal analysis to optimize protein lyophilization. Cryobiology 29:632-656 (1992).Google Scholar
  24. 24.
    S. Ablett, M. J. Izzard, and P. J. Lillford. Differential scanning calorimetric study of frozen sucrose and glycerol solutions. J. Chem. Soc. Far. Trans. 88:789-794 (1992).Google Scholar
  25. 25.
    B. Luyet and D. Rasmussen. Study by differential thermal analysis of the temperature of instability of rapidly cooled solutions of glycerol, ethylene glycol, sucrose and glucose. Biodynamica 10:1167-1191 (1968).Google Scholar
  26. 26.
    A. Pyne, R. Surana, and R. Suryanarayanan. Crystallization of mannitol below Tg′ during freeze-drying in binary and ternary aqueous systems. Pharm. Res. 19:901-908 (2002).Google Scholar
  27. 27.
    B. D. Herman, B. D. Sinclair, N. Milton, and S. L. Nail. The effect of bulking agent on the solid-state stability of freeze-dried methylprednisolone sodium succinate. Pharm. Res. 11:1467-1473 (1994).Google Scholar
  28. 28.
    C. Ahlneck and G. Zografi. The molecular basis of moisture effects on the physical and chemical stability of drugs in the solid state. Int. J. Pharm. 62:87-95 (1990).Google Scholar

Copyright information

© Plenum Publishing Corporation 2003

Authors and Affiliations

  1. 1.College of PharmacyUniversity of Minnesota
  2. 2.Lilly Research LaboratoriesEli Lilly and CompanyIndianapolis

Personalised recommendations