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Freeze-drying of aqueous solutions: Maximum allowable operating temperature
CRYOBIOLOGY,
9,
559-561 (1972)
Freeze-Drying of Aqueoue Allowable Operating RICHARD Department
J. BELLOWS
of Chemical Engineering,
Solutions : Maximum Temperature
AND
University
Freeze-drying (Iyophilization) is a two-step dehydration process by which a substance is first frozen, and then water is removed by sublimation under vacuum. Freeze-drying has a diverse range of application, including the preparation of biological samples, pharmaceutical manufacture, and food preservation. Freeze-dried liquids show an abrupt, loss in product quality when the temperature of the subliming interface during drying is maintained above a critical value, t’he “collapse” temperature (6). Drying above this temperature gives a distorted (collapsed) product with poor solubility, low drying rates, uneven drying, and loss of texture and volatile substances. We find that, in the absence of solute crystallization, the collapse temperature depends principally upon the viscosity of the unfrozen port,ion of the solution. -4 simple theory, presentcd below, shows that increasing solute molecular weight gives higher collapse temperatures, and predict’s observed collapse temperatures quantitatively. Det>ails of experimental procedure, further background for the theory, and comparison with other reported experimental collapse temperatures are given elsewhere (1). Freezing causes a microseparation of an aqueous solution into a two-phase mixture of ice and a solute-rich phase. Some solutes crystallize readily, and for such solutions a eutectic mixture of ice and crystallized solut,e forms. Ito (3) has shown that solutions of eutectic-forming solutes collapse at their eutectic temperature during sublimation. Commonly, however, many organic solutes and especially mixed solutes such as occur in natural substances are kinetically inhibited from exhibiting eutect)ic behavior. Freezing these solutions gives a microscalc mixt,ure of ice needles and concentrated amorphous solution (CaS). The CAS remains an amorphous aqueous solution or a glass. cl-en below -78C (1,5).
of Cnlijornia,
@ 1973 by Academic Press, of reproduction in any form
Inc. reserved.
Berkeley,
Calijornicr gC,iZO
In freeze-drying ice nearest the outer surface sublimes first, leaving n relat,ivel>. sharp boundary between ice-free and frozen regions. Heat is conducted from a sourre to the point of sublimat,ion, and vapor flows outward through pores left b!- sublimed ire crystals. The operating temperature of t)he frozen zone is determined by the czhamber pressure and the heat source temperature through the int,eract,ion of simultaneous heat and milss transfer processes. similar in principle to the wet-bulb thermometer (11. CM forms a matrix between the ict= needles and r&ins water for some t,ime after local sublimation of the ice. For frozen-zone temperatures below the collapse temperature. this matrix is highly viscous and acts as a structurally rigid solid. Above t,he collapse temperature the CAS becomes more fluid, and the surface tension of the moist CAS matrix causes distortion and collapse of the product. In nearly all c;ises surface tension will dominate over gravity in cnusing collapse (I). il simple equation may bc derived from an energy balance between surfarc cncrgy dccrcase and viscous dissipation for a single empty capillary in an infinite medium i 1 I : (1) where t is the t’ime for collapse, y is the surface tension, p is the viscosity of the CAS. and R is the initial capillary (ice needle) radius. Typical values for frozen liquid foods are R = 20 pm and 7 = 70 dynes/cm. Assuming that the evaporative drying time of the lamellae between ice crystals (between 1 and 10” Fee in commercial freeze-dryers) must be less than t. this analysis predicts that collapse will occur when the CBS viscosity is of the order 107-10’o cP. For other applications, such as histology with optical or electron microscopy, different freezing techniques may cause R to be considerably less, thereby increasing the critical viscosity required for collapse. CAS viscosity tends to increase with decreas-
Received September 19.1972 Copyright All rights
C. JUDSON KING
559
560
BELLOWS
AND KING (-60 to -lOC), background pressure (60-300 ,um Hg), and sample weight being recorded at regular intervals (1). Power input to an overhead radiant heater was continually increased, causing a progressive increase in the frozen zone TABLJ<: 1 COLLAPSE TEMPERATURES OF VARIOUS SOLUTIONS
Solute, initial wt y0
Collapse
Temperature
FIG. 1. Equilibrium viscosity tions of various sugars.
PC)
curve for solu-
ing temperature, increasing solute concentration and increasing solute molecular weight. If the chemical potential of water is equal between the ice and CAS phases and if the ice crystals are large enough so tallat, curvature effects may be ignored, the CAS concentration is uniquely determined thermodynamically by the temperature and by the molecular weight and nonideality of the solutes. Therefore, for a given solute or mix of solutes the CAS has a unique viscosity-temperature relationship, which represents the effect of temperature, both directly and through changing solute concentration. Figure 1 shows this relationship for various solutes (dark circles), based upon measured viscosities (1). The sucrose-glucose mixture is 50 wt % of each: results are also presented for a 10 wt % NaCl-90 wt % sucrose mixture. These “equilibrium \-iscosity” curves are exceedingly strong functions of temperature, because lowering temperature concentrates the CAS. Consequently, an order-of-magnitude viscosity analysis is suffirient for quantitative prediction of the collapse temperature. Sugar solut’ions show markedly different equilibrium viscosity curves depending upon whether they are composed of mono-, di-, or trisaccharides, with higher molecular-weight solutes having higher viscosities, primarily because the CAS concentration (wt %) is higher at any given temperature. Sample slabs of numerous aqueous solutions were frozen in sample holders and freeze-dried with frozen-zone temperature experimentally.
Sugars Xylose 257 1,E’ruc;ose,‘25”; u-Glucose, 255; Sucrose
Average Observed collapse molecular temperawt ture (C) 150 180 180
-49 -44 -41.5
15% 25% 35% 455; 55% 12.5% Glucose-12.5c;
342 342 34:! 342 342 237
-22.5 -24 -27 -29 -29.5 -33.5
sucrose Maltose, 257; Xaffinose, 259’ ,r
342 504
-23 -18
167a
-33.5
1100 145
-43 -47.5
164
-46
238
-37.5
Sugars, with additives 2.5yc NaCl-22.5% SLIcrose 57; NaCl-200/;, sucrose 10yc Dimethyl sulfoxide15yc sucrose loo/; Glycerine-157; sucrose 12.5cc Sorbitol-12.57; sucrose lc;C Pect,in-240j, fructose 2”/ Gelatin-234; frrlctose Food liquids Orange juice 23%* Grapefruit j;ice, lV’ih Lemon juice 9ycb Apple juice, ‘22ycb Concord grape juice,
-180 -180
-34 -27.5
-277 -228 -194 -180
-24 -30.5 -36.5 -41.5 -46
-261
-33.5
-180 -180
-41.5 -35.0 -20
-194
:6%* Sweetened concord grape juice, 23y( * Pineapple juice, 10%* Prune extract 20?‘* < Coffee extract: 2555 a NaCl
assumed
* Approximate concentration).
completely ionized. (derived from natural
level of
OPERATING
TEMPERATURE
temperature during drying. Drying rates increased in rough proportion to the vapor pressure of ice at the frozen zone temperature, until collapse, when an abrupt discontinuity in the drying rnt,e was observed along with incipient surface irregularities. Measured collapse temperatures are reported in Table 1 for various solutions of approximately 25 wt r/o solutes, and for sucrose solutions of sel-era1 concent,rat’ions. Measured collapse temperatures are included as open circles in Fig. 1 and show good agreement with the collapse range predicted from Eq. (I). The solutes in fruit juices are over 85% monoand disaccharides (2). Pure sugars, mixtures of sugars and fruit juices all show that the collapse temperature increases with increasing molecular weight, as predict,ed. Other reported rollertions of observed collapse temperatures are given by MacKenzie (6) and Ito (3) ; thrse show the same molecular weight trend. Two different effects of nonsaccharide components on collapse temperature are evident. When molecules are small relative to sucrose-such as NaCl, DMSO, glycerine, or sorbitol-and are added to sucrose, they lower the CBS caoncentrat’ion by depressing t’he freezing point and hence lower t)he equilibrium viscosity at any temperature. This causes the observed collapse temperat,ure to decrease. Small amounts of large molecules, such as pectin and gelatin, do not affect t,he C.4S concentration much, but do directly increase the CAS viscosity and hence the collapse temperature. Similar behavior has been observed experimentally by MacKenzie (6,7). Increasing the initial concentration of solutes before freezing decreases the collapse temperature somewhat, because the volumet,ric ratio of CAS to ice increases and the CBS dries more slowly. This allows more time for collapse and requires a somewhat higher equilibrium Gcosity t’o prevent collapse [Eq. (l)].
FOR FREEZE-DRYING
561
Very high initial solute concentrations (i.e., ah-e 70 wt 5/o sucrose) do not nucleat’e ice readily (1). Unless ice is formed during freezing the product must dry by evaporation rather than by sublimation. Evaporative drying will give inferior product quality and los-: of st’rucbeyond ture; thus, the solute concentration w-hi& ice will not nucleate represents an upper limit on the concentrat’ion of solutions to be freeze-dried. ACKNOWLEDGMES’T This work was supported by the Western Marketing and Nutrition Research Division, Agricultural Research Service, U. S. Dept. of Agriculture, Berkeley. CA, as part of Grant 12-14-100~9906(74). REFERENCES 1. Bellows. R. J.. and King, C. J. Paper presented at Annual Meeting, American Instit.ute of Chemical Engineers, New York, NY, Nov. 28, 1972; Chem. Eng. Progr. Symp. Series, in press; Bellows, R. J., Ph.D. Dissertation, University of California, Berkeley, 1972. 2. Copley, M. J., and Van Arsdel. W. B. “Food Dehydration-Vol. II, Products and Technology,” p. 521, AVI Publishing Co., Westport, CT (1964). 3. Ito, K.. Chem. Pharm. &11., 18, 139-1518, 151g-1525 (1970); 19,1095-1102 (1971). 4. King. C. J., CRC Critical Reviews in Food Technology, 1, 379-451 (1970) ; “Freeze-Drying of Foods,” Chemical Rubber Publ. Co.. Cleveland, Ohio, 1971. 5. Lusena, C. B., Ann. X. Y. Acad. Sci., 85, 541548 (1960) ; Luyet. B. J. Z~L“Ciba Foundation Symposium on the Frozen Cell>’ (G. E. W. Wolstenholme and M. O’Connor, eds.). pp. 27-50, Churchill. London. 1970. 6. MacKenzie, d. P., Bull. Parentera D,xg Ass. 20, 101-129 (1966); Ann. h;. Y. Acad. Sci. 125, 522-547 (1965); Cryobiology 3, 387 (1967). 7. MacKenzie, A. P., private communication.
9,
559-561 (1972)
Freeze-Drying of Aqueoue Allowable Operating RICHARD Department
J. BELLOWS
of Chemical Engineering,
Solutions : Maximum Temperature
AND
University
Freeze-drying (Iyophilization) is a two-step dehydration process by which a substance is first frozen, and then water is removed by sublimation under vacuum. Freeze-drying has a diverse range of application, including the preparation of biological samples, pharmaceutical manufacture, and food preservation. Freeze-dried liquids show an abrupt, loss in product quality when the temperature of the subliming interface during drying is maintained above a critical value, t’he “collapse” temperature (6). Drying above this temperature gives a distorted (collapsed) product with poor solubility, low drying rates, uneven drying, and loss of texture and volatile substances. We find that, in the absence of solute crystallization, the collapse temperature depends principally upon the viscosity of the unfrozen port,ion of the solution. -4 simple theory, presentcd below, shows that increasing solute molecular weight gives higher collapse temperatures, and predict’s observed collapse temperatures quantitatively. Det>ails of experimental procedure, further background for the theory, and comparison with other reported experimental collapse temperatures are given elsewhere (1). Freezing causes a microseparation of an aqueous solution into a two-phase mixture of ice and a solute-rich phase. Some solutes crystallize readily, and for such solutions a eutectic mixture of ice and crystallized solut,e forms. Ito (3) has shown that solutions of eutectic-forming solutes collapse at their eutectic temperature during sublimation. Commonly, however, many organic solutes and especially mixed solutes such as occur in natural substances are kinetically inhibited from exhibiting eutect)ic behavior. Freezing these solutions gives a microscalc mixt,ure of ice needles and concentrated amorphous solution (CaS). The CAS remains an amorphous aqueous solution or a glass. cl-en below -78C (1,5).
of Cnlijornia,
@ 1973 by Academic Press, of reproduction in any form
Inc. reserved.
Berkeley,
Calijornicr gC,iZO
In freeze-drying ice nearest the outer surface sublimes first, leaving n relat,ivel>. sharp boundary between ice-free and frozen regions. Heat is conducted from a sourre to the point of sublimat,ion, and vapor flows outward through pores left b!- sublimed ire crystals. The operating temperature of t)he frozen zone is determined by the czhamber pressure and the heat source temperature through the int,eract,ion of simultaneous heat and milss transfer processes. similar in principle to the wet-bulb thermometer (11. CM forms a matrix between the ict= needles and r&ins water for some t,ime after local sublimation of the ice. For frozen-zone temperatures below the collapse temperature. this matrix is highly viscous and acts as a structurally rigid solid. Above t,he collapse temperature the CAS becomes more fluid, and the surface tension of the moist CAS matrix causes distortion and collapse of the product. In nearly all c;ises surface tension will dominate over gravity in cnusing collapse (I). il simple equation may bc derived from an energy balance between surfarc cncrgy dccrcase and viscous dissipation for a single empty capillary in an infinite medium i 1 I : (1) where t is the t’ime for collapse, y is the surface tension, p is the viscosity of the CAS. and R is the initial capillary (ice needle) radius. Typical values for frozen liquid foods are R = 20 pm and 7 = 70 dynes/cm. Assuming that the evaporative drying time of the lamellae between ice crystals (between 1 and 10” Fee in commercial freeze-dryers) must be less than t. this analysis predicts that collapse will occur when the CBS viscosity is of the order 107-10’o cP. For other applications, such as histology with optical or electron microscopy, different freezing techniques may cause R to be considerably less, thereby increasing the critical viscosity required for collapse. CAS viscosity tends to increase with decreas-
Received September 19.1972 Copyright All rights
C. JUDSON KING
559
560
BELLOWS
AND KING (-60 to -lOC), background pressure (60-300 ,um Hg), and sample weight being recorded at regular intervals (1). Power input to an overhead radiant heater was continually increased, causing a progressive increase in the frozen zone TABLJ<: 1 COLLAPSE TEMPERATURES OF VARIOUS SOLUTIONS
Solute, initial wt y0
Collapse
Temperature
FIG. 1. Equilibrium viscosity tions of various sugars.
PC)
curve for solu-
ing temperature, increasing solute concentration and increasing solute molecular weight. If the chemical potential of water is equal between the ice and CAS phases and if the ice crystals are large enough so tallat, curvature effects may be ignored, the CAS concentration is uniquely determined thermodynamically by the temperature and by the molecular weight and nonideality of the solutes. Therefore, for a given solute or mix of solutes the CAS has a unique viscosity-temperature relationship, which represents the effect of temperature, both directly and through changing solute concentration. Figure 1 shows this relationship for various solutes (dark circles), based upon measured viscosities (1). The sucrose-glucose mixture is 50 wt % of each: results are also presented for a 10 wt % NaCl-90 wt % sucrose mixture. These “equilibrium \-iscosity” curves are exceedingly strong functions of temperature, because lowering temperature concentrates the CAS. Consequently, an order-of-magnitude viscosity analysis is suffirient for quantitative prediction of the collapse temperature. Sugar solut’ions show markedly different equilibrium viscosity curves depending upon whether they are composed of mono-, di-, or trisaccharides, with higher molecular-weight solutes having higher viscosities, primarily because the CAS concentration (wt %) is higher at any given temperature. Sample slabs of numerous aqueous solutions were frozen in sample holders and freeze-dried with frozen-zone temperature experimentally.
Sugars Xylose 257 1,E’ruc;ose,‘25”; u-Glucose, 255; Sucrose
Average Observed collapse molecular temperawt ture (C) 150 180 180
-49 -44 -41.5
15% 25% 35% 455; 55% 12.5% Glucose-12.5c;
342 342 34:! 342 342 237
-22.5 -24 -27 -29 -29.5 -33.5
sucrose Maltose, 257; Xaffinose, 259’ ,r
342 504
-23 -18
167a
-33.5
1100 145
-43 -47.5
164
-46
238
-37.5
Sugars, with additives 2.5yc NaCl-22.5% SLIcrose 57; NaCl-200/;, sucrose 10yc Dimethyl sulfoxide15yc sucrose loo/; Glycerine-157; sucrose 12.5cc Sorbitol-12.57; sucrose lc;C Pect,in-240j, fructose 2”/ Gelatin-234; frrlctose Food liquids Orange juice 23%* Grapefruit j;ice, lV’ih Lemon juice 9ycb Apple juice, ‘22ycb Concord grape juice,
-180 -180
-34 -27.5
-277 -228 -194 -180
-24 -30.5 -36.5 -41.5 -46
-261
-33.5
-180 -180
-41.5 -35.0 -20
-194
:6%* Sweetened concord grape juice, 23y( * Pineapple juice, 10%* Prune extract 20?‘* < Coffee extract: 2555 a NaCl
assumed
* Approximate concentration).
completely ionized. (derived from natural
level of
OPERATING
TEMPERATURE
temperature during drying. Drying rates increased in rough proportion to the vapor pressure of ice at the frozen zone temperature, until collapse, when an abrupt discontinuity in the drying rnt,e was observed along with incipient surface irregularities. Measured collapse temperatures are reported in Table 1 for various solutions of approximately 25 wt r/o solutes, and for sucrose solutions of sel-era1 concent,rat’ions. Measured collapse temperatures are included as open circles in Fig. 1 and show good agreement with the collapse range predicted from Eq. (I). The solutes in fruit juices are over 85% monoand disaccharides (2). Pure sugars, mixtures of sugars and fruit juices all show that the collapse temperature increases with increasing molecular weight, as predict,ed. Other reported rollertions of observed collapse temperatures are given by MacKenzie (6) and Ito (3) ; thrse show the same molecular weight trend. Two different effects of nonsaccharide components on collapse temperature are evident. When molecules are small relative to sucrose-such as NaCl, DMSO, glycerine, or sorbitol-and are added to sucrose, they lower the CBS caoncentrat’ion by depressing t’he freezing point and hence lower t)he equilibrium viscosity at any temperature. This causes the observed collapse temperat,ure to decrease. Small amounts of large molecules, such as pectin and gelatin, do not affect t,he C.4S concentration much, but do directly increase the CAS viscosity and hence the collapse temperature. Similar behavior has been observed experimentally by MacKenzie (6,7). Increasing the initial concentration of solutes before freezing decreases the collapse temperature somewhat, because the volumet,ric ratio of CAS to ice increases and the CBS dries more slowly. This allows more time for collapse and requires a somewhat higher equilibrium Gcosity t’o prevent collapse [Eq. (l)].
FOR FREEZE-DRYING
561
Very high initial solute concentrations (i.e., ah-e 70 wt 5/o sucrose) do not nucleat’e ice readily (1). Unless ice is formed during freezing the product must dry by evaporation rather than by sublimation. Evaporative drying will give inferior product quality and los-: of st’rucbeyond ture; thus, the solute concentration w-hi& ice will not nucleate represents an upper limit on the concentrat’ion of solutions to be freeze-dried. ACKNOWLEDGMES’T This work was supported by the Western Marketing and Nutrition Research Division, Agricultural Research Service, U. S. Dept. of Agriculture, Berkeley. CA, as part of Grant 12-14-100~9906(74). REFERENCES 1. Bellows. R. J.. and King, C. J. Paper presented at Annual Meeting, American Instit.ute of Chemical Engineers, New York, NY, Nov. 28, 1972; Chem. Eng. Progr. Symp. Series, in press; Bellows, R. J., Ph.D. Dissertation, University of California, Berkeley, 1972. 2. Copley, M. J., and Van Arsdel. W. B. “Food Dehydration-Vol. II, Products and Technology,” p. 521, AVI Publishing Co., Westport, CT (1964). 3. Ito, K.. Chem. Pharm. &11., 18, 139-1518, 151g-1525 (1970); 19,1095-1102 (1971). 4. King. C. J., CRC Critical Reviews in Food Technology, 1, 379-451 (1970) ; “Freeze-Drying of Foods,” Chemical Rubber Publ. Co.. Cleveland, Ohio, 1971. 5. Lusena, C. B., Ann. X. Y. Acad. Sci., 85, 541548 (1960) ; Luyet. B. J. Z~L“Ciba Foundation Symposium on the Frozen Cell>’ (G. E. W. Wolstenholme and M. O’Connor, eds.). pp. 27-50, Churchill. London. 1970. 6. MacKenzie, d. P., Bull. Parentera D,xg Ass. 20, 101-129 (1966); Ann. h;. Y. Acad. Sci. 125, 522-547 (1965); Cryobiology 3, 387 (1967). 7. MacKenzie, A. P., private communication.