General Principles of Freeze Drying (The Lyophilization Process)

Introduction Application and Uses Freeze Drying, or lyophilization as it is referred to in the Pharmaceutical and Diagnostic Industries, is a dehydration technique, which enables liquid or slurry products, which have previously been frozen to be dried under a vacuum. The applications of freeze drying are numerous, but is generally employed when the requirements demand:
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Preservation of temperature sensitive products, particularly those of biological origin, such as enzymes, blood plasma, vaccines, etc. To achieve a chemical balance, such as for biological reagents. To provide a practical solution for certain delivery problems, for example, the packaging of constituents that cannot be mixed in the liquid state, but which are solidified in successive stages and then freeze dried. To implement an important stage of a product (such as concentration). To improve storage life and improved marketing of the end product. To resolve certain filling problems. It may be difficult, for instance, to divide several milligrams of powder into precise vial dosages, due to the difficulty of measuring tiny amounts, homogeneity, granulation, static electricity etc.. The distribution of the product from the liquid state eliminates such production problems. Product temperature sensitivity, and its relation to taste.

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Description of the Operation Generally, the Freeze Drying, or Lyophilization cycle is divided in three phases: An initial freezing process, carried out in such a way that:
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The product exhibits the desired crystalline structure. The product is frozen below its eutectic temperature. A primary drying (sublimation) phase during which:

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The partial pressure of the vapor surrounding the product must be lower than the pressure of the vapor from the ice, at the same temperature. The energy supplied in the form of heat must remain lower than the product's eutectic temperature (the highest allowable product temperature during the conditions of sublimation.) A secondary drying aimed at eliminating the final traces of water which remain due to absorption, and where:

The partial pressure of the vapor rising from the product will be at its lowest levels.

At the completion of the process, the treated product will have retained its form, volume and original structure-as well as all its physical, chemical and biological properties. It can then be stored (provided packaging is effective to the reduction of moisture migration) for an almost indefinite period of time. As the product is porous, it can be re-dissolved by the simple addition of a proper solvent. From this description of the process of freeze drying, three facts emerge:
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The sublimation characteristics of the product are greatly dependent of the frozen structure. This structure cannot be altered during the process. Product temperature plays an active role in all three phases, and in execution it is upon this that the choice of other parameters (vaccuum, heat rate, etc.) is based.

Theoretical Basis of Freeze Drying The theoretical principle of freeze drying is clearly defined in the diagram "Pressure Temperature" (Fig A). In order to avoid the liquid phase, it is absolutely essential to lower the partial pressure of water, below the triple point pressure. A freeze drying cycle is shown in this diagram, which has been designed to conform to a typical example (described below):
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Freezing of a product from 20 C to -20 C at atmospheric pressure. Sublimation of the product at -20 C. Transfer of evolved vapor to the condenser at low temperature. Vacuum release. Defrost.

Freeze Drying is a complex operation, and all facets cannot be addressed in this type of explanation. Instead, certain aspects will be highlighted which play a part in the development of a freeze drying operation:
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Freezing. Drying. Vacuum influence. The liquid shelf on which the product is placed. Essential control aspects during freeze drying.

Freezing
Upon completion of product freezing, the product will have acquired a frozen structure, which cannot be changed during freeze drying. Sublimation, and the qualities of the finished product are greatly dependent on this crystal structure. In fact, it is considered the most crucial stage of the freeze drying process.

Speed of Freezing

On the pilot level, fast or very fast freezing is relatively easy to achieve. However, for industrial production settings, freezing at the same rates is unrealistic because of the problems of product preparation (filling, loading time) and larger systems costs will dictate compromises in the same process. From Fig 1 we observe: As soon as the product reaches 0 C (Point A on the curve of Fig 1), some of the particles transform to ice. This is the nucleation process. Generally, biological products contain between 80% and 95% water. Observe that the temperature of the product stabilizes after time period at about 0 C. At Point B, the ice crystals previously formed have expanded, and consist practically of pure water. At Point C, the crystals have grown larger, and now occupy 80% to 90% of the initial volume of the solution. The crystallization of the free water is nearly complete. These crystals seem to be contained in an interstitial state, still liquid, but which constitutes the principal active element of the solution. At Point D, the interstitial component itself has reached freezing temperature, and the amorphous appearance is even more apparent, and a barely visible skin has formed on the surface. This structure is ideal for sublimation. We now have a paradoxical situation: a slow cooling which can lead to a rapid coagulation of the constituent water. In many cases, freezing induced by these conditions may be necessary to achieve successful freeze drying of a sensitive product.

Freezing Temperature
In these examples, Point D on the curve, Fig 1 and Fig 2 represent the temperature of the complete freezing of the product. The establishment of this eutectic zone is very important. Between C and D, the concentration and consistency of the liquid phase is increased, and in the case of biological products, may produce a change in the bacteria as a result of the hyper concentration of the active ingredient, and the mechanical effect of the ice crystals. During the sublimation phase, signs of melting appear in the product, which induces a temperature higher than that corresponding to the eutectic point. Temperature analysis permits determination of the onset of the fusion temperature. This acts as an indicator to help prevent melt back, or other such accidents during the course of freeze drying. Generally, a structure of large crystals presents more difficulties with regard to freeze drying, in that a thick crust forms at the surface. The appearance of the freeze dried product is heterogeneous. This very often makes dissolution difficult. A product structure of fine crystals freezes more easily. The freeze dried product has an amorphous appearance, and re-dissolves more quickly. Obtaining the desired crystalline structure is not always easy, as the formation of this type of crystal depends on several factors:

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The nature of the product. The processing. Freezing speed. Type of freezing.

Given the diverse range of products, which may be treated in the dryer, the desired freezing rate, and the type of final packaging that may used, the manufacturer must consider:
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The quantity of product per container. The form of the container. The type of freezing. And, plan for a freeze dryer that is flexible to these different demands.

Freeze Drying
Once the product is properly frozen, it must be sublimated (evaporated) at a low temperature under reduced pressure. The ideal curve of lyophilization is depicted on Curve A of Fig. 3, with temperature displayed on the ordinand line, and time displayed on the abscissa. With the product maintained at a constant temperature, it will be necessary to supply the energy of sublimation, a combination of the latent heat of fusion (which supplies the transformation of the liquid to the ice state) and the sublimation energy (about 700 calories per gram of ice evaporated). The ability of vapor release from the matrix is a function of molecular agitation inside the matrix. Ideally, the temperature of the frozen product should be brought to the highest temperature compatible with the frozen condition, without exceeding it, which will lead to the irreparable production of foam (commonly known as melt back) and product deterioration. On the other hand, if the heat energy is insufficient, the product will sublime at too low a temperature (Curve E), and the length of the freeze drying cycle will become abnormally long. After the disappearance of the final ice crystals, the temperature of the product rapidly increases, and must be maintained at the most maximum permissible temperature to liberate the lowest residual moisture embedded in the matrix (secondary drying). The liquid shelves on which the product is loaded transfer the required energy of sublimation. (Curves B, C, or D represent the heating rate) A chilled surface known as the ice condenser collects the vapor from the evolving product. (Curve F) During lyophilization, the pressure in the drying chamber follows the fluctuations identified by Curves P or P1.

Importance of Vacuum in the Freeze Drying Process

Freeze Drying can only take place if the partial pressure of the vapor in the drying chamber is lower than the water vapor pressure above the product. By strict definition, the vacuum in the

chamber is not essential, as sublimation can take place at atmospheric pressure by passing dehydrated air above the product. By artificially lowering the pressure in the drying chamber the time can be reduced. For example, a product sublimated at 20 C (Fig 4) at the given vapor pressure shown as 0.8 torr. As soon as the value in the chamber reaches below 0.8 torr, the ice begins to sublime. Flosdorff has shown that as soon as the pressure in the chamber is reduced, evaporation increases, but the rate of evaporation is not without limit, and reaches a maximum when the pressure in the chamber has a value equal to about 50% of the vapor pressure above the product. The curve in Fig. 5 represents the changes in evaporation rate for a product which is undergoing sublimation at 20 C, as a function of the pressure in the chamber; the rate reaching a maximum of 0.4 torr. Contrary to widely held opinion, it is not necessary to have a very low vacuum during the sublimation period, because below the limit defined, the evaporation rate is not improved, and that too low a pressure acts as a barrier to effective heat transfer.

Freeze Drying
Once the product is properly frozen, it must be sublimated (evaporated) at a low temperature under reduced pressure. The ideal curve of lyophilization is depicted on Curve A of Fig. 3, with temperature displayed on the ordinand line, and time displayed on the abscissa. With the product maintained at a constant temperature, it will be necessary to supply the energy of sublimation, a combination of the latent heat of fusion (which supplies the transformation of the liquid to the ice state) and the sublimation energy (about 700 calories per gram of ice evaporated). The ability of vapor release from the matrix is a function of molecular agitation inside the matrix. Ideally, the temperature of the frozen product should be brought to the highest temperature compatible with the frozen condition, without exceeding it, which will lead to the irreparable production of foam (commonly known as melt back) and product deterioration. On the other hand, if the heat energy is insufficient, the product will sublime at too low a temperature (Curve E), and the length of the freeze drying cycle will become abnormally long. After the disappearance of the final ice crystals, the temperature of the product rapidly increases, and must be maintained at the most maximum permissible temperature to liberate the lowest residual moisture embedded in the matrix (secondary drying). The liquid shelves on which the product is loaded transfer the required energy of sublimation. (Curves B, C, or D represent the heating rate)

A chilled surface known as the ice condenser collects the vapor from the evolving product. (Curve F) During lyophilization, the pressure in the drying chamber follows the fluctuations identified by Curves P or P1. Conclusion: By controlling chamber pressure, you can reduce the heat transfer coefficient between the warm shelves and the product. This simple control will greatly improve the energy transfer and reduce primary drying times. The curve of shelf temperature is shown in schematic form in B of Fig. 3, and the vacuum curve in P. The control of vacuum in the lyophilization process can become a useful means of controlling heat transfer, and the means of getting energy to the product. A laboratory example illustrates the influence of pressure in heat transfer: A specially fitted freeze dryer, equipped with the necessary measurement and control equipment was used to simulate these sublimation phenomena. Freeze drying of the product was carried out at 20 C, with a controlled pressure in the chamber in the range of 0.4 torr, and a shelf temperature of 30 C. When the injection of non-condensable gas was terminated, the product cooled rapidly, and the vapor removal rate slowed. To regain the sublimation temperature of 20C, it was necessary to bring the shelf temperature to 125 C, a difference of 95 C in the heating source to produce the same sublimation temperature of the product, and virtually the same evaporation rate! The chamber pressure acts as a thermal regulator, which can, in the space of a few moments can produce the same effect as raising the temperature 95 C.

Rate of Vapor Transfer to the Condenser:

Thermodynamic formulae enable the calculation of values of the volume occupied by vapor as a function of chamber pressure. For one gram of water, this volume is in the region of 100m3 at a pressure of 10-2 torr, and 1000m3 at a pressure of 10-3 torr. Consider a product with a eutectic of about 20C, freeze dried under the following conditions:
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500 ml bottle filled with 300ml liquid product. A water content of approximately 250 grams. An evaporation surface of about 0.02 sq. meters. A chamber pressure of 2 x 10-2 torr.

The length of primary drying (sublimation of the ice) around ten hours. One gram of water occupies, under this pressure, a volume of 50m3, and the amount of water vapor which sublimates from the product per second can be calculated from the following formula:
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50m3 x 250 = 0.35m3 per/second

10 hrs x 3600

The surface area inside the bottle being around 0.02m2, the vapor leaves the product at a speed of:
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0.350m3/s = 17.5 m/s 0.02 m2

Inside the neck of the bottle, which has a diameter of about 3mm, or a section of 7cm2 (7 x 10-4m2), the speed is calculated at:
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0.350m3/s = 500 m/s 7 x 10-4 m3

This illustrates a very high rate advantageous to shorter drying times. But, as the product becomes lighter (the kinetic energy of the vapor is proportional to the square rate), the crossing of an already dry boundary carries product in the flow (as much as 10 to 20%), and deposits itself on the condenser where it is virtually irrecoverable. In addition, these light product particles can pollute the chamber area, where sensitive sensing equipment may be located. When a non-condensable gas is introduced to raise the pressure level, say to 0.2 torr, the volume occupied by one gram of water (about 5m3), the speed through the neck of the container is about 50 m/s. In short, the kinetic energy of the vapor is divided by 100, and the amount of product entrained in the vapor flow is reduced to negligible amounts.

With Pressure x Volume Constant:

For one (1) gram of water at:
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Atmospheric pressure (760 torr) we derive: 760 x 22.4 (l) 1000 At P=1 torr, V=1000 l or 1m3 At P=10-1, V=1000, = 10,000 l, or 10m3 At P=10-2 V=100m3 At P=10-3 V=1000m3

Pressure Vapor Velocity, Vapor Velocity, Kinetic Energy

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Product container neck vapor (1/2mV2) 2x10-2 torr 17.5 m/s 500 m/s 100X 2x10-1 torr 1.75 m/s 50 m/s 1000X At the end of the sublimation phase, there is sufficient time to re-establish very low pressure to create the pressure differential conditions needed for terminal drying.

The Role of the Condenser:

Given the thousands of cubic meters of water vapor emitted from the product in a freeze dryer, no vacuum pump, regardless of rate, could remove these volumes directly. Therefore, a vapor trap, or condenser is essential to condense the evolving water molecules. As the role of the vacuum pump is limited to eliminating the tiny traces of non-condensable gases, one could conclude that the real vacuum effect is derived from the ice condenser (via the pressure differential created by the surface temperature of the product and the condenser). If you examine the curve of Fig 4, you can note that each temperature value for condensed ice corresponds to a vapor pressure, the value of which is always lower than the chamber pressure. For example, if a machine is equipped with an ice condenser for which the lowest temperature limit is 40 C (Fig 4), it would be impossible to lower the pressure below the vapor pressure corresponding to 40 C, i.e. 10-1 torr. This value is irrespective to the type or design of the vacuum pump. A failure in the refrigeration system leads to a chain reaction:
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A rise in condenser temperature. A rise in chamber pressure. A rise in product temperature. An irreversible eutectic melting and the boiling of liquid fractions. Product failure.

Because of the importance of the refrigeration system, a freeze dryer must be equipped with a condenser designed and constructed with the ability to:
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Condense all vapors from the product. Provide a vapor route of minimum distance to avoid hindering vapor flow. Permit easy defrosting after the run. Prevent vapors from contaminating the oil in the vacuum pump. Provide a simple cleaning operation. Provide the necessary BTU output under load to condense vapor at a maximum rate without disturbing the products selected primary sublimation temperature. Insure the necessary low temperature (saturated suction) during the secondary drying to deliver the lower vacuum levels needed for this phase. Provide a high degree of reliability.

Heating Control in the Freeze Drying Process:

The transmission of energy to the product needs to be carefully controlled for three important considerations:
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To avoid transferring too much heat and passing the safe primary drying temperature. To avoid supplying insufficient heat, hence prolonging the sublimation period.

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To achieve a homogeneous temperature in the total batch, as the lyophilization cycle will be determined by the temperature of the coldest product area. The higher the sublimation temperature, the faster the drying cycle. The conditions of low temperature and high vacuum, can have, if they are unnecessarily prolonged, an effect on living organisms, which form the great bulk of freeze dried products.

Production Application:
In industrial freeze drying applications it is essential to reproduce indentical drying protocols for a wide range of products. To accomplish this goal, it is vital to insure the same parameters for each operation. The essential criteria are:
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The temperature of the product. The duration of the primary and secondary drying phases. Heat input. Chamber pressure.

By a series of reproducible runs on a scaleable the optimum cycle can be established. If the process is automated it is essential that:
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Cycle is performed with a guarantee of functional security. Validation is simple. Allows maximum flexibility for a variety of product drying protocols. In the pilot development phase, it is critical that the favorable conditions for freezing and the optimum parameters of primary and secondary drying be established.

Secondary Drying:
At the end of the sublimation phase (primary drying), all the ice will have disappeared. The product will begin to rise in temperature, and will tend to approach the control temperature of the shelf. However, at this stage the product is not sufficiently dry for long term storage. For most products, the residual moisture is in the region of 5% to 7%. The product now enters the desorption phase, during which the last traces of water vapor are removed, along with traces of the bound water within the product matrix. This phase is identified as secondary drying. The aim of this final phase is to reduce the product to the acceptable moisture levels needed for long term storage (3% to 1%). The reasons for drying the product to these levels are desirable for several benefits:
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When the water content is higher than these levels, the product will denature. When the residual moisture is forced lower than these levels, many products may undergo chemical or enzymatic changes.

Residual moisture in the product is generally dependent on four factors:

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The product matrix (both in frozen and sublimation mode.) The vacuum in the drying chamber.

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The duration of secondary drying (isothermic phase). The maximum temperature allowed in the product during secondary drying.

Note: Products with excipients may exhibit tendencies to retain water in the matrix, and are hygroscopic in ambient air. The final residual moisture obtainable is dependent on more conditions than the vacuum and moderate temperature generally employed for this secondary phase. Good vacuum is essential so that the mean free path of water molecules from the product matrix is not impeded by pressure. The lowest possible pressure is not necessarily desirable (below 10-2 torr) because below this pressure, desorption produces an assympotic rate (Fig. 7), and the phenomenon of backstreaming the vacuum pump becomes more significant, thus risking pollution the product matrix.

Factors affecting secondary (terminal) drying phase:

By a series of reproducible runs on a scaleable the optimum cycle can be established. If the process is automated it is essential that:
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Cycle is performed with a guarantee of functional security. Validation is simple. Allows maximum flexibility for a variety of product drying protocols. In the pilot development phase, it is critical that the favorable conditions for freezing and the optimum parameters of primary and secondary drying be established.

Secondary Drying:
At the end of the sublimation phase (primary drying), all the ice will have disappeared. The product will begin to rise in temperature, and will tend to approach the control temperature of the shelf. However, at this stage the product is not sufficiently dry for long term storage. For most products, the residual moisture is in the region of 5% to 7%. The product now enters the desorption phase, during which the last traces of water vapor are removed, along with traces of the bound water within the product matrix. This phase is identified as secondary drying. The aim of this final phase is to reduce the product to the acceptable moisture levels needed for long term storage (3% to 1%). The reasons for drying the product to these levels are desirable for several benefits:
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When the water content is higher than these levels, the product will denature. When the residual moisture is forced lower than these levels, many products may undergo chemical or enzymatic changes.

Residual moisture in the product is generally dependent on four factors:

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The product matrix (both in frozen and sublimation mode)

The vacuum in the drying chamber

Production Lyophilization
In considering a freeze dryer for production use, the following general guidelines should be a part of your determinations:
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The products lyophilization profile. (A vital stage in making a determination of the production freeze dryers performance characteristics). This work should be carried out on scalable pilot equipment.

Laboratory units, especially those equipped with air cooled refrigeration systems cannot provide the depth of data needed to determine a production cycle. If the user does not have access to an industrial pilot unit, this work can be accomplished on an outsource basis at relatively modest cost. Determine the total length of the process cycle, including all of the following factors:
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Preparation of the equipment. Dryer loading time. Freezing time. Freeze Drying time. Unloading time. Defrosting time. Clean up time required to make the area ready for the next batch.

Heating Power, General Rules

A standard industrial freeze dryer should permit sublimation of approximately one kilogram of water per square meter of surface per hour, considering that the energy of sublimation is about 800 kcal/kg of water sublimated. The heating value of kw/hr=860 kcal/hr. Therefore, the freeze dryer should be furnished with heating capacity equivalent to 1 kw/sq.meter of surface area.

Refrigeration Power, General Rules

The freeze dryers condensing system should be capable of condensing the sublimated vapor liberated from the product at a temperature of 55 C to 65 C, at a rate of 800cal/kg of water, having a theoretical refrigeration capacity of about 1000kcal/hr at -50 C/sq.meter of surface area. It is important that at the end of secondary drying, the refrigeration system must be capable of lowering the temperature of the ice condenser to a temperature low enough to obtain the necessary residual moisture in the product.

Production Cycle Duration, General Rules

Assuming the optimum rate of flow, an estimate for a dryer of approximately 100 square foot capacity would be:
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100 kg of product in bulk (liquid loaded on trays). 1000 kg to 1500kg of stainless steel (product shelves, hoses, etc). 300kg to 500 kg of thermal medium (heated and cooled).

All of the above to be chilled or heated during the drying cycle. If the dryer is equipped with 10kw heating employed 80% of the cycle, the shelves and heat exchangers represent:
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1000kg of stainless steel with a specific heat of .12cal/kg per degree C 500kg of thermal fluid with a specific heat of 0.36kcal/kg per degree C (Silicone) A set point freezing temperature of 40 C and a terminal temperature of 40 C (Shelf)

The amount of energy required would be: Q=100kg x 800kcal + 1000kg x 0.12kcal x 80 C + 500kg x 0.36kcal x 80= 100,000kcal The time of sublimation would be: T= 100,000 --------------------------- = 15 hours 10kw x 860kcal x 0.8 The secondary drying time must be added to this total to deduce the total cycle time. Secondary drying time is dependent on the desired residual moisture content. Addenda Nota

Bulk Drying, General Rules

As the interface separating the dried and frozen regions increases, the frozen portion of the matrix moves toward the lower surface of the frozen material. This upper dry layer is highly insulating to heat flux, but is less sensitive at low pressures to the phenomena of mass transfer (pressure difference between product and condenser induced by a temperature differential). Heat transfer is increased by a pressure rise, but at the expense of an increased resistance to mass transfer. Water vapor migrating through the upper surface of the matrix and the thermal conductivity of the ice layer is two times higher than that of the dry layer. In this condition, water vapor flux is decreased as pressure is increased.