Lyophillization

1. Dr. Prashant L. Pingale
Associate Professor, Dept. of Pharmaceutics
GES’s Sir Dr. M. S. Gosavi College of Pharm. Edu. & Research,
Nashik-422005, INDIA

2. CONTENT
 Introduction,
 Advantages / Disadvantages,
 Characteristics of products
 Principle,
 Steps involved and
 Application of Freeze drying process.
 Component,
 Parameters,
 Construction and Working of Lyophilizer/ Freeze dryer
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3. INTRODUCTION
 Lyophilization means freeze drying.
 The term “lyophilization” describes a process to produce a
product that “loves the dry state”.
 This process comprises three steps: freezing, primary
drying, and secondary drying.
 The most common form of sterile parenteral powder is
freeze dried or lyophilized powder.
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4. Advantages
Stored in dry state, so stability problem is few.
Product is dried without elevated temp.
Good for O2 & air sensitive drugs.
Rapid reconstitution time.
Constituents of dried material remain homogenously dispersed.
Product is process in the liquid form.
Storage of dry material is less expensive than solution form.
In some specialized laboratories, scientists are developing more sophisticated processes that combine
freeze-drying technology with electron microscopy, biochemistry, and refined surgery.
At the same time, the cosmetics industry is increasing its use of Lyophilization to help prepare beauty
masks, hair dyes, and sophisticated supports for face creams.
Chemical industries also are beginning to use freeze-drying to prepare refined chemicals, catalysts, and
selective filters.
Freeze-drying can preserve food and make it very lightweight.
Lyophilization can also be used to increase the shelf life of some pharmaceuticals for many years.
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5. Disadvantages
 Volatile compounds may be removed by high vacuum.
 Expensive unit operation because pumps are more expensive.
 Stability problems associated with individual drugs.
 Some issues associated with sterilization & sterility assurance of dry chamber & aseptic loading
of vials into chamber.
 Freeze-drying is facing difficult challenges as the sensitivity, complexity, and price of treated
products steadily rise.
 New antibiotics and drugs, immunological products, substances derived from genetic
engineering, high molecular weight proteins, and sophisticated peptides are very fragile, difficult
to freeze, and all highly sensitive to residual moisture content.
 Amorphous (glassy) materials do not have a eutectic point, but do have a critical point, below
which the product must be maintained to prevent melt-back or collapse during primary and
secondary drying.
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6. Characteristics of Freeze Dried
Products
 Intact cake
 Sufficient strength to prevent cracking, powdering or collapse
 Uniform color & consistency
 Sufficient dryness to maintain stability
 Sufficient porosity & surface area to permit rapid reconstitution
 Sterile
 Free of pyrogens & particulates
 Chemically stable
 Long-term stability
 Short reconstitution time
 Elegant cake appearance
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7. Characteristics of Freeze Dried Products
 Maintenance of the characteristics of the original dosage form upon reconstitution, including
solution properties; structure or conformation of proteins; and particle-size distribution of
suspensions
 Isotonicity upon reconstitution (in some cases, for bulk solution)
 Today, considering all these issues, we can say that lyophilization;
 Is an increasingly essential tool for the pharmaceutical industry
 Although a highly sophisticated technology, still is far from mature and deserves substantial
fundamental and applied research presents constant challenges for equipment
manufacturers that must provide instruments that can process, in a reproducible and reliable
way, large batches of high therapeutic and material value.
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8. Principle of Lyophilization
 The fundamental principle in freeze-drying is sublimation, the shift from a solid directly into a gas.
 Just like evaporation, sublimation occurs when a molecule gains enough energy to break free from
the molecules around it.
 Water will sublime from a solid (ice) to a gas (vapor) when the molecules have enough energy to
break free but the conditions aren't right for a liquid to form.
 There are two major factors that determine what phase (solid, liquid or gas) a substance will take:
heat and atmospheric pressure.
 For a substance to take any particular phase, the temperature and pressure must be within a
certain range.
 Without these conditions, that phase of the substance can't exist.
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9. Principle of Lyophilization
 We can observe from fig. 1 that, water can
take a liquid form at sea level (where
pressure is equal to 1 atm) if the
temperature is in between the sea level
freezing point (32 degrees Fahrenheit or 0
degrees Celsius) and the sea level boiling
point (212oF or 100oC).
 But if we increase the temperature above
32oF while keeping the atmospheric
pressure below 0.06 atmospheres (ATM),
the water is warm enough to thaw, but there
isn't enough pressure for a liquid to form.
 It becomes a gas.
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10. Sublimation
 Sublimation is when a solid (ice)
changes directly to a vapor without first
going through a liquid (water) phase.
Thoroughly understanding the concept
of sublimation is a key building block to
gaining knowledge of freeze drying.
 As shown in figure, the phase diagram
for water, low pressures are required
for sublimation to take place.
 Sublimation is a phase change and heat
energy must be added to the frozen
product for it to occur.
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11. Sublimation
Sublimation in the freeze drying process can be
described simply as:
FREEZE - The product is completely frozen, usually
in a vial, flask or tray.
VACUUM - The product is then placed under a
deep vacuum, well below the triple point of water.
DRY – Heat energy is then added to the product
causing the ice to sublime.
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12. LYOPHILIZATION CYCLE
Step1]:- Sample Preparation
Step2]:- Freezing
Step3]:- Primary drying
Step4]:- Secondary drying
Step5]:- Final Product
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13. Annealing
 Annealing is an optional step, occasionally used to
crystallize the formulation component. If the solute
separates out in crystalline form, it is known as the
eutectic temperature. In contrast, if an amorphous form is
formed, the temperature is referred to as the glass
transition temperature (Tg).
 Annealing is an additional step involved in freeze drying
process which will help to accelerate primary drying.
 This process involve holding the product at a temperature
for defined period.
 Annealing reduced primary drying rate.
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14. The steps required to lyophilize
Pretreatment / Formulation
Loading / Container (Bulk, Flask, Vials)
Freezing (Thermal Treatment) at atmospheric pressure
Primary Drying (Sublimation) under vacuum
Secondary Drying (Desorption) under vacuum
Backfill & Stoppering (for product in vials) under partial vacuum
Removal of Dried Product from Freeze Dryer
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15. The steps required to lyophilize
In addition to providing an extended shelf-life, successful freeze-drying should
yield a product that has a short reconstitution time with acceptable potency
levels.
The process should be repeatable with well-defined temperature, pressure
and time parameters for each step.
Visual and functional characteristics of the dried product are also important
for many applications.
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16. Freeze drying process
The freeze drying process consists of three stages:
◦ Freezing,
◦ Primary drying, and
◦ Secondary drying.
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17. Freezing
 The freezing process consists of freezing the material.
 In a lab, this is often done by placing the material in a freeze-
drying flask and rotating the flask in a bath, called a shell
freezer, which is cooled by mechanical refrigeration, dry ice and
methanol, or liquid nitrogen.
 On a larger-scale, freezing is usually done using a freeze-drying
machine.
 In this step, it is important to cool the material below its
eutectic point, the lowest temperature at which the solid and
liquid phases of the material can coexist.
 This ensures that sublimation rather than melting will occur in
the following steps.
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18. Freezing
 Larger crystals are easier to freeze-dry.
 To produce larger crystals, the product should be frozen slowly or can be cycled up and
down in temperature.
 This cycling process is called annealing.
 However, in the case of food, or objects with formerly-living cells, large ice crystals will
break the cell walls (discovered by Clarence Birdseye).
 Usually, the freezing temperatures are between −50°C and −80°C.
 The freezing phase is the most critical in the whole freeze-drying process, because the
product can be spoiled if badly done.
 Amorphous (glassy) materials do not have a eutectic point, but do have a critical point,
below which the product must be maintained to prevent melt-back or collapse during
primary and secondary drying.
 Large objects take a few months to freeze-dry.
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19. Freezing
The freezing point can be determined by means of,
◦ Theoretical thermodynamic value
◦ Cryo-microscope
◦ DSC (Differential Scanning Calorimetry)
◦ Measurement of temperature and resistance during the freezing phase
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20. Primary drying
During the primary drying phase, the pressure is lowered (to the
range of a few millibars), and enough heat is supplied to the
material for the water to sublimate.
The amount of heat necessary can be calculated using the
sublimating molecules’ latent heat of sublimation.
In this initial drying phase, about 95% of the water in the material
is sublimated.
This phase may be slow (can be several days in the industry),
because, if too much heat is added, the material’s structure could
be altered.
In this phase, pressure is controlled through the application of
partial vacuum.
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21. Primary drying
The vacuum speeds sublimation, making it useful as a deliberate
drying process.
Furthermore, a cold condenser chamber and/or condenser plates
provide a surface(s) for the water vapor to re-solidify on.
This condenser plays no role in keeping the material frozen; rather,
it prevents water vapor from reaching the vacuum pump, which
could degrade the pump's performance.
Condenser temperatures are typically below −50°C (−60°F). It is
important to note that, in this range of pressure, the heat is
brought mainly by conduction or radiation; the convection effect
can be considered as insignificant.
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22. Secondary drying
The secondary drying phase aims to remove unfrozen water
molecules, since the ice was removed in the primary drying
phase.
This part of the freeze-drying process is governed by the
material’s adsorption isotherms. In this phase, the
temperature is raised higher than in the primary drying
phase, and can even be above 0 °C, to break any physico-
chemical interactions that have formed between the water
molecules and the frozen material.
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23. Secondary drying
Usually, the pressure is also lowered in this stage to
encourage desorption (typically in the range of microbars,
or fractions of a pascal).
However, there are products that benefit from increased
pressure as well.
After the freeze-drying process is complete, the vacuum is
usually broken with an inert gas, such as nitrogen, before
the material is sealed.
At the end of the operation, the final residual water
content in the product is around 1% to 4%, which is
extremely low.
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24. Remember….
Freezing (annealing)
Primary drying (sublimation)
Secondary drying (desorption)
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25. Freeze drying equipment
The main components of freeze drying equipment
are:
◦ Refrigeration System
◦ Vacuum System
◦ Control System
◦ Product Chamber or Manifold
◦ Condenser
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26. Freeze drying equipment
The refrigeration system cools the (ice) condenser located
inside the freeze dryer.
The refrigeration system can also be employed to cool
shelves in the product chamber for the freezing of the
product.
The vacuum system consists of a separate vacuum pump
connected to an airtight condenser and attached product
chamber.
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27. Freeze drying equipment
Control systems vary in complexity and usually include
temperature and pressure sensing ability. Advanced
controllers will allow the programming of a complete
“recipe” for freeze drying and will include options to
monitor how the freeze drying process is progressing.
Choosing a control system for the freeze dryer depends on
the application and use (i.e. lab vs. production).
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28. FREEZE DRYING EQUIPMENT
Product chambers are typically either a manifold with attached flasks,
or, a larger chamber with a system of shelves on which to place the
product.
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29. FREEZE DRYING EQUIPMENT
 The purpose of the condenser is to attract the vapors being sublimed
off of the product.
 Because the condenser is maintained at a lower energy level relative
to the product ice, the vapors condense and turn back into solid form
(ice) in the condenser.
 The sublimated ice accumulates in the condenser and is manually
removed at the end of the freeze drying cycle (defrost step).
 The condenser temperature required is dictated by the freezing point
and collapse temperature of the product.
 The refrigeration system must be able to maintain the temperature
of the condenser substantially below the temperature of the product.
 In shelf freeze dryers, the condenser can be located inside the
product chamber (internal condenser) or in a separate chamber
(external condenser) connected to the product chamber by a vapor
port.
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30. LYOPHILIZATION CONTAINER
REQUIREMENTS
Thermal conductivity
Minimize the moisture
Tight sealing
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31.  This technique widely used for pharmaceuticals to improve
stability and long term storage of labile drugs.
 Lyophilization or Freeze-drying fills an important need in
pharmaceutical manufacturing technology .
 The most common application of pharmaceutical freeze drying is
in the production of injectable dosage forms.
 This process is also used in the production of diagnostics .
PROCESS TO PRODUCE A PRODUCT
THAT “LOVES DRY STATE”
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32. TYPICAL LYOPHILIZATION PROCESS
 The first step that takes place in lyophilization process is component
preparation i.e. the sterile solution should be prepared, compound, mixed,
filtered.
 The filtered solution is filled into containers (vials). Partially insert a
special designed rubber closure onto the vials.
 Aseptically load the vials into a freeze dry chamber.
 Freeze every single solution in every vial below a pre-determine critical
temperature.
 Using appropriate application of temperature and pressure, sublime the
ice from the product.
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33. Solution Powder
33

34. DESIGN OF LYOFREEZER
Fig: Lyofreezer
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35.  Chamber
 Shelves
 Process condenser
 Shelf fluid system
 Refrigeration system
 Vaccum system
 Sensors
 Control system
The Essential Components of lyofreezer include:-
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36. EXCIPIENTS USED IN LYOPHILIZED
FORMULATION
 The design of a lyophilized formulation is dependent on the
requirements of the active pharmaceutical ingredient (API) and
intended route of administration.
 A formulation may consist of one or more excipients that perform
one or more functions.
Excipients for lyophilization usually fit one of the following
categories:
1] Bulking agents
2] Stabilizers
3] Buffering agents
4] Tonicity modifiers, surface-active agents
5] Collapse temperature modifiers
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37. Buffers
Buffers are required in pharmaceutical formulations to stabilize pH. E.g.
Phosphate buffers, especially sodium phosphate, Tris, citrate, and
histidine buffers.
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38. Bulking Agents
The purpose of the bulking agent is to provide bulk to the
formulation.
This is important in cases in which very low concentrations
of the active ingredient are used.
Crystalline bulking agents produce an elegant cake structure
with good mechanical properties.
However, these materials often are ineffective in stabilizing
products such as emulsions, proteins, and liposomes but
may be suitable for small-chemical drugs and some
peptides e.g. Mannitol, Sucrose or one of the other
disaccharides.
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39. Stabilizers
 In addition to being bulking agents, disaccharides form an
amorphous sugar glass and have proven to be most effective in
stabilizing products such as liposomes and proteins during
lyophilization. Sucrose and trehalose are inert and have been used in
stabilizing liposome, protein, and virus formulations. Glucose,
lactose, and maltose are reducing sugars and can reduce proteins by
means of the Mailard reaction. Two hypotheses have been
postulated to explain the stabilizing effects of the disaccharides.
 The water replacement hypothesis: Disaccharides have been found
to interact with these products by hydrogen bonding similarly to the
replaced water.
 The vitrification hypothesis: Disaccharides form sugar glasses of
extremely high viscosity. The drug and water molecules are
immobilized in the viscous glass, leading to extremely high activation
energies required for any reactions to occur.
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40. Tonicity Adjusters
The need for such a formulation may be dictated by either the stability
requirements of the bulk solution or those for the route of
administration. Excipients such as mannitol, sucrose, glycine, glycerol,
and sodium chloride are good tonicity adjusters.
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41. Collapse temperature (Tc) modifiers
The 1˚ drying temperature for amorphous materials should
be below the Tc of the formulation, while some Excipients
have very low values of Tc which increases the 1˚ drying
time hence to reduce the 1˚ drying temperature Collapse
temperature modifiers are used which increases the Tc of
the formulation, making the process economical without
compromising the quality of product.
Examples of collapse temperature modifiers includes
dextran which have a collapse temperature of -9°C,
hydroxyethyl starch having a collapse temperature of -5°C.
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42. Effects of the ingredients on the
lyophilization process
 The process of lyophilization will be determined by the formulation For
example, the use of disaccharides will result in a low collapse
temperature, which causes primary drying to be performed at low
temperatures and implying a long process.
 A large volume fill or high solids content in the formulation will provide
increased resistance to mass transfer, hence a longer process.
 The process also can determine the properties of the formulation.
 The freezing process can influence crystallization of excipients such as
mannitol and glycine.
 Incomplete crystallization will depress the collapse temperature
Significant crystallization of the bulking agent will reduce drying time.
 However, large amounts of crystalline bulking agent can reduce
stabilizing effects of the amorphous stabilizer especially with proteins.
 Dextran is often used as excipient during lyophilization as a bulking
agent and/or a collapse temperature modifier.
42

43. CYCLE OPTIMIZATION
 In addition to designing a recipe that successfully dries a product, it is
also extremely valuable to optimize (shorten) the length of the cycle,
especially if there is potential for process repetition or scale-up for
production. Freeze drying can be a multi-day process. The cycle time can
often be substantially reduced by investigating several factors:
 Freezing and annealing – maximize crystal size and crystallization to increase
drying rates.
 Thickness of product - water vapor molecules experience resistance as they
exit from the dried portion of the product. Thinner samples yield less
resistance to vapor flow and lead to faster drying. Shell freezing can help
when drying bulk product in flasks.
 Critical Collapse Temperature – this is the most important piece of
information for cycle optimization. The ability to run primary drying at higher
product temperatures greatly reduces drying time by creating a larger
pressure differential between the vapor pressure over ice in the product and
the pressure at the condenser. Each 1oC increase in product temperature can
decrease primary drying time by 13%.
43

44. Freeze dryer classification
Freeze dryers can be informally classified by the type of product
chamber: (1) Manifold dryers where the product is typically pre-frozen
& in flasks (2) Shelf dryers where the product is placed in a tray or
directly on a shelf (3) Combination units with both drying options.
44

45. LYOPHILIZATION EQUIPMENT:-
There are essentially three categories of freeze dryers:
Manifold freeze-dryer
Rotary freeze dryer
Tray style freeze-dryer
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46. Freeze dryer typesFreeze-dryers can also be grouped by size & use: (1) laboratory
bench-top units for R&D (2) pilot units for process development
and scale-up, and (3) larger production-sized units. It should be
noted that in addition to process scale-up work, pilot-sized freeze
dryers are often used for product R&D as well as small volume
production applications.
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47. Manifold freeze dryers
 Manifold freeze dryers rely on ambient conditions to provide the
heat of sublimation to the product. This heat input does not melt
the product because an equivalent amount of heat is removed by
vaporization of the solvent. Advanced shelf freeze dryers can provide
a heat source to control/expedite the drying process and they can
also employ the refrigeration system to allow freezing of product
inside the unit.
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48. How to choose freeze dryer??
Choosing a freeze dryer depends on the product
characteristics as well as many other application-based
variables including the container that the product will be
dried in, the shelf area or number of ports required to
accommodate the quantity to be dried in each batch, the
total volume of ice to be condensed and whether there are
any organic solvents.
The type and shape of product being dried and its end-use
also need to be considered.
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49. APPLICATIONS OF LYOPHILIZATION
TECHNOLOGY
Industrial applications:
1] Pharmaceutical industry:
2] Food industry:
3] Other industries:
49

50.  Lyophilization (freeze-drying) is often used to prepare dry
pharmaceutical formulations to achieve commercially viable shelf lives.
 The process comprises three steps: freezing, primary drying, and
secondary drying.
 Under appropriate lyophilization conditions, the ice is removed by
sublimation during primary drying.
 Residual water in the freeze concentrate is removed in the secondary
drying step.
The knowledge of how to control, or at least manipulate, the freezing
step will help to develop more efficient lyophilization cycles and
biopharmaceutical products with an improved stability.
CONCLUSION
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