Lyophilization

J . J eff Schwegman, Ph.D.

AB BioTechnologies
Bloomington, Indiana
jjschwegman@gmail.com
Case Studies of Failed Lyophilized
Products
Failed Products
When a failure occurs for a lyophilized
product, the formulation/process
development scientist plays a major role
in identifying the reason(s) for the failure,
and in correcting the problems that
caused the failure.
Failed Products
1. Differentiate between equipment and
formulation failures
Chart from lyophilization cycle
Visual observation of samples
Location of failure
Thermal Analytical data (Tg, Te)
Stages of Freeze-Drying
Freezing
Primary
Drying
Secondary
Drying
Failed Products
2. Correct identification of product
failure
Collapse
Meltback
Partial collapse
Phase separation
Analytical data (potency/purity/moisture)
Active/Excipients/Container
Collapse
Complete Collapse of
the Product
Melt Back
Melt Back
Failed Products
3. Correcting the problems
Longer/shorter processing times
Warmer/colder shelf temperatures
Lower/higher vacuum levels
Less throughput to condenser
Reformulation
Definitions
A eutectic mixture is an intimate mixture
of 2 (or more) crystalline species that are
in such close contact that they melt like a
single, pure substance.
A glass is an amorphous species
(randomly oriented molecules) which
forms a solid, non flowing mass below its
Tg
Freezing of Aqueous Systems
Solution
Solute crystallizes?
Ice Nucleation
Ice crystal growth, freeze
concentration
Solute Crystallizes,
eutectic formation
Metastable
Glass
Stable Glass
Lyotropic
Liquid Crystal
Thermal Transition Terms Important
in Freeze-Drying
Eutectic temperature (Te)
Refers to crystalline systems and measured by thermal
or thermoelectric analyses
Glass transition temperature (Tg and Tg)
Refers to amorphous systems and also measured by
thermal or thermoelectric analyses
Collapse temperature (Tc)
Essentially the same as Tg, but measure by freeze-dry
microscopy.
Eutectics and Glasses
Why are These Important?
The eutectic or glass transition
temperatures determine maximum
temperature that the product can
withstand during primary drying
without loss of structure
Formulation 1
Melt Back (90% of Batch)
Formulation 1
API: Small molecule,
crystalline 10mg/mL
Excipients:
10 mM phosphate
buffer
pH 6.8
10 mg/mL mannitol
Formulation and Analytical Data
Te: -5C
Potency/Purity Post Lyo:
10.1 mg-mL/99.9%
Moisture: 5%
Fill Volume: 20 mL
Vial Size: 50 mL
Formulation 1
Dryer Parameter Set Points
Freezing:
25C to 40C
Primary Drying:
Shelf Temp: -5C
Vacuum: 50mTorr
Condenser: -77C
Secondary Drying:
Shelf Temp: 65C
Vacuum: 50mTorr
Condenser: -77C
Formulation 1
-90
-80
-70
-60
-50
-40
-30
-20
-10
0
10
20
30
40
50
60
70
300 400 500 600 700 800 900 1000 1100 1200 1300 1400 1500
Time, minutes
T
e
m
p
e
r
a
t
u
r
e
,

C
0
100
200
300
400
500
600
700
800
900
1000
P
r
e
s
s
u
r
e
,

M
T
o
r
r
Shelf
Condenser
Tc1
Tc2
Tc3
Tc4
Tc5
Pirani
Formulation 1
Recommended Cycle/Formulation
Modifications
Increase primary drying time (ensure all vials complete
primary drying
Formulation 2
Complete Collapse of
All Vials in Batch
Formulation 2
API: Peptide,
Amorphous 1mg/mL
Excipients:
15 mM HEPES buffer
pH 7.5
15 mg/mL glycine
5 mg/mL trehalose
Formulation and Analytical Data
Tg: -45C
Potency/Purity Post Lyo:
0.73 mg-mL/72.8%
Moisture: 20%
Fill Volume: 4mL
Vial Size: 10 mL
Formulation 2
Dryer Parameter Set Points
Freezing:
5C to 45C
Primary Drying:
Shelf Temp: -40C
Vacuum: 250mTorr
Condenser: -77C
Secondary Drying:
Shelf Temp: 25C
Vacuum: 150mTorr
Condenser: -77C
Formulation 2
-90
-80
-70
-60
-50
-40
-30
-20
-10
0
10
20
30
40
0 200 400 600
Time, minutes
T
e
m
p
e
r
a
t
u
r
e
,

C
0
50
100
150
200
250
300
350
400
450
500
550
600
650
700
750
800
850
900
950
1000
V
a
c
u
u
m
,

m
T
o
r
r
Shelf, C
Thermocouple 1
Thermocouple 3
Thermocouple 4
Condenser, C
Pirani, mTorr
Formulation 2
Recommended Cycle/Formulation
Modifications
1. Product temperature exceeds Tg at the start of primary
drying look at lowering shelf temperature
2. Product Tg is likely too low to be able to freeze-dry
3. Need to look at the formulation and try Tg modifying
excipients
Formulation 3
95% of Vials in Batch
Broken
Formulation 3
API: Small Molecule,
Crystalline 3mg/mL
Excipients:
50 mM Citrate buffer
pH 5.0
50mg/mL mannitol
Formulation and Analytical Data
Te: -5C
Potency/Purity Post Lyo:
3.0 mg-mL/100%
Moisture: 0.8%
Vial Size/Fill Volume:
10mL/8mL Fill
Formulation 3
Dryer Parameter Set Points
Freezing:
5C to 45C
Primary Drying:
Shelf Temp: 0C
Vacuum: 200mTorr
Condenser: -80C
Secondary Drying:
Shelf Temp: 45C
Vacuum: 200mTorr
Condenser: -80C
Formulation 3
-90
-80
-70
-60
-50
-40
-30
-20
-10
0
10
20
30
40
50
60
0 200 400 600 800 1000 1200 1400 1600 1800
time (minutes)
t
e
m
p

(
C
)
0
100
200
300
400
500
600
700
800
900
1000
p
r
e
s
s
u
r
e

(
m
i
c
r
o
n
s
)
Shelf
TP01
TP02
Condenser
Pirani
Formulation 3
Recommended Cycle/Formulation
Modifications
1. Decrease the amount of mannitol in the formulation
2. Decrease the fill volume
Formulation 4
Complete Collapse of
Samples on Stability at
+40C
Formulation 4
API: Peptide,
Amorphous 200g/mL
Excipients:
40 mM Citrate buffer
pH 4.5
10 mg/mL mannitol
1 mg/mL sucrose
Formulation and Analytical Data
Tg: -32C
Potency/Purity Post Lyo:
202 g-mL/100%
Moisture: 3.5%
Fill Volume: 1mL
Vial Size: 3 mL
Formulation 4
Dryer Parameter Set Points
Freezing:
5C to 50C
Anneal at 10C
Cool to 45C
Primary Drying:
Shelf Temp: -45C
Vacuum: 50 mTorr
Condenser: -80C
Secondary Drying:
Shelf Temp: 25C
Vacuum: 50 mTorr
Condenser: -80C
Formulation 4
Recommended Cycle/Formulation
Modifications
1. Increase secondary drying time
2. Increase drying time of stoppers under vacuum to
remove moisture
Formulation 5
Specifics:
Ring of Meltback on
Bottom
Only batch out of 10
previous to have problem
5% of Batch Rejected
Mottled Bottom and Top
Spotty Discoloration
Formulation 5
API: Small Molecule, 173 g/mol
Solution Concentration: 450 mg/mL
Eutectic Melting Temperature: -12.3C
Vial Size: 20 mL x 20 mm
Lyophilization Cycle Ran Flawlessly
Significant Degradation Products Detected by
HPLC
HPLC Potency: 150%
Formulation 5
-90
-80
-70
-60
-50
-40
-30
-20
-10
0
10
20
30
40
50
60
70
0 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400
Time, minutes
T
e
m
p
e
r
a
t
u
r
e
,

C
0
100
200
300
400
500
600
700
800
900
1000
SHELF IN
Thermocouple 1
Thermocouple 2
Thermocouple 3
Thermocouple 4
Thermocouple 5
Condenser
Pirani, mTorr
Formulation 5
Recommended Cycle/Formulation
Modifications
1. Glitch in Filling Equipment Various Vials Getting
Double Fill.
2. Engineering Contacted and Corrected the Problem
Summary
Troubleshooting and correcting failed lyophilized
products can be a challenge as many different
factors (formulation and equipment) can be
involved in a failure. By taking a systematic and
empirical approach to troubleshooting, it is
possible for the development scientist to
understand why the failure occurred and suggest
changes (formulation or equipment) to prevent
further failures from occurring in future batches
AB BioTechnologies, LLC
J . J eff Schwegman, Ph.D., CEO
Providing Consulting, Teaching and Expertise in:
Parenteral Preformulation and Formulation Development
Lyophilization Cycle Design and Optimization
Troubleshooting Failed Cycles and Formulations
Solutions, Emulsions, Suspensions and Lyo Products
Formulation and Process Transfer and Scale-Up
Contact Dr. Schwegman at
812-327-6898 or jjschwegman@gmail.com