1. Jian-Hwa Han
USP Expert Panel Meeting
May 29-30, 2013
Using Enzymes for
Dissolution - The
Understandings from
Gelatin to Enzymes
SUB-TITLE, DATE
2. • Dissolution Equipment and Apparatus
• Gelatin Dissolution in Various Media
• Reference Compound – Acetaminophen
o Tablet vs. OE Capsule (Over-encapsulated Tablet sample)
• Tier-2 Testing Performance Evaluation
o Enzymes Information
o Control of Experiments
o Setting the Reference Point - <711> Pepsin
o Equivalent Performance of other Enzymes
o Reproducibility of Bromelain and Papain
o Disintegration Evaluation
• Summary & Next Step
Presentation Outline
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
2
3. Dissolution by FO-UV System
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
3
• System 1: Distek Opt-Diss FO-UV Dissolution System
• System 2: Sotax with Rainbow FO-UV System
• Advantage: Direct detection with details profiles
(and w/o Sampling errors)
• Apparatus 2 @ 50 rpm
• Samples
• Hard Gelatin Shell
• Tylenol Tablet
• OE Tylenol Sample
• Cross-linked OE Tylenol Sample
5. Gelatin Dissolution – Typical Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
5
• Data clearly
show much
faster
dissolution rate
in 0.1N HCl
• The dissolution
rates in pH 4.5,
pH 6.8, and
water are
similar
Dissolution of Hard Gelatin Shell - App 2 @ 50 rpm
in Typical Dissolution Media
0.0
20.0
40.0
60.0
80.0
100.0
120.0
0.0 10.0 20.0 30.0 40.0 50.0 60.0
Time, Minutes
%LC
0.1N HCl Acetate, pH 4.5 Phosphate, pH 6.8 Water
6. Gelatin Dissolution –
Surfactant Impact in Water
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
6
• Surfactants do
slow down the
dissolution of
gelatin
• The impact
from POE and
CTAB are not
significant
• SDS posts more
impact, but it
still within the
variability
Dissolution of Hard Gelatin Shell - App 2 @ 50 rpm
Surfactant Impact in Water
0.0
20.0
40.0
60.0
80.0
100.0
120.0
0.0 10.0 20.0 30.0 40.0 50.0 60.0
Time, Minutes
%LC
Water 0.5% SDS in Water 0.5% POE in Water 0.2% CTAB in Water
7. Gelatin Dissolution –
Surfactant Impact in pH 6.8 Phosphate Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
7
• No significant
impact
observed in
pH 6.8
Phosphate
media with
either SDS or
POE10LE
Dissolution of Hard Gelatin Shell - App 2 @ 50 rpm
Surfactant Impact in pH 6.8 Phophaste Buffer
0.0
20.0
40.0
60.0
80.0
100.0
120.0
0.0 10.0 20.0 30.0 40.0 50.0 60.0
Time, Minutes
%LC
0.5% SDS in pH 6.8 pH 6.8 Buffer 0.5% POE in pH 6.8
8. Gelatin Dissolution –
Surfactant Impact in 0.1N HCl Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
8
• No significant
impact
observed for
POE in
0.1N HCl media
• Significant
impact
observed for
SDS in 0.1N HCl
media!!!
Dissolution of Hard Gelatin Shell - App 2 @ 50 rpm
Surfactant Impact in 0.1N HCl
0.0
20.0
40.0
60.0
80.0
100.0
120.0
0.0 10.0 20.0 30.0 40.0 50.0 60.0
Time, Minutes
%LC
0.5% SDS in HCl 0.1N HCl 0.5% POE in HCl
9. Comparison of Gelatin Dissolution Time in
Various Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
9
Note – In general, the capsule opens within 2 minutes in 0.1N HCl.
Gelatin Dissolution Comparison
0
5
10
15
20
25
30
0.1
N
HC
l,900
m
L
0.5%
SDS/0.1N
H
C
l
0.5%
PO
E/0.1N
H
Cl
Phosphate,pH
6.8
0.5%
SDS/pH
6.8
0.5%
PO
E/pH
6.8
W
ater,900
m
L
0.5%
SDS/W
ater
0.5%
PO
E/W
ater
0.2%
CTAB/W
ater
Time,Minutes
>~25% Dissolved
>~50% Dissolved
11. Reference Compound – Acetaminophen
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
11
• Tylenol 250 mg
Tablet was used as
the test drug
• Size 0 HGC Used for
the OE sample
without filler
12. Tablet vs. OE Capsule Dissolution – Typical Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
12
Gelatin Dissolution Research - Acetaminophen, 0.1N HCl
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
HGC Shell Tylenol OE Capsule Tylenol Tablet
Gelatin Dissolution Research - Acetaminophen, Water
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
HGC Shell Tylenol OE Capsule Tylenol Tablet
Gelatin Dissolution Research - Acetaminophen, pH 6.8
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
HGC Shell Tylenol OE Capsule Tylenol Tablet
Gelatin Dissolution Research - Acetaminophen, pH 4.5
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
HGC Shell Tylenol OE Capsule Tylenol Tablet
Full release (>85%) within
3 minutes from Tablet
(for all conditions)
Once capsule opens, the
residual gelatin does not
impact dissolution for this
product.
Full release (>85%) within
10 minutes from OE
Capsule (for all conditions)
0.1N HCl pH 4.5
pH 6.8 Water
13. Acetaminophen OE Capsule Dissolution –
Surfactant Impact (in Water)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
13
• Surfactant
somewhat
delays the drug
release
• SDS: high
capsule-to-
capsule
variability
• However, all
conditions
release >85% in
10 minutes
Gelatin Dissolution Research - Acetaminophen OE Capsule,
Surfactant Impact (in Water)
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
Water Only 0.5% Tween-80 0.5% POE 0.5% CTAB 0.5% SDS
14. Acetaminophen OE Capsule Dissolution –
Surfactant Impact (in 0.1N HCl)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
14
• Surfactants do
NOT slow down
the drug release
in 0.1N HCL
• CTAB and POE
show slightly
faster drug
release rate
(Very low gelatin
dissolution rate in
0.5% SDS/HCl does
not impact the
drug release from
OE Capsules – see
slide #16)
Gelatin Dissolution Research - Acetaminophen OE Capsule,
Surfactant Impact (in 0.1N HCl)
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
0.1N HCl only 0.5% SDS 0.5% CTAB 0.5% POE Gelatin_SDS/HCl
15. Acetaminophen OE Capsule Dissolution –
Surfactant Impact (in pH 6.8 Phosphate)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
15
• CTAB delays the
capsule opening
(> 5minutes)
• SDS shows very
high capsule-to-
capsule
variability, and
did not release
>85% at 10
minutes
Gelatin Dissolution Research - Acetaminophen OE Capsule,
Surfactant Impact (in pH 6.8 Phosphate)
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
pH 6.8 Phosphate only 0.5% SDS 0.5% CTAB 0.5% POE
16. Acetaminophen OE Capsule Dissolution –
SDS (at 0.5%) Impact in Different Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
16
• More variable
in water and pH
6.8 Phosphate
• SDS/pH 6.8: did
not release
>85% at 10
minutes
• However, all
conditions can
release >85% of
drug in 15
minutes
Gelatin Dissolution Research - Acetaminophen OE Capsule,
SDS Impact in Various Media
0
20
40
60
80
100
120
0 5 10 15 20 25 30
Time, Min.
%LC
Water Only SDS in Water SDS in HCl SDS in pH 6.8
17. Comparison of Acetaminophen Dissolution Time:
Tablet vs. OE Capsule in Various Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
17
• The Tablet full releases the drug within 3 minutes (> 90% for all media tested)
• The OE Capsule releases >85% drug within 10 minutes for all conditions tested
Acetaminophen Dissolution Comparison - Tablet vs. OE Capsule (Tier-1)
0
1
2
3
4
5
6
7
8
9
10
0.1N
H
Cl, Tablet
0.1N
H
Cl, O
E
C
apsule
0.5%
SDS/0.1N
H
C
l_C
ap
pH
4.5, Tablet
pH
4.5, O
E
Capsule
pH
6.8, Tablet
pH
6.8, O
E
Capsule
0.5%
CTAB/pH
6.8_C
ap
W
ater, Tablet
0.5%
Tween-80, Tablet
W
ater, O
E
C
apsule
0.5%
Tween-80, O
E
C
ap
0.5%
SDS, O
E
C
apsule
0.5%
PO
E, O
E
C
apsule
0.5%
CTAB, O
E
C
apsule
Time,Minutes
>=5% Release >=85% Released
19. Enzymes Information (Currently in USP <711>)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
19
Pepsin:
• Manufacture: American
Laboratories Inc.
• Product: Pepsin, 1:10,000
Powder
• Lot No: 00511010
• Pepsin Activity: 1:10,400
Pacreatin:
• Manufacture: American
Laboratories Inc.
• Product: Pancreatin 8X USP
Powder
• Lot No: 52320708
• Activity:
Protease: 8.1X USP (203 USP units/mg)
Amylase: 10.2X USP
(255 USP units/mg)
Lipase: 33.7 USP Units/mg
20. Enzymes Information (New)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
20
Bromelain:
• Manufacture: American
Laboratories Inc.
• Product: Bromelain 2400 GDU
• Lot No: 55920701-A
• Proteolytic Activity:
2,482 GDU/g
Papain:
• Manufacture: American
Laboratories Inc.
• Product: Papain Full Strength
• Lot No: 70310665-B
• Proteolytic Activity:
66,900 USP units/mg
21. Control of Experiments
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
21
Need to get Reproducible results! (Consistency!)
• Same Equipment and Apparatus
• Same Analyst
• Consistent Execution of Dissolution Runs
• Direct Detection – Avoid sampling errors
• Precise operation for generating cross-linked capsule
samples: 2-hour Treatment with Formaldehyde
22. <711> Tier-1 vs. Tier-2:
Non-cross-linked Capsules
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
22
• No significant
difference after 10
minutes
Normal Capsule - 0.1N HCl
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
0.1N HCl/Pepsin 0.1N HCl
Normal Capsule - pH 6.8
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
pH 6.8 pH 6.8/Pancreatin
Q: NLT 80% in 30 min.
23. Tier-1, 0.1N HCl:
Non-cross-linked Capsule vs. Cross-linked Capsule
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
23
• Fails Stage-3!
• The ~same
level of cross-
linked capsule
samples (i.e. 2-
hour treated)
were used for
all Tier-2
experiments
Tier-1, 0.1N HCl: Normal Capsule vs. X-linked Capsule
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_Tier-1_HCl Xlinked_Tier-1_HCl
Q: NLT 80% in 30 min.
24. Tier-1, pH 6.8:
Non-cross-linked Capsule vs. Cross-linked Capsule
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
24
• Fails Stage-3!Tier-1, pH 6.8: Normal Capsule vs. X-linked Capsule
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_Tier-1_pH 6.8 Xlinked_Tier-1_pH 6.8
Q: NLT 80% in 30 min.
25. <711> Pepsin, 750,000 Units/L
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
25
• Current <711>
750,000 units/L
is equivalent to
16,000,000 FCC
units/L
• For X-linked
capsule (2-hour
treated with
formaldehyde),
Tier-2 result at
30 minutes is
92% (± 6.2%)
May Pass S-1
of Tier-2
<711> Pepsin @ 750,000 Units/L
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_Tier-1_HCl Xlinked_Tier-2_HCl
26. <711> Pancreatin, 1750 Units/L
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
26
• Current <711>
1750 units/L
• For X-linked
capsule (2-hour
treated with
formaldehyde),
Tier-2 result at
30 minutes is
83% (± 10.6%)
Will go to
Stage-2 of
Tier-2
<711> Pacreatin @ 1750 Units/L
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_Tier-1_pH 6.8 Xlinked_Tier-2_pH 6.8
27. Finding Equivalent Pancreatin Level to Match
Pepsin Performance
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
27
• Current <711>
1750 units/L
(1X)
• Levels tested:
[Units/L]
1750 (1X), 2000,
2500, 3500 (2X),
and 5250 (3X)
• For 2000 Units/L,
Tier-2 result at
30 minutes is
93% (± 4.0%)
May Pass S-1
of Tier-2
pH 6.8 / Pancreatin
0
20
40
60
80
100
120
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_Tier-2 Normal_Tier-1 Xlinked_Tier-1
Xlinked_Tier-2_3X Xlinked_Tier-2_2X Xlinked_Tier-2_1X
Xlinked_Tier-2_2500U Xlinked_Tier-2_2000U
1750 Units/L
2000 Units/L
5250 Units/L (3X)
32. Screening Papain at pH 4.5 to Match Pepsin
Performance
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
32
• Starting point:
9 mg/L (i.e.
~600,000 U/L)
• Levels tested:
[KU/L]
600, 550, 525,
and 500 KU/L
• Very small
amount of
Papain Used:
pH 4.5 / Papain
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_T1 X_T1 X_Tier-2 (600KU)
X_Tier-2 (550KU-1) X_Tier-2 (550KU-2) X_Tier-2 (525KU-1)
X_Tier-2 (525KU-2) X_Tier-2 (500KU)
Units/L mg / 6L
600,000 53.8
550,000 49.3
525,000 47.1
500,000 44.9
33. Screening Papain at pH 4.5 to Match Pepsin
Performance (cont.)
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
33
• The results seem
more variable
than Bromelain
• For 550 KU/L,
Tier-2 result at
30 minutes is
95% (± 3.0%)
Too Good?
• For 525 KU/L,
Tier-2 result at
30 minutes is
85% (± 9.9%)
Will go to
Stage-2 of Tier-2
pH 4.5 / Papain
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Normal_T1 X_T1 X_Tier-2 (600KU)
X_Tier-2 (550KU, N=12) X_Tier-2 (525KU, N=12)
525 KU/L
35. Stability of Papain Medium at 550,000 Units/L
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
35
• X-linked Capsule
/ Tier-2 Testing:
Levels =
550 KU/L
• After 7 hours,
Tier-2 result at
30 minutes is
78% (± 15.3%)
May Fail the
Tier-2
pH 4.5 / Papain, ~550 KU/L
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Papain 550 kU-1 Papain 550 kU-2 Papain 550 kU_7 Hours
36. Stability of Papain Medium at 525,000 Units/L
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
36
• X-linked Capsule
/ Tier-2 Testing:
Levels =
525 KU/L
• Up to 4 hours,
the dissolution
rate did not slow
down
• Overall, it shows
high variability
from run-to-run
using Papain at
550 KU/L and
525 KU/L
pH 4.5 / Papain, ~525 KU/L
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Papain 525 kU-1 Papain 525 kU-2 Papain 525 kU_4 Hours
37. Stability of Bromelain Medium at 30 GDU/L
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
37
• X-linked Capsule
/ Tier-2 Testing:
Levels =
30 GDU/L
• The dissolution
rate are very
consistent up to
4 hours
• After 7 hours,
Tier-2 result at
30 minutes is
70% (± 9.6%)
May Fail the
Tier-2
pH 4.5 / Bromelain, ~30 GDU/L
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Bromelain 30GDU-1 Bromelain 30GDU-2
Bromelain 30GDU_4-hour Bromelain 30GDU_7-hour
38. Solution Stability of Enzyme Media
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
38
• Enzyme activity will drop over time in solution
• From the observations of Bromelain and Papain media, it
suggests that the dissolution run should start within 4 hours
after the media is made. The media is no longer good after 7
hours.
• Pepsin and Pancreatin had not been tested for solution stability,
but the general rule from above may apply to them
39. Disintegration Evaluation:
Another Look from Different Hydrodynamics
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
39
• Bromelain @30 GDU/L
and Papain @525
KU/L have much
longer disintegration
time than Pepsin
• @6X works well (6
capsules in 1 beaker)
Disintegration of Cross-linked Capsules
(Individual Data)
0
5
10
15
20
25
30
W
ater
SG
F
SIF
Pepsin,1XPancreatin,1XBrom
elain,1X
Papain,1XBrom
elain,6X
Papain,6X
DisintegrationTime,Min.
0
2
4
6
8
10
12
14
16
18
20
W
ater
SG
F
SIF
Pepsin, 1X
Pancreatin, 1X
Brom
elain, 1X
Papain, 1X
Brom
elain, 6X
Papain, 6X
Disintegration of Cross-linked Capsules (Mean, N=6)
• Water can not open the X-linked
capsules within 15 minutes
• Pancreatin @1750 units/L is
weaker than SGF and Pepsin
(similar to dissolution results)
40. Summary: Recommended Levels of these four
Enzymes
Recommended
levels of
Enzymes:
•Pepsin: 750,000
Units/L (<711>),
i.e. 16,000,000
FCC units/L
•Pancreatin:
2000 Units/L
•Bromelain:
30 GDU/L
•Papain: 550,000
Units/L
(525,000 U/L may
be slightly too
weak and too
variable.)
40
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
Tier-2 Performance Comparison - All 4 Enzymes (N=12)
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Pepsin/750000 U Pancreatin/2000 U
Bromelain/30 GDU Papain/550000 U
Normal Capsule (pH 6.8)
Tier-2 Performance Comparison - All 4 Enzymes (N=12)
0
20
40
60
80
100
0 10 20 30 40 50 60
Time, Minutes
%LC
Pepsin/750000 U Pancreatin/2000 U Bromelain/30 GDU Papain/525000 U
41. Summary and Next Step
• The gelatin dissolves faster in 0.1N HCl than pH 4.5 & pH 6.8
• SDS may interact with gelatin and show more variable results. The
impact is not significant for fast dissolving formulations
• Pepsin in 0.1N HCl is used as the reference, and identified the
appropriate levels for the other three enzymes
Next Steps:
• Verify these experiment results from other labs
• Verify working pH ranges for each enzymes
• Find the best way to express the enzyme activity, and be consistent
from various suppliers
41
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
42. • The design, study conduct, and financial support for the study
were provided by Abbvie. Abbvie participated in the
interpretation of data, review, and approval of the
presentation.
• Abbvie is the sole source of funding for the related studies in
this presentation. No other entity has provided any funding
for the related studies in this presentation.
Disclosures
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
42
43. Acknowledgement
• Dr. Jian-Hwa Han is employed at Abbvie. Dr. Han is the primary
investigator for this study.
• Mr. Daniel Bonilla is employed at Abbvie. Mr. Bonilla provided
technical assistance for this study.
• Ms. Susan George and Dr. Paul D. Curry, Jr. are employed at Abbvie
who have contributed for scientific discussions and reviewing the
presentation materials.
• Mr. Thomas Langdon of American Laboratories, Inc. provided all
four enzymes for this study, and the technical information about
these materials.
43
USP Dissolution Workshop, Rio de Janeiro-Brazil, 2013
