paolo avalle discovery chemistry congress2012

Application of imaging techniques to oral dosage forms.

Examples of in-situ imaging.Paolo Avalle

Merck Sharp & Dohme (UK’CH)

Introduction• Direction of modern formulation efforts:

– improve the solubility of the drugs with effective formulation– tune the drug release profile to meet the desired Pharmacokinetic criteria.

• Focus of the talk: use of imaging techniques as a characterization tool of drug-polymers system

• Remit of imaging characterization techniques– Provide surface and internal chemical imaging of the whole dosage form or of individual components

on a macro-, micro- or nanoscale.

– Temporal and spatial mapping of the drug release from the tablet matrix and obtaining novel mechanistic insights into the drug liberation phenomena.

– Understanding the interplay between the underlying diffusion and erosion mechanism of release and how these can be related to the solubility of the drug.

Agenda

•Mechanism of dissolution explored by imaging techniques

– NIR microscopy– MRI

•Conclusions & Acknowledgements

NIR microscopy• Applications & Case studies:

1. Distribution of API and excipients.– Formulation development: CR pellets– Formulation troubleshooting CR pellets

2. In-situ NIR: Imaging the dissolution mechanism.– Diffusion – based systems (“high” solubility)– Erosion –based systems (low solubility)– Failure mode of erosion based matrices

3. In-situ MRI: gel layer formation and drug mobilization

NIR spectroscopy• High content imaging I achieved by rasterized acquisition of spectra. • A single spectrum is acquired at each location (pixel) using a moveable stage.

• From the collection of location tagged spectra a map can be generated in various ways:

- The integrated absorbance of a specific peak- Intensity of a specific peak- PCA- PLS

pixel: 25 x 25 m

1

2

3

4

1

2

3

4

pixel: 25 x 25 m

1

2

3

4

1

2

3

4

GMS-900

PVP

Optical Image

PVP

Avicel Ethocel TEC

API Lactose

Reconstructed image with background suppression

Artifact.No coating present

in this part of the pelletcfr. Optical Image

GMS-900

PVP

Optical Image

PVP

Avicel Ethocel TEC

API Lactose

GMS-900

PVP

Optical Image

PVP

Avicel Ethocel TEC

API Lactose

Reconstructed image with background suppression

Artifact.No coating present

in this part of the pelletcfr. Optical Image

presented at UKICRS 2010

Distribution of API and excipients.

1. Distribution of API and excipients

1: Horizontal sample support made in-house. 2: Adhesive disc. 3: Microscope cover slip. 4: Glue. 5: Sample pellets. 6: Rotating tungsten carbide blade. 7: Vertical Axis of cutting: this ensures a flat surface 8: Horizontal movement of the blade.

• The distribution of different chemicals is represented in colour-coded intensity maps. • The maps of two chemicals can be compared in a pixel-to-pixel fashion to generate a correlation chart • Avicel and API shows a positive correlation indicating potential co-location. • The association of APIand Lactose is somewhat less evident.• API and PVP appear to be anti-correlated.



Distribution of API and excipients: Co-localization

•Maps obtained by Least square fitting of the NIR spectra of the 7 pure components. •Maps were masked to remove the background spectra from the Least square fitting of the Map. •Background of the image is showed in blue on the left side and in white on the Reconstructed image.•This maps isolated only 6 out of 7 components. TEC could not be detected

Note 1) Negative-correlationbetween the spatial distributionof LACTOSE and API

2) Positive-correlationbetween the spatial distribution of AVICEL and API

3) Negative-correlationbut more noisy betweenGMS-900 and MK-1



Distribution of API and excipients: Co-localization

Scatter plot• The intensity of Each pixel of the API map is used as X-coordinate.• The intensity of Each Pixel of the AVICEL map is used as Y-coordinate.• Two identical maps (for example API vs. API) would generate a straight line with positive correlation. • The colour code of the points indicate their position on the map.• The position is indicated approximately by the colour key in the border of the graph. • White points are located at the centre of the original image.

Avicel

API

Two positively correlated clustersa) White and Green, located in

the centre of the picture indicates a degree of matching in the coating pixels intensities between the two maps.

b) Yellow-red, located on the right side of the map indicate degree of matching intensities between the AVICEL and MK-0941 maps

a

b

Controlled Release pellets MK-1.

Spatial Co-location


Date: 18/04/2006

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.0256

-0.020

-0.015

-0.010

-0.005

0.000

0.005

0.010

0.015

0.0196

cm-1

A

Date: 18/04/2006

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.0139

-0.012

-0.010

-0.008

-0.006

-0.004

-0.002

0.000

0.002

0.004

0.006

0.008

0.010

0.012

0.014

0.016

0.018

0.020

0.022

0.024

0.026

0.0278

cm-1

A

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.0108

-0.008

-0.006

-0.004

-0.002

0.000

0.002

0.004

0.006

0.008

0.010

0.012

0.014

0.016

0.018

0.0199

cm-1

A

Date: 18/04/2006

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.0256

-0.020

-0.015

-0.010

-0.005

0.000

0.005

0.010

0.015

0.0196

cm-1

A

Date: 18/04/2006

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.180

-0.16

-0.14

-0.12

-0.10

-0.08

-0.06

-0.04

-0.02

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.140

cm-1

A

Date: 18/04/2006

6100.0 5800 5600 5400 5200 5000 4800 4600 4400 4200 4000 3800.0-0.0157

-0.014

-0.012

-0.010

-0.008

-0.006

-0.004

-0.002

0.000

0.002

0.004

0.006

0.008

0.010

0.012

0.014

0.0156

cm-1

A


Formulation troubleshooting CR pellets: PLS data representation

FITACTUAL

FITACTUAL

FITACTUAL

FITACTUAL

FITACTUAL

FITACTUAL


Scheme for tablet hydration.- swelling. - drug migration vs. dissolution - polymer dissolution within a controlled release formulation.

The particle labelled ‘A’ indicates the drug. While it is commonly accepted that swelling is subsequent to permeation and hydration of the tablet the extent of those event and the extent of drug migration vs. dissolution is largely dependent on the solubility of the drug and the viscosity of the polymer. Together, these parameters modulate the release profile.

2. In-situ NIR: Imaging the dissolution mechanism

Basic Concepts

TABLETS: 400 mg, 8 mm, flat-faced tablets containing 125 mg dose of a low solubility drug (MK-1), IN FLOW ANALYSIS: A bespoke tablet hydration cell enabled the acquisition of NIR data during the dissolution process.

RESULTSEXPERIMENT SET-UP

• Image Size: 3000 x 1000 µm• Pixel Size: 25 µm• Scan Time: ~ 18 min• Scan Frequency: Every 30 min

•Medium: Deionised Water•Temperature: 37°C•Flow Rate: 10 mL/min


The subsequent acquisition of spectral map and their processing Allow to follow the evolution of the API signal and the HPMC signals.

0’ 10’ 20’ 30’ 40’ 50’ 60’ 180’120’90’dry

HPMC

API


European Journal of Pharmaceutical Sciences 43(5) 400-408

Diffusion based systems:: comparing two diffusion-based formulation

Fast

Slow

Fast

Slow

Fast

Slow

The hydration profiles exhibited several trends: 1. An apparent high intensity plateau, corresponding to a uniform distribution of HPMC (dry tablet core)

2. A sloped region indicative of a decreasing drug/HPMC concentration across the pseudo-gel layer 3. A plateau of low intensity arising from the bulk of the hydration medium.

Drug and HPMC profiles from the fast and slow formulations As the tablet was exposed to the hydration media, polymer relaxation occurred and the HPMC began to swell and hence the increment in the intensity profile became progressively sloped as a consequence.

2. In-situ NIR: Imaging the dissolution mechanismDiffusion based systems: comparing two diffusion-based formulation


The erosion, swelling and API dissolution front for both formulation SLOW AND FAST with data modellingThe data were modelled using the equation first proposed by Peppas and Sahlin to describe solute release from polymeric devices, where FP indicates the Front Position (either (i) the erosion front, (ii) the swelling front or (iii) the API front). FP is expressed in microns.

2. In-situ NIR: Imaging the dissolution mechanismDiffusion based systems:: comparing two diffusion-based formulation


mm tktkFP 221

mm tktkFP 221

:THIN GEL LAYER

: THICK GEL LAYER

• Image Size: 6000 x 2000 µm• Pixel Size: 25 µm• Scan Time: ~ 18 min• Scan Frequency: Every 30 min

•Medium: Deionised Water•Temperature: 37°C•Flow Rate: 10 mL/min

Component %

API (MK-1) 31.25

HPMC K4M 20.00

Avicel PH102 47.75

Magnesium Stearate 1.00

In-situ NIR: Imaging the dissolution mechanism

In-situ and in-flow imaging experiments

Erosion –based systems: Low solubility formulationIN-FLOW Imaging

STEP by STEP processing roadmap

Black = experimental spectrum (PLS target) Blue = PLS Fit

Load *.fsm in Transmittance

Load *.fsm in Transmittance

Reload

*.fsm file

Load masked .fsm

in Absorbance

Process /

DerivativeProcess / PCA

“20 factors”

Review Targets

Reload

masked .fsm file

Process / LSQ Fit

Load Target

SpectraProcess / Subtract Average 7800-3800

Process / Range

7800-3800

Process / Derivative

Load Target

SpectraProcess / Subtract Average 7800-3800

Process / Range

7800-3800

Review LSQ Fit

Masking

PLS Fitting

API HPMC AVICEL

Since the fitting obtained from the PLS is very good the representation of API, HPMC, and AVICEL maps Is to be considered valid and accurate.

Process / PCA “10 factors”

Process / Mask

Process / PCA “20 factors” and spatial masking “ALL”


Presented at UKPharmSCI 2011, paper in progress

API Maps from PLS30’ 60’ 90’ 120’ 150’ 180’ 210’ 240’

#1

#2

#3

30’ 60’ 90’ 120’ 150’ 180’ 210’ 240’HPMC Maps from PLS


• Undissolved API is present up to 3 hours and seems to migrate in the gel Layer• Can we zoom in and follow closely the fate of API particles?



API depletion and hydration profiles from PLS API mapsselective enhancement

8 bit Blue Channelconversion R3 240’

8 bitGrayscaleconversion R3 240’

R1

R2

R3

R1

R2

R3

API depletionHydration



Mechanism of release: Swelling and Erosion fronts.

Physical tablet boundary

PLS API mapR2 30’

PLS API mapR2 240’

Is it possible to further explore the NIR maps and gain a better understanding at a more microscopic level of the mechanism of release?



R1 R2 R3

ZOOM-IN on EROSION AND DISSOLUTION FRONTS

Mechanism of release: Swelling and Erosion fronts. Source of error bars


R2 30’

EROSION FRONT

SWELLINGFRONT


API / HPMC

Spatial Co-location of API and HPMC• API and HPMC are clearly co-located •Their distribution is not mutually exclusive within the section sampled by NIR• co-location can be applied to the PLS images to create a superposition map of API and HPMC.•Thresholding can be judiciously chosen to optimize the API particles separation contrast

PLS IMAGESAPI

HPMC

http://rsbweb.nih.gov/ij/plugins/colocalization.html

Contrast enhancement of PLS API/HPMC intensity maps


• This approach enables the editing of a sequence of images • The gel layer region (dark green) show low API concentration, and progressive

dissolution.• However larger aggregates of API (yellow) remains unchanged even when fully

“immersed” in the gel Layer (240’).• This colour band discrimination makes the images amenable to further analysis.

30’ 60’ 90’ 120’ 150’ 180’ 210’ 240’

GELerosion

swelling

Contrast enhancement of PLS API/HPMC intensity maps


1 1

NIR post PLS

NIR post PLS

API-HPMCcorrelation

API-HPMCcorrelation

2 2 2 2 2 2

API distribution time course

• From the correlation maps it is possible to further filter out the signal of the pure APIgenerating a highly contrasted image that enable single particle tracking.

• The comparison between the post-PLS (A), post Colocalization (B) and thresholding (C) shows that the signal of API distribution is retained throughout the processing.

• The highly contrasted images enable the study of the evolution of single API particle (or clusters of)

A AB BC C

1

30’ 60’ 90’ 120’ 150’ 180’ 210’

2

240’Two different particles are tracked (1) and (2)

2

Contrast enhancement of PLS API/HPMC intensity maps: Single particle tracking


30’ 60’ 90’ 120’

150’ 180’ 210’ 240’

Single particle tracking : Particle 1


30’ 60’ 90’ 120’

150’ 180’ 210’ 240’

1. Aggregation

2. Migration 3. Disintegration

Blue contour: indicated the particle frame at 120’ minutes chosen as reference. The aggregation process occurring in the first 90 minutes, seems to have stopped and the migration process of the whole particle will dominate for the subsequent 90 minutes before significant erosion will takes place.



30’ 60’ 90’ 120’ 150’ 180’ 210’ 240’

Particle -1

dissolved

Particle -1 show a a dissolution behaviour, fragmentation is also visible.Blue line: initial particle outline. Red area: actual particle at each time point.

water



Ref. frame

Because of the complex aggregation and migration the perimeter is not a good descriptor of the time course as the area

aggregation migration disintegration

Reference frame

Perimeter

Cluster -2

2

240’30’

1. Aggregation 2. Migration3. Disintegration

water



•In-situ MRI- Acquisition of MRI data during the dissolution, - Hyphenation of USP-IV dissolution with MRI.

Hyphenation of USP-IV dissolution with MRI.

NMR spectrometerPeristaltic

pump SGF

USP-IVFlow-through cell

x

yz

Imaging plane

FaSSIF

22.6 mm

Lescol® XL tablet

In-situ MRI

0.0

0.2

0.4

0.6

0.8

1.0

0 5 10 15 20

Standard USP-IV experiment

In line UV measurement

UV

abs

orpt

ion

(a.u

.)

Time (h)

IN PRESS: Journal of Controlled Release

In-situ MRI

Fluvastatin

1

2

3

Fluvastatin

1

2

3

pKa1 = 4.27pKa2 = 13.98pKa3 = 14.96

Formulation composition- 84.24 mg Fluvastatin Sodium, - 8.42 mg Potassium Bicarbonate, - 111.26 mg Avicel, 4.88 mg Povidone, - 16.25 mg HPC (Klucel HXF),- 97.5 mg HPMC (K100 LV), - 2.44 mg MgSt, - 9.75 mg Opadry Yellow (coating)

Hyphenation of USP-IV dissolution with MRI. LESCOL XL


In-situ MRIHyphenation of USP-IV dissolution with MRI. LESCOL XL

Water Maps

0

5

10

15

20

0 10 20 30 40 50

Dry core diameterGel layer thicknessTablet diameter

Radi

al th

ickne

ss (m

m)

Time (h)

0

5

10

15

20

0 10 20 30 40 50

Dry core diameterGel layer thicknessTablet diameter

Radi

al th

ickne

ss (m

m)

Time (h)

0 100%[H2O]



T2 relaxation mapsT2-relxation maps shown indicate a quite different behaviour:

- The structural integrity of the tablet remains intact, even after 42 hours.

- This indicates that the gel erosion process is now slow and evenly distributed.

- Collectively, figures 3 and 4 show that after 42 hours the gel matrix was highly hydrated and distributed.


(a) (b)


19F Signals: Combining imaging with high resolution spectroscopy


Conclusions

• Imaging: Seeing is believing.• Current and more complex formulation do

require more sophisticated analysis techniques.

• API dissolution need to be supported by more sophisticated test to ensure that the mechanism of drug dissolution is known and stable over time.

Acknowledgements & Credits

• Rob Saklatvalawww.linkedin.com/pub/robert-saklatvala/8/6a4/406

• Brett Cooperwww.uk.linkedin.com/pub/brett-cooper/3a/759/20b

• Sam Pygallwww.uk.linkedin.com/pub/samuel-pygall/10/139/532

• Agnieszka Jamstreszka• Katryn Bradleywww.uk.linkedin.com/pub/kathryn-bradley/20/789/8a

• Nick Gower• Jonathan Pritchard• James Mannwww.uk.linkedin.com/pub/james-mann/16/680/757

• Dr. Mick Mantle

www.uk.linkedin.com/pub/dr-mick-mantle/0/5a0/462

• Qilei Zhang

www.uk.linkedin.com/pub/qilei-zhang/1a/584/b33

• Prof. Lynn Gladdenwww.uk.linkedin.com/pub/lynn-gladden/39/221/617

BACKUP

Optical Microscopy

• Widely used!– Paolo Colombo demonstrated the gel layer

growth and the evidence of – Colin Melia

Swelling rate of: HPMC K4M, HPMC K100LV, PEO301, PEO1105

Correlation with in the physicochemical properties of the polymers.

Based on the approach of Gao and Meury and Paolo Colombo

(Colombo et al, 1999, Colombo et al, 1996, Li et al, 2005, Kiil and Dam-Johansen, 2003, Gao and Meury et al. 1996)

Optical Microscopy

HPMC K4M HPMC K100LV PEO 301 PEO 1105

60’

120’

180’

0’

(Colombo et al, 1999) (Avalle, Pygall Pritchard, Jamstrenzka - MSD, Unpublished)

Previous work findings:1. For a low solubility drug (MK-1) There is differing behaviour with respect to drug

release from CR matrices based on PEO and HPMC

2. The mechanism and the extent of drug and polymer dissolution varies greatly upon the polymer used

•Evaluating the performance of poly(ethylene oxide) (PEO), hydroxyethylcellulose (HEC) and hydroxypropy methylcellulose (HPMC) in erosion-based hydrophilic matrices for low solubility drugs –

(Pygall et al. in preparation)Time (hours)

0 5 10 15 20

% d

rug

rele

ase

0

10

20

30

40

50

PEO 1105PEO 301HEC 250HPMC K4MHPMC K100LV

Drug release from matrices- 125 mg of compound (MK-1 )-40% polymer

USP apparatus II dissolution test at 100 rpm, 37±1°C. Mean values (n=3) ± 1SD

Optical Microscopy

original tablet boundary

Raw Image: Light intensity expressed as a gray scale 0-255 (black to white)

Numerical average: each column of pixel is averaged for each position. Each column give one point on the chart. A &B indicates swelling and erosion front positions

-10

10

30

50

70

90

110

-300-200-1000100200300400500

position (microns)

Norm

alize

d sig

nal i

nten

sity

Physical tablet boundary @ t0

Time course: The process is repeated for each time point.From this plot we can calculate the erosionand swelling front A & B

60’

0

20

40

60

80

100

120

-400-300-200-1000100200300400500600

POSITION

Physical tablet boundary @ t0Erosion front

Swelling front

DISSOLUTION AND EROSION fronts fro PEO1105 as function of time

-300

-200

-100

0

100

200

300

400

500

600

0 50 100 150 200

time (minutes)

POSI

TIO

N

SWELLING FRONT

EROSION FRONT



0’

149’

49.5’

(not all curves are displayed)

The movement of the front position can be plotted as function of time.

A

B

The fronts position are taken fromThe mean intensity of the 2 inflection points (A) and (B)

Optical Microscopy

PEO1105PEO301

HPMC K100LVHPMC K4M

HPMC K4M HPMC K100LVPEO301 PEO1105

•The gel layer for each polymer matrix system clearly shows the development of different gel layer morphologies •With concomitant discrimination of swelling and erosion front profiles

•The PEO polymers expand rapidly and continue to expand over the next 2 hours.•The HPMC polymers expand then slows down after the first hour.

•The larger particle size and lower compressibility of PEO leads to faster gel layer formation compared to HPMC with smaller particle sizes and higher compressibility.

•The fast initial wetting and swelling of PEO implies that they are more hygroscopic than HPMC. This may be due to the hydrophobic methoxyl group in HPMC, or to the lower compressibility index, allowing the polymer to hydrate

faster.

Optical microscopy RESULTS

Optical microscopy: VALIDATION

IN SITU NIR OF HIGH SOLUBILITY DRUGS

• Chloropheniramine maleate• MK-2: Understanding Failure mode

Chloropheniramine maleate

CHLOROPHENIRAMINE MALEATE vs ACETYL SALICYLIC ACIDDissolution of 6.4 mm Flat Disc Tablets into Water Using Baskets

0

20

40

60

80

100

120

0 5 10 15 20

Time (hrs)

% C

laim

Case 2: highly soluble drug in a non-homogeneous formulations

5.5 g/L solubility in waterFormulation:10% drug loading, 20% HPMC


Mapping chloropheniramine maleate

0’

60’

4175μm

3250μm

PLS images for HPMC Water front movement

Erosion –based systems: High solubility / non homogeneous formulation

Mapping chloropheniramine maleate

Time (minutes)

Posi

tion

(mic

rons

)

Posi

tion

(mic

rons

)

-800

-700

-600

-500

-400

-300

-200

-100

0

0 10 20 30 40 50 60 70

WATER FRONT MOVEMENT AREA & PERIMETER DEPLETION

Time (minutes)

AREA

Perim

eter

Frame 1

Frame 1

y = 0.1143x + 98.557R2 = 0.9951

0

20

40

60

80

100

120

-800 -600 -400 -200 0

water front movement

% o

f par

ticle

dep

lete

d (p

erim

eter

)

0

10'20'

30'40'50'

60'

PERIMETER & FRONT MOVEMENT

Erosion –based systems: High solubility / non homogeneous formulation

Understanding Failure Mode

0

10

20

30

40

50

60

70

80

90

100

110

0 2 4 6 8 10 12 14 16 18 20Time / hours

% L

abel

Cla

imUnderstanding Failure modes

The problem

□ NON STRESSED● STRESSED 1 week 40°C / 75% RH

• MK-2 is an erosion based controlled release formulation, with a high load of a highly soluble, but slowly dissolving molecule.• Hypothesis: given the high drug loading the shift in dissolution could be caused by changes in the API.

Failure mode of erosion based matrices

Understanding Failure modes

Approach: replace the API with another of similar properties and study the dissolution

0

10

20

30

40

50

60

70

80

90

100

110

0 2 4 6 8 10 12 14 16 18 20Time / hours

% L

abel

Cla

im

6 % K100M_ Niacin - Initial6 % K100M_ Niacin - 1 week at 40/75 open6 % K100M_Caffeine - 1 week at 40/75 open6 % K100M_Caffeine - Initial6 % K100M_Caffeine - 1 week at 50/75 open

MK-2 NON STRESSEDMK-1 STRESSED 1 w 40°C / 75% RHCaffeine STRESSED 1 w40°C / 75% RHCaffeine NON STRESSEDCaffeine 1 week at 50/75


• The stressed formulation showed a much more rapid and deeper -initial- erosion which is then followed by a long period in which the dissolution appear slower.

• This is confirmed by both MK-2 release signal and water penetration signal

Understanding Failure modes

NIACIN SIGNAL

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

TABLET CORE

TABLET CORE

NIACIN SIGNAL

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

TABLET CORE

TABLET CORE

MK-2 signal: faster tablet erosion in stressed tablets


MK-2: Rate of Water penetrationfaster in stressed tablets


Understanding Failure modesFailure mode of erosion based matrices

Caffeine: Stressed vs Unstressed tablets

Caffeine was used as a control for MK-2 NIR in-situ work; data processing was applied as for Niacin-based formulation (MK2) And data showed no difference in API and water penetration rates between stressed and unstressed


-1- Significant erosion for stressed Niacin formulations-2- No gross changes in the position of dissolution and erosion fronts for Caffeine


Conclusion (seeing is believing)

NON STRESSED MK-2 STRESSED MK-2

Time lapse photography: Schematics

Pharmaceutical Research, Vol. 17, No. 10, 2000

Sequential Layer model Siepman Peppas 2000

NON STRESSED MK-2 STRESSED MK-2

Time lapse photography: Schematics

Pharmaceutical Research, Vol. 17, No. 10, 2000

Sequential Layer model Siepman Peppas 2000


paolo avalle discovery chemistry congress2012

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