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Malvern Usp 429
Malvern Usp 429
Malvern Usp 429
Measures of precision:
Repeatability – same group of particles measured consecutively in a set
of measurements: intra-assay precision
Machine + Method (changes in material during measurement: time)
Reproducibility – another sample; more samples: batch heterogeneity
Machine + Method + Material
Robustness – how a key variable affects the end result (Club selection?
Imaginary part of RI)
3 R‘s sorted out in method development
But bulk versus primary/dispersed?
Dry powder – PST - Pressure-Size Titration – DP of
venturi
No plateau
Compare with dispersed wet result
Sample heterogeneity - reproducibility
Wet – BDAS – before, during and after sonication
(wetting, separation, stabilization)
Plateau possible
Microscopy to assist
Repeatability/intra-assay precision (single sample)
Calculation of mean and RSD/CV for 10 or more
results
Effect of time
Stability, stability, stability
Heywood again – myth of sensitivity
A number counter isn‘t necessarily more sensitive to the fine material –
otherwise a 100 nm latex would be measured just fine on a visual light
image analyzer or microscope
Bottom-end imposes a mathematical as well as a physical barrier – the
harder we look then the more we see:
Dr. H Heywood: Dr. Bronowski asked whether anyone could comment upon
automatic star counting methods; although I cannot say what methods were used for
this purpose there was an interesting similarity between star and particle counting
results when the limit of visibility was approached. Early records of microscopical
counts, when extended to the extreme limit of resolution, showed a false maximum
frequency just greater than the lower limit, whereas it is now known that the
numerical frequency for normal dusts increases continuously as the particle size
decreases. Star counts made by unaided vision about 300 years ago showed a
maximum frequency at about 4th magnitude, the limit being around 6th magnitude.
With the aid of telescopes it was easily shown that the numerical frequency of stars
increased as the magnitude increased, i.e. brightness decreased. The important
feature of this comparison was that false effects might be observed if any counting
process was worked to the extreme limit of its capabilities.
Taken from Table 1.5 page 38, T.Allen Particle Size Measurement 5th Edition Volume 1
Chapman and Hall 1997 ISBN 0 412 72950 4
~ 6300 g
―I wish to detect a small amount of
agglomeration in my system‖
n e^-0.1 0.1^n n! P
0 0.905 1 1 0.9050
1 0.905 0.1 1 0.0905
2 0.905 0.01 2 0.0045
3 0.905 0.001 6 0.0002
4 0.905 0.0001 24 0.0000
5 0.905 0.00001 120 0.0000
6 0.905 0.000001 720 0.0000
80
Transmission [%]
30
70
Dv(50) [µm]
60
50 20
40
Manual sampling
30
10
20
10 Set Point
0 0
0% = Lots of particles 0 50 100 150 200 250 300 350 400
80
70 Transmission Dv(50)
Transmission (%)
60
Particle Size (µm)
50
40
30
20
10
0
0 100 200 300 400 500 600
alan.rawle@malvern.com
+ 1 508 768 6434
Questions?
Appendix
References
Basic Principles
T Allen Particle Size Measurement Volume 1 Powder sampling and particle
size measurement Fifth Edition Chapman and Hall (1997)
R D Cadle Particle Size Determination Interscience New York (1955)
G Herdan Small Particle Statistics Second Revised Edition Butterworths
London (1960)
J M DallaValle Micromeritics The Technology of Fine Particles Second
Edition Pitman New York (1948)
Clyde Orr, Jr. Particulate Technology The Macmillan Company New York
(1966)
Clyde Orr, Jr., J M Dallavalle Fine Particle Measurement Size, Surface and
Pore Volume The Macmillan Company New York (1959)
H Heywood The Scope of Particle Size Analysis and Standardization‖ in
Symposium on Particle Size Analysis Supplement to Transactions,
Institution of Chemical Engineers, Vol. 25, 14 – 24 (1947)
R R Irani, C F Callis Particle Size: Measurement, Interpretation and
Application John Wiley & Sons Inc., New York (1963)
Appendix
References
Basic Principles
H E Rose The Measurement of Particle Size in Very Fine Powders London
Constable & Company (1953)
American Society for Testing Materials Symposium on New Methods for
Particle Size Determination in the Subsieve Range Washington Spring
Meeting March 4, 1941 ASTM, Philadelphia, U.S.A.
A Jillavenkatesa, A J Dapkunas, L-S H Lum Particle Size Characterization
NIST Practice Guide Special Publication 960-1 U S Government Printing
Office Washington (2001)
M J Groves Particulate Matter Sources and Resources for Healthcare
Manufacturers Interpharm Press Buffalo Grove, Illinois (1993)
H G Schroeder, P P DeLuca ―Theoretical Aspects of Particulate Monitoring
by Microscopic and Instrumental Methods‖ Journal of the Parenteral Drug
Association Vol. 34, No. 3 183 – 191 (May - June 1980)
S J Borchert et al Particulate Matter in Parenteral Products: A Review
Journal of Parenteral Science & Technology Vol. 40, No. 5 212 – 241,
(September – October 1986)
Appendix
References
Basic Principles
H G Merkus Particle Size Measurements Fundamentals, Practice, Quality
Springer (2009)
Clive Washington Particle size analysis in pharmaceutics and other
industries: theory and practice Ellis Horwood (1992)
A F Rawle Instrument Qualification and Performance Verification for
Particle Size Instruments Chapter 12 in Practical Approaches to Method
Validation and Instrument Qualification Ed. C C Chan, H Lam and X M
Zhang 255 – 298 Wiley (2010)
A F Rawle Analytical Tools for Suspension Characterization Chapter 6 in
Pharmaceutical Suspensions: From Formulation Development to
Manufacturing A K Kulshreshtha et al (Eds.) Springer Science + Business
Media, 177 – 230, DOI 10.1007/978-1-4419-1087-5_6 AAPS (2010)
A F Rawle Particle Morphology and Characterization in Preformulation
Chapter 3.2 in Preformulation in Solid Dosage Form Development M C
Adeyeye, H G Brittain (Eds.) Volume 178, 145 – 184, Informa Healthcare
(2008)
A F Rawle The Basic Principles of Particle Size Analysis Malvern
Application Note MRK-034 (1993) Downloadable:
http://www.nbtc.cornell.edu/facilities/downloads/Basic%20principles%20of%20particle%20size%20analysis.pdf
ISO 9276-2:2001 Representation of results of particle size analysis -- Part
2: Calculation of average particle sizes/diameters and moments from
particle size distributions
Appendix
References
Basic Principles
R A Mugele, H D Evans ―Droplet Size Distribution in Sprays‖ Ind. Eng. Chem. 43(6),
1317 – 1324 (June 1951)
M Alderliesten: 5 part series in Part. Part. Syst. Charact. (from 1990 – 2005) [Part 6 in
press]
M Alderliesten Mean Particle Diameters Part I: Evaluation of Definition Systems Part.
Part. Syst. Charact., 7 (1990) 233-241
M Alderliesten Mean Particle Diameters Part II: Standardization of Nomenclature Part.
Part. Syst. Charact., 8 (1991) 237-241
M Alderliesten Mean Particle Diameters Part III: An Empirical Evaluation of
Integration and Summation Methods for Estimating Mean Particle Diameters from
Histogram Data Part. Part. Syst. Charact., 19 (2002) 373-386
M Alderliesten Mean Particle Diameters Part IV: Empirical Selection of the Proper
Type of Mean Particle Diameter Describing a Product or Material Property Part. Part.
Syst. Charact. 21 (2004) 179 – 196
M Alderliesten Mean Particle Diameters Part V: Theoretical Derivation of the Proper
Type of Mean Particle Diameter describing a Product or Process Property Part. Part.
Syst. Charact. 22 (2005) 233 – 245
ASTM E 2578 – 07 Standard Practice for Calculation of Mean Sizes/Diameters and
Standard Deviations of Particle Size Distributions
The ISO terminology for particle size
A F Rawle The Basic Principles of Particle Size Analysis Malvern Application Note
MRK-034 (1993) Downloadable:
http://tinyurl.com/2c4unfn
Continuous and discontinuous distributions
Source of confusion
Contrast a screen with light scattering
The number of size classes in a histogram
affects the vertical axis - a density
distribution is required in the vertical axis
A frequency curve is a continuous
distribution
Can pass through any point in a histogram bar
An infinitely small increment in size means an
infinitely small quantity in the vertical axis
An undersize plot avoids the confusion in
horizontal or vertical axes
Reconciliation
Pitard/Gy:
MS = 18 r f d3/σ2
(where d refers to 95% passing)
f = shape factor (= p/6 for spheres, 1 for cubes)
Rawle:
MS = 20 r (p/6) d3/σ2
(for x95; spheres considered)
Above: Henry A Vezin‘s calculations in Richards‘ Ore Dressing (Volume II page 850)
Interestingly Reference ‗14‘ in Richards is to H O Hofman Metallurgy of Lead‖ (1899)
where on page 45 (not 42 – the start of the chapter), Vezin is named with an 1866 comment
but no actual text reference is given….. (Hofman was a MIT Professor like Richards)
Appendix – Sampling
Rawle-Vezin comparison
d (mm) d/10000 d^3 Density (g/cm3) 10000*100*(PI/6) Minimum Mass (g) Vezin (1865/1866) Vezin/Rawle
1 0.0001 1E-12 2.6 523598.7756 0.000001
10 0.001 1E-09 2.6 523598.7756 0.001361
100 0.01 0.000001 2.6 523598.7756 1.36
125 0.0125 1.9531E-06 2.6 523598.7756 2.66 2.92 1.10
200 0.02 0.000008 2.6 523598.7756 10.89
250 0.025 1.5625E-05 2.6 523598.7756 21.27 23.3 1.10
500 0.05 0.000125 2.6 523598.7756 170.2 186.7 1.10
1000 0.1 0.001 2.6 523598.7756 1361.4 1493 1.10
1500 0.15 0.003375 2.6 523598.7756 4595
2000 0.2 0.008 2.6 523598.7756 10891 11950 1.10
4000 0.4 0.064 2.6 523598.7756 87127 95570 1.10
5000 0.5 0.125 2.6 523598.7756 170170
8000 0.8 0.512 2.6 523598.7756 697015 764600 1.10
10000 1 1 2.6 523598.7756 1361357
Neal Zupec
Research Scientist/Baxter Healthcare Corporation
• instrument considerations
• Remove four portions, not less than 5 mL each, and count the
number of particles equal to or greater than 10 µm and 25 µm.
Disregard the result obtained for the first portion, and calculate
the mean number of particles for the preparation to be examined.
≥10 µm ≥25 µm
Calibration Validation
Limits
• stir bar – can be used with either sampler type (as long as it is
not a source of particulate matter itself)
Laser diode
Laser diode
0 100 200 300 400 500 600 700 800 900 1000
Channel No.
Baxter Healthcare Corporation
Sources of Erroneous Count Data
• particle overconcentration
• demonstrates compliance
Questions?
• Method Scope
• Equipment & Lab
• Operator Training
• Important Steps
• Blank Control
– USP 788 blank requirement
– Lab control blanks
• Counting and Sizing: full and partial
– Quantitative practices
– Additional information
• Calculations
• Limits
Method Scope
• Method-specific
design
• Linear scale
graduated in 10µm
increments
• Certified at
installation using a
NIST calibrated stage
micrometer
• Standardizes the
source of calibration
across manufacturing
sites
• Facilitates partial-
counting
Microscope Alignment
• Quantitative practices
– Full count for all samples is preferred
– Partial count for high level of retained particles - above 1000
• Additional information may be sought
– Size thresholds
– Particle type assessment
Sizing and Tabulation
• The microscopic test is flexible in that it can count from very few to
many particles over the entire membrane
• The operator counts all particles on the membrane surface or
effective filtration area (EFA). The graticule field of view (GFOV),
defined by the large circle of graticule, may be ignored and the
vertical crosshair used as the counting plane.
• Scan the entire membrane in a path that adjoins but does not
overlap the first scan path
• Repeat the procedure, moving the membrane under the
objective until all particles on the membrane are counted
• Tabulate:
– Total number of particles that are ≥10µm - 25µm = P10-25
– And the number that are ≥ 25µm or larger = P25
Total Count <788>
The ophthalmic solution meets the requirements of the test if the average
number of particles present in the units tested does not exceed the
appropriate value listed in Table 4
A
B
C
Dynamic Light Scattering (DLS): a good (high-
resolution) inversion algorithm is needed to
analyze “difficult” PSDs (e.g., multimodals)
B C
Single-Particle Optical Sizing (SPOS), Part I:
Conventional Light Extinction (LE) is effective
for counting/sizing particles above 2-um
USP<729>
Single-Particle Optical Sizing (SPOS), Part II:
Light scattering (LS) provides added sensitivity,
needed to count/size particles down to 0.5-um
Emulsion LD:
1.01 x 106/mL
Emulsion LM:
4.47 x 106/mL
Emulsion LN:
7.37 x 106/mL
FX-Nano: powerful tool for analyzing submicron
agglomeration of proteins and other biologicals
in injectable pharmaceutical formulations
Kevin Powers
USP Workshop on Particle Size:
Particle Detection and Measurement
December 10, 2010
Always Look at your Particles under a Microscope!
No Exceptions!
• Particle Count
• Particle Size Distribution
• Particle Shape
• Other information
With Respect to USP 788/789
INTERNATIONAL ISO
STANDARD 13322-1
Particle size analysis — Image analysis methods —
Part 1:
Static image analysis methods
Analyse granulométrique — Méthodes par analyse d'images —
Partie 1: Méthodes par analyse d'images statiques
Dynamic Image Capture
Resolution is a function of both the microscope and the Image capture device. The
microscope’s resolving power is dependent on its optics and physical principles.
The image can also be captured or digitized at varying resolutions.
d 0.6
no sin
• In general optical size and shape information is difficult to obtain below 3-5mm
• Electron microscopes can resolve much smaller particles with greater
depth of field but suffer from lower contrast levels STED
Eva Rittweger, Kyu Young Han, Scott E. Irvine, Christian Eggeling, Stefan W. Hell (2009). "STED microscopy reveals crystal
colour centres with nanometric Resolution.". Nature Photonics 3: 144–147.
Magnification 2- 10 mm?
Recommended Magnifications for selected size ranges
(ISO 13322-1 Static Image Analysis)
Sample size range Resolution (mm) Field size (mm) Total Magnification
0.1 – 1.0 0.01 10 x 10 20Kx
0.3 – 3.0 0.03 30 x30 8Kx
1 –10 0.1 100 x 100 2Kx
3 – 30 0.3 300 x 300 800x
2000
1800
Population 10293
120.00% Morphology
100.00%
1600
Comparison of Laser Diffraction Data and Optical Analysis
1400 Frequency for GCP 3-30 Carbon (10196 particles)
80.00%
Cumulative % 14
1200
Frequency
1000 60.00% 12
800
10
Differential Volume %
40.00%
600
8
400 Optical GCP 3-30 carbon
20.00%
Instrument A
200
6
0 .00%
4
2
8
4
2
1
9
2
6
1
3
6
7
3
3
8
2
6
6
9
07
92
55
00
4
04
11
17
28
45
72
14
83
65
42
.8
.8
.0
.9
.4
1.
4.
9.
3.
6.
12
20
0.
2.
11
18
30
47
76
12
19
30
49
78
0.
0.
0.
0.
0.
0.
1.
1.
4.
7.
Diameter (microns)
2
0
1 10 100
X MAX
4 Area
XA X MIN
p
USP 788
The projected area of each particle can be converted to the
area equivalent circular diameter, xAi.
4 Ai X = 10µm
x Ai
p A=78.54 µm2
Is 100x enough?
Image Analysis Software – See like a computer
Image analysis software can differ in capabilities and may vary in the way
operations are defined. Operators have a wide variety of image
manipulation, sorting, and measurement options which can aid or hinder
statistical reliability.
• Border definition
• Threshold levels Contrast!
• Area definition (holes – euler number)
• Filters and image processing operations
• Splitting agglomerates or touching particles
• Size and length Measurements
• Discarding agglomerates
• Roundness and shape factors
• Data Presentation and Statistics
Area and aggregates
2000 120.00%
1800
100.00%
Optical Image Analysis
1600
1400 Frequency
on Spherical Reference
80.00%
1200
Cumulative %
Powder 3-30 microns
Frequency
1000
800
60.00%
vs Laser Diffraction
40.00%
600
400
20.00% Comparison of Laser Diffraction Data and Optical Analysis
200 for GCP 3-30 Carbon (10196 particles)
14
0 .00%
4
2
8
4
2
1
9
2
6
1
3
6
7
3
3
8
2
6
6
9
07
92
55
00
04
11
17
28
45
72
14
83
65
42
.8
.8
.0
.9
.4
1.
4.
9.
3.
6.
12
20
0.
2.
11
18
30
47
76
12
19
30
49
78
12
0.
0.
0.
0.
0.
0.
1.
1.
4.
7.
Diameter (microns)
10
Differential Volume %
Number distribution is derived 8
Optical GCP 3-30 carbon
distribution 2
0
1 10 100
Diameter (microns)
Comparison Image Analysis and Light
Scattering (1-10micron spheres)
MBP 1-10 Glass Powder
(4038count)
12
Relative Volume %
8
0
0.1 1 10 100 1000
Diameter (microns)
MMD GSD
Instrument A 4.73 1.38
Instrument B 4.77 1.42
Optical 4.46 1.46
Shape
INTERNATIONAL ISO
STANDARD 9276-6
Representation of results of particle size analysis —
Part 6:
Descriptive and quantitative representation of particle shape and
morphology
Représentation de données obtenues par analyse granulométrique —
Partie 6: Description et représentation quantitative de la forme et de la
morphologie des particules
Pop Quiz EMA 5008 - What is Spherical?
A Little Help?
More Help
54 93 42 77 81 7 58 36 39 105 61 46 59
What percentage of these
particles are spherical?
11 68 62 111 41 74 8 82 38 89 53 116 12
102 76 72 26 3 22 6 71 44 32 47 37 73 94
Water Formulations
PBS
Drug Bloodstream
pH Isotonic Buffers Blood
Culture media Salts Cells
Surfactants Proteins/Opsonins
Rheology Modifiers Filtration
Shear!
How do Particles Behave?
Image Analysis for Complex systems
• The same issues with other Particle Size Analysis methods also
apply to sizing by Image Analysis - plus a few more!
Council of the
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Board Expert
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USP Council of Experts – General Information
Professional Background
2% 1%1%1%
4%
8% Industry - 175
Academe - 62
Consultant - 36
Government - 25
11% 53% Practitioner - 15
Other - 7
Association - 5
Pharmacopeias - 5
Retired - 2
19%
USP Council of Experts – Expert Committees
• Examples:
• Unfractionated Heparin Expert Panel
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Requirements to serve as expert volunteers