Liquidborne Particle Counting using Light Obscuration and Light Scattering Methods

What has been . . .

Focus has been on injectable liquids

• Possibility to block capillaries and arteries –

Red Blood cells are about 5 µm

– –

Capillary (5 to 10 µm) Large veins (10 to 50 µm)

• Threat of microbial infection • Allergic reaction to foreign substances 2

Definition of Particulate Contaminants Unwanted insoluble matter that exist as “randomly-sourced extraneous substances”

• Excludes homogeneous monotonic materials that exist as a precipitate or suspension • i.e. colloids, drug degradation or otherwise derived from a defined source and can be analyzed by chemical means

Regarded as “contamination” and “adulteration” under Food and Drug Act

• the chemical composition of the particulate is varied, and would not be declared on the label –

Examples: bits of paper fiber, fragments of filler material, etc

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Liquid Particle Counting Applications Final Product Testing – USP <788>

• SVP or SVI (Small Volume Parenteral/Injectable) –

Ampoules, Vials

• LVP or LVI (Large Volume Parenteral/Injectable) –

IV (Intravenous) solutions Process contamination studies Decomposition studies (stability) DI or WFI Water Precision Cleaning – Medical Devices

• Aqueous • Other Chemicals 5

Other Applications for Particle Counting Medical Devices

• Cleanliness of manufacturing environment • Cleanliness of device before implantation –

pacemakers, stents, artificial arteries

• Cleanliness of reclaimed devices

Design of particulate-based medicines

• Inhalation therapies • Intentional occlusion of blood flow to cancers • Time-based dosages • Transdermal absorption 6

Global Regulations: Particles in Liquids USP <788>, EP 2.9.19, JP XV, KP, CP Primary method

• Optical Particle Counter [OPC] –

Light Obscuration Counter Secondary method

• Optical microscope –

Subjective

–

Labor intensive

–

Requires more time to process samples

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Proposed: USP 787, USP 1787 USP <787> Under discussion Focused on reducing necessary test volumes due to concerns of biotechnology manufacturers of cost for test Expensive and often very small dose factory

–

for example: 500 uL pre-filled syringe

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USP <787> Primary method ?

• Optical Particle Counter [OPC] –

Light Obscuration Counter Proposed: USP 787, USP 1787 Secondary method ?

• Optical microscope –

Subjective

–

Labor intensive

–

Requires more time to process samples

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USP <787> Small sample volume - 1 mL ??

Concerns with variability - within production lots - in analytical methods Proposed: USP 787, USP 1787

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Optical Particle Counter Optical Instrument

• Must move fluid through sensor • Can quantify particles from 100 nm to 5000 µm • Counts particles individually (one at a time) • Cannot tell you composition • But results are immediate 11

Many shapes and sizes

Alumino-silicate with K and Ti Talc 12

Largest Dimension d

Sizing Particles by Microscope

Martin’s Diameter d Area A Area B Ferret’s Diameter Projected Area d d 13

Challenges of Protein-based Products Handling can change material !!!

• Agitation • Heat and Light • Contaminates • Container: Vials versus syringes/cartridges • Shear forces

Key concern is Aggregation

• Reduction of native form (impacts efficacy) • Introduction of homogeneous aggregates • Introduction of heterogeneous aggregates 14

Challenges of Protein-based Products Transparency of most proteineous entities

• Refractive index • NIST working on calibration material

Not “contamination” but instead a shift from native form

• Not a solution as with small-molecule therapeutics • Formation of quaternary structures [dimers, etc.] • Protein complexes

Reconstitution of lyophilized product

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Refractive Index Key is the ability to distinguish between the particle and the surrounding fluid - needs to be great enough Optical response is proportional to comparative index

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Refractive Index Key is the ability to distinguish between the particle and the surrounding fluid - needs to be great enough Optical response is proportional to comparative index

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Refractive Index NIST working on protein-like calibration material

• Probably 2 years away • Exploring 2 methods of manufacture • Need thread-like material • Indices near water • Stable over reasonable period 18

II. Sample Handling

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Settling/Agitation Entrained gas - sonication probably not ideal with protein structures - light vacuum seems to work OK Settling Limits collection of particles - especially of greater mass - dependent on time and viscosity - improved collection with slanted containers

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Temperature Settling Probe position Consistency of sample characteristics

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Issues with Sampling Particles in Liquids Sampling Errors Account for most problems

Accidental Contamination or Alteration by Technician

1. System Preparation

Initial Cleanliness Calibration

2. Sample Preparation

Contamination - Particles - Gases - Liquids

3. Sample Handling

Aggregation Settling Cavitation 22

Largest Dimension d

Sizing Particles by Microscope

Martin’s Diameter d Area A Area B Ferret’s Diameter Projected Area d d 24

HIAC Liquid Particle Counters Example: HIAC 9703

• The industry standard liquid particle counter since 1997 • USP <788> was written specifically around HIAC technology • Every major manufacturer of particle calibration standards uses the HIAC 9703 25

 Reproducibility  Repeatability

HIAC Liquid Particle Counters Example: HIAC 9703+

• Improved sample mounting method for small vials or containers • Detection of usual conditions such as bubbles or contamination • Proven syringe sampler • SVI and LVI sampling • Addresses non-compendial applications, e.g. R&D and other low frequency, small sample volume applications 26

Detection Ranges 0.1µm 1µm 10µm 100µm Light Obscuration Light Scattering 1 nm 10 100 1000 10000 100000 1000000

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Light Obscuration Light Obscuration Sensors and system

• also known as Light Extinction • also known as Light Blocking 28

Principles: Light Obscuration Detector Output

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Principles: Light Obscuration Detector Output

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Principles: Light Obscuration

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Particle Detection Like an air particle counter, the larger the particle, the larger the pulse that is created

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Light Trap

Principles: Light Scattering

Detector

Detector Output

Mirror Laser Diode 33

Light Trap

Principles: Light Scattering

Detector Particle Laser Diode Mirror 34

Advantages: Light Scattering Good sensitivity from 0,1µm to 50µm Wide range of sample concentration Good rejection of false counts High sample flow rates

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Disadvantages: Light Scattering More complicated construction = higher cost Characteristics of particle surface (shiny, color) affect response

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Effect of colors and surfaces on Light Scattering Dark Light Shiny

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Alumino-silicate with K and Ti Talc 38

Largest Dimension d

Sizing Particles by Microscope

Martin’s Diameter d Area A Area B Ferret’s Diameter Projected Area d d 39

General Comments on Liquid Counting Particle Counters Report Size

–

But measure an Optical Response

–

Difference in reported size compared to microscope

Projected Area d

Calibration Relates the Optical Signal to Size

–

Difference between calibration material characteristics and “real world” particles

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Particle Counters Report Size

• But measure an Optical Response • Differences in reported size compared to microscope

Calibration Relates the Optical Signal to Size

• Difference between calibration material characteristics and “real world” particles

General Comments on Liquid Counting

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LO results versus LS results Light Obscuration [LO]

• Good immunity to variations of surface and morphology • • Very stable Limit of quantitation circa 1.2 – 1.3 microns

Light Scattering [LS]

Results affected by surface characteristics and coloring Good stability Limit of quantitation sub-micron

Problem can occur in the attempt to correlate results of these two methods above 1 micron

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System Preparation 2-step Verification - optional:

• Run 2 test solutions –

Blank (“particle-free” water)

–

Aqueous solution containing known counts

at 10 µm or 15 µm In the range of 1000 to 3000 counts per mL • Frequency – based on risk analysis –

Each morning

– – –

Shift change Change of operator Other interval

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System Preparation Check for bubbles in sample lines and syringe

• Affects flow rate and calibration

Verify correct calibration curve installed

• Different flow rates for same sensor • Change of syringe size • Some companies have multiple sensors

Verify calibration is current

• Sensor resolution and response curve • IST tests conducted [USP, JP] 45

System Preparation Instrument Standardization Tests [IST]

• Five tests of system –

Volume accuracy

– – – –

Flow rate accuracy Calibration of sensor Resolution Count accuracy

• Required by USP and JP but not EP –

USP <788> 31

“…at intervals of not more than six months.” –

JP <24>

“…at least once a year.” 46

Sample Preparation Removing residue of previous sample

• If previous sample contained particles, may be good plan to run a “particle-free” blank before the next sample • Use liquid that is compatible with sample fluid –

An aqueous blank could trigger false counts in an oil-based sample by causing immiscible droplets

–

Potential residue from previous sample can cause change of counts

Data from first run of series is often discarded 47

Contamination • Particles • Gases • Liquids

Sample Preparation

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Sample Preparation

True Particle Sources • Residue on glassware and equipment • Ambient air • Paper dust • Glass • Diluent • Residue from previous sample • Colloidal suspensions False Particle Sources • RF signals or other electronic interference • Bubbles from entrained gases 49

Sample Preparation Work in controlled environment Use particle-free gloves Let water flow for 200 mL or more after opening a valve / tap

• Opening / closing valve generates particles • Wipe the outside of containers before sampling Particles on surface of vials or ampoules

Open vials and ampoules away from beaker or flask

• Particles from activity can fall into open container • Wash outside of containers to reduce potential particle source 50

Three common methods

• Allowing to stand in ambient air Risk of large particles settling • Sonification [ultrasound] 80 to 120 watts [USP <788>] 30 seconds [USP <788>] • Vacuum Bell jar or dessicator 0.6 – 0.8 atmospheres for 2 to 10 minutes

Degassing sample

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Sample Preparation Possibility of decreasing true particle counts

• Settling • Lack of agitation • Position of probe in sample vessel • Remaining material from previous sample run – –

Sample with lower counts Blank

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Aggregation Settling Entrained gases

Sample Handling

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Sample Handling Aggregation

• Samples held over time or at extremes of temperature can develop aggregates of smaller particles • • Exposure to light can also trigger this reaction • Sub-micron particles can thus add to particle counts above 10 µm

Suggestion:

–

Profile counts under 10 µm [e.g. at 2 or 5 µm] in addition to standard count sizes at 10 and 25 µm

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Sample Handling Settling

• Undercounting caused by –

Gravitational settling

–

Failure to suspend particulate matter adhering to walls or stopper of container

• Standards have recommended procedure for agitation –

Multiple inversions of original container before decanting

–

Constant motion of liquid during sampling

“Gently stir the contents of the container by hand-swirling or by mechanical means…” USP <788> 55

Optical Particle Counter Instrument & sensor

• Must move fluid through sensor • Can quantify particles from 100 nm to 5000 µm • Counts particles individually (one at a time) • Cannot tell you composition • But results are immediate 56

Questions?

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Patent Pending

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New Hardware: 9703+ Key features

Auto stop for sensor elevator arm Small vial holding clamp Sample probe with reduced dead volume Back-flush and forward flush from front panel Supports MC-05 sensor (0.5 micron sensitivity) 59

Software: PharmSpec 3 Key new features

Compendial test support continues

USP, EP, JP, KP looks same as previous PharmSpec versions

Uses same log on as for Windows Improved Report format Improved Error Detection and Display 60

Still the HIAC 9703 you know and trust – only better!

Syringes

• 1 ml, 10 ml, 25 ml

Flow rate settings

• 10 to 100 ml

Sensors

• MC-05 is added

Sampling Probes

• added shorter small-bore probe

Instrument size / shape

• 50%+ of instruments are placed in laminar flow cabinets.

• Smooth, curved surfaces create less turbulence for the air flow 61

Easier, Faster and Confident Sampling Use less sample, save valuable time - protect your investment

• Small vial clamp ensures that sample does not spill during testing • Probe needle safety switch prevents probe damage • New small needle probe with industry’s smallest tare volume 62

Easier, Faster and Confident Sampling Reduce uncertainty from data anomalies

• Bubble alarm notification • Contamination alarm notification • Advanced notification when service or calibration is due • Invalid configuration notification 63

HIAC just got easier!

Less time needed for clean-up

• Automated flushing and cleaning routines • Push a button, walk away and return to a clean sensor

Export your data with ease

• Select one, several or all of your historical data records with our batch export utility • Select PDF, Word, Excel, or text files

Save time with electronic signature

• Stricter interpretation of 21CFR Part 11 electronic signature process…. WITHOUT more manual inputs • Remembers user Login ID 64

HIAC 9703+ Flexibility Interchangeable sampling probes, syringes, and sensors

Ensure you have one instrument to manage all applications Now supports MC05 sub-micron sensor Change configuration with no impact to instrument validation

Customized reporting

Customize the number of reviewers and approvers for compendial test reports Add company logo, user-defined descriptors

Customized test recipes

Procedure Builder enables the development of unique test recipes for your application Enables testing to marketing license-specific applications 65

3 probes available

• Tare

¼” ID =1.2 ml tare volume

1/16

” ID = 0.172 ml tare volume

• New small / short probe 1/16

” ID = 0.09 ml tare volume

Tests can be performed on 1 ml of product!

New Sampling Probe

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Small Vial Clamp Platform

• Can be retrofitted • Can be removed

Ease of Use

• Use one hand to compress lever arms • Use other hand to place sample in central location • Decompress hand • Clamp auto-centers and holds sample container

Small Vial Clamp

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Docking Module

• Enables removal/disabling of the stir bar mechanism • Enables field installation of small vial clamp outside of the laminar flow cabinet –

Avoid potential of re qualification that can occur if instrument is moved Future developments to expand 9703 applications Docking Module

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Sampling safety switch

• Ensures the sampling probe does not crash (and bend or break) into the docking module • Ensures the probe does crash into or tip the sample container

Sampling Safety Switch

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Liquidborne Particle Counting using Light Obscuration and Light Scattering Methods

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