Liquid Bourne Particle counter (LBPC)
Objective: To lay down the procedure for Cleaning, Operation, Maintenance, and Analysis of samples using LBP counter.
Scope: This SOP is applicable for the Operation, Cleaning, Maintenance and Analysis of LBPC samples in the manufacturing department.
Responsibility: Microbiologist / IPQA personnel are responsible for follow the procedure.
Accountability: Head – Quality Assurance /Nominee is accountable for overall compliance of SOP.
Procedure:
Make: Particle Measuring Systems, Model: LS 200
Collection of samples during the rubber stopper washing:
Rinse water sample of rubber stoppers shall be collected during the washing cycle of the rubber stoppers
Collect around 100 ml sample of the rinse water of rubber stoppers from the sampling port provided after the completion of the PW wash (Wash – 2).
Transfer the collected sample to the IPQC room for further testing/Analysis.
Pipette out around 5ml of the sample to the Micro for BET analysis.
Subject remaining sample of the rinsed water sample of rubber stoppers for LBP testing.
Place the rinse sample of the rubber stoppers on the sample port.
Perform the LBPC testing of the rinse water of rubber stoppers placed under the sample port.
Collection of samples during the Vial washing:
Initially Collect 25 washed vials for (10 ml, 15 ml, and 20 ml) and 12 washed vials for (50 ml and 100 ml) testing. Collect the samples using forcep, wrap the collected vials in a sterilized aluminum foil, and transfer to the IPQC room.
Fill the vial with suitable quantity of WFI using pipette and collect in a bottle, till we get approximately 100 ml washed vial rinse sample for visible and sub-visible particulate matter testing.
Place the sample collected from washing on the sample port.
After every four hours Collect 05 washed vials for (10 ml, 15 ml, 20 ml, 50 ml and 100 ml), fill the vial with suitable quantity of WFI, till we get approximately 50 ml washed vial rinse sample for visible and sub-visible particulate matter.
Collection of samples during the Vial filling
Collect filled vials for visible and sub-visible particulate matter testing
Reconstitute the Vials with WFI during initial and for subsequent containers thereafter check the solubility and clarity of the solution for visible and sub visible particulate matter and perform LBPC testing.
Take the reconstituted vials for visible testing and then collect the reconstituted volume to perform LBPC testing
For Test Sample preparation, testing of visible and sub-visible particles.
Interpretation:
The average cumulative counts, which are obtained in the three runs, are to be taken for calculation.
Calculations:
Pooled samples (small-volume injections): – average the counts from the three runs. calculate the number of particles in each container by the formula:
PVT / VAn,
Which,
P is the average particle count obtained from the portions analyzed,
VT is the volume of pooled sample, in ml,
VA is the volume in ml, of each portion analyzed, and n is the number of containers pooled
Individual samples (small-volume injections): – average the counts obtained from the three runs from each separate unit analyzed, and calculate the number of particles in each container by the formula:
PV / VA ,
Which,
P is the average particle count obtained from the portions analyzed,
V is the volume, in ml, of the tested unit, and VA is the volume in ml, of each portion analyzed.
Individual unit samples (large-volume injections): – average the counts obtained for two or more 10 ml aliquot portions taken from the solution unit. calculate the number of particles in each ml of injection taken by the formula:
P / V,
Which,
P is the average particle count for an individual 10 ml sample volume, and V is the volume, in ml, of the portion taken.
For all types of product, if the tested material has been diluted to effect a decrease in viscosity or expected counts are very high, the dilution factor must be accounted for in the calculation of the final test result
Acceptance Criteria:
The product meets the requirement of the test if the average number of particles present in the samples tested does not exceed 6000 particles of ≥ 10 microns and 600 particles of ≥ 25 micron size per container in case of small volume injections (or) as per in-house specifications.
In case of large volume injections ≥ 10 micron size particles should not exceed 25/ml and ≥ 25 micron size particles should not exceed 3/ml (or) as per in-house specifications.
In case of rinse samples ≥ 10 micron size particles should not exceed 60 particles /ml and ≥ 25 micron size particles should not exceed 6 particles/ml.
Operation of LBPC:
Ensure that the LAF working area is clean and check the differential pressure of the LAF is within limit.(8-15 mm of water column)
Ensure that the instrument is properly calibrated and has proper calibration tag.
Switch on the mains power of computer and then the instrument, liquid syringe sampler, laser, CPU, and monitor.
Double click on sampler sight pharma icon on the desktop.
Monitor displays the standard format of the analysis window.
Go to File in the main menu and click on the Log on and enter the user name and password.
Click on Configure> Instrument on the toolbar and click on Enable tab to enable the sensors.
Clean the syringe with MilliQ water/particle free water/water for Injection by inserting the sampling tube in a beaker filled with MilliQ water/ particle free water and then clicking on the flush icon on the Perform menu.
Blank check for ‘test environment’:
To perform Blank Check for ‘Test Environment’, use a cleaned bottle of the type and the volume representative of that to be used in the test.
Take 5 cleaned bottles/particle-free containers and fill them with approximately 25 mL of particle-free water (Ensure that the temperature of the particle-free water should not be more than 400 C) and agitate the water sample in the cleaned glassware by inversion or swirling.
Pool 5 cleaned bottle samples in one particle-free container to make up the volume of 100 ml and agitate the pooled sample by inversion or swirling.
Degas by sonicate for about 20 minutes or by allowing it undisturbed until it is free from air bubbles.
Swirl the bottle containing the water sample by hand or agitate by mechanical means to suspend particles.
Place the bottle under the sampler and operate as per the operating procedure.
Measure the blank sample by clicking on the start button. The system will start measuring the sample 6 times of 5mL each and an average count of different micron sizes is displayed.
Click on Configure > Sample on the toolbar.
Select the Sample configuration name to Blank Determination
Click on Modify
Set the system parameters to the following.
- Repeat Count – 6
- Discard Count – 1
- Sample Delay (secs) – 0
- Containers Pooled – 1
- Container Volume (ml) – 100
- Drain Rate (ml/min) – 50
- Fill/Flow Rate (ml/min) – 20
- Syringe Volume (ml) – 10
- Tare Volume (ml) – 0
- Sample Volume (ml) – 5
If more than 25 particles of > 10-micron size are observed for the combined 25 mL sample, either the environment is not suitable for particulate analysis or the filtered water and glassware have not been properly prepared.
In this case, if cleaned bottles are used for the test then send all the bottles for rewashing, and then clean the exterior and interior surface of glassware with particle-free water. After cleaning take fresh particle-free water and repeat the process until the above criteria are met.
Perform a Blank Check for “Test Environment” start of the testing day.
Test Procedure:
Test preparation:
Switch on the LAF 30 minutes before the sample analysis.
Prepare the test specimens in the following sequence.
Wear powder-free gloves while doing analysis.
Out side of the laminar enclosure, remove outer closures, sealing bands, any loose or shedding paper labels etc.
Rinse the exteriors of the Containers with particle free water and mop with lint free cloth.
Mop the LAF with 70% IPA and place the containers under the laminar air flow to protect the container from environmental contamination until analyzed.
Withdraw the contents of containers under test in a manner least likely to generate particles.
Sample Analysis:
Click on Configure> Sample on the tool bar
Select the appropriate Sample configuration name.
Click on Modify
Set the system parameters as requirement.
Click on the Channels tab.
Set and enable the channels 10 and 25 µm.
Click on OK until returned to the Sampler Sight-Pharma main screen.
Click on Configure.
Click on Display tab.
Select Cumulative and Counts per Milliliter/container from the drop down selection list.
Set the precision to 4 significant level.
Click on Labels tab.
Enter the Three desired labels.
Click on OK until returned to the Sampler Sight-Pharma main screen.
Reconstitute the product using particle free water/ water for injection tested for blank check. (for finished product analysis.)
Keep the product container on the slab and place the stirrer into the container and switch on the Magnetic stirrer and adjust its speed.
- Insert the sample tubing in the product container, without touching the lower half of the tube.
- Click on the Start button on the main screen.
- Enter the Labels for the product
- Click on the GO button to start the analysis.
- After the analysis is over click on the File menu.
- Click on Print Sample to take a printout of the sample.
- After completion of the test clean the sampler with MilliQ water/particle-free water/ water for injection by pressing the flush on the Perform menu.
- Enter the details in the logbook.
- Clean the working area after completion of the analysis.
Precautions:
Do not Halt/Stop the instrument unless it is required.
Ensure the instrument communication status every time in the display during the analysis.
After each sample analysis flush the syringe with particle free purified water.
Always keep the sampler tube in particle free water when not in use.
Don’t change the set parameters; it can be done only by administrator (if required).
First switch ON main and later switch ON the instrument main only.
Frequently change the user password for the security of the system.
Cleaning:
Clean the syringes weekly using either the detergent/bleach process or the acid/base process.
Clean the capillary using the sensor capillary cleaning kit and a standard glassware cleaning solution, when the DC light reading reaches 7.0.
Maintenance:
Cleaning of the syringe and capillary should be performed regularly.
Do not run syringe more than a few cycles without fluid in them.
Always flush the syringe with distilled or deionized water after use.
If the plunger is removed from the syringe barrel, it should be wiped with alcohol before replacing it.
Do not operate the instrument in an environment affected by excessive Electromagnetic Interference.
Calibration frequency: Calibration should be done yearly by an outside party.
Note: In report format, the first name indicates the person involved in the operation of the instrument and the second name indicates the person who is verifying the report results.
Reconstituted Volume
| Strength | Vials Quantity | Reconstituted Volume per Vial |
| 1.5 g | 05 | 15 ml |
| 1.0 g | 05 | 9.6 ml |
| 750 mg | 05 | 8.3 ml |
| 500 mg | 10 | 4.8 ml |
| 250 mg | 15 | 2.4 ml |
| 06 g | 10 | 26ml |
