Standard Operating Procedure for Oxygen Mass Transfer Rate Determination in Biosimilar Manufacturing
| Department | Biosimilars |
|---|---|
| SOP No. | SOP/BS/141/2025 |
| Supersedes | SOP/BS/141/2022 |
| Page No. | Page 1 of 10 |
| Issue Date | 04/05/2025 |
| Effective Date | 06/05/2025 |
| Review Date | 04/05/2026 |
1. Purpose
To establish a validated procedure for determining the oxygen mass transfer rate (kLa) in biosimilar bioreactors to support process optimization, scale-up, and robust control of dissolved oxygen (DO) levels.
2. Scope
This SOP applies to all pilot-scale and production bioreactors used in upstream processing of biosimilars where DO is a critical process parameter. It includes both static and dynamic gassing-out methods for kLa determination.
3. Responsibilities
- Process Development: Design experiments and calculate kLa values for different bioreactor configurations.
- Engineering: Support setup, instrumentation, and parameter adjustments.
- QA: Review validation reports and ensure traceability of all records.
4. Accountability
The Head of Upstream Manufacturing is accountable for ensuring accurate kLa assessments are performed, documented, and incorporated into process development reports and scale-up protocols.
5. Procedure
5.1 Selection of Method
- Use either:
- Dynamic gassing-out method (preferred)
- Sodium sulfite chemical method (alternative)
- Select based on availability of DO sensors, reactor size, and automation capability.
5.2 Dynamic Gassing-Out Method
- Fill bioreactor with WFI or buffer to working volume (no cells).
- Deoxygenate media by sparging nitrogen until DO = 0%.
- Switch to air sparging and record DO values at 10-second intervals until saturation.
- Record airflow rate, agitation RPM, backpressure, and temperature.
- Calculate kLa using:
ln(C* – C) = -kLa × t
Where C = DO concentration at time t, C* = saturated DO level. - Plot data on semi-logarithmic scale and derive slope for kLa.
5.3 Sodium Sulfite Method (if applicable)
- Add 0.5 g/L sodium sulfite + 50 µM cobalt catalyst to WFI in the reactor.
- Start air sparging and monitor oxygen uptake using DO probe or titration.
- Calculate oxygen uptake rate and derive kLa based on oxygen consumption kinetics.
5.4 Documentation and Reporting
- Record all raw readings, sensor calibration data, and calculations in Annexure-1: kLa Determination Log.
- QA to review and archive validation data in equipment qualification file.
6. Abbreviations
- kLa: Oxygen mass transfer coefficient
- DO: Dissolved Oxygen
- WFI: Water for Injection
- QA: Quality Assurance
7. Documents
- kLa Determination Log – Annexure-1
- Sensor Calibration Certificate – Annexure-2
8. References
- ICH Q8 – Pharmaceutical Development
- FDA Guidance on Process Validation: General Principles and Practices
- USP <1046> – Cellular and Tissue-Based Products
9. SOP Version
Version: 2.0
10. Approval Section
| Prepared By | Checked By | Approved By | |
|---|---|---|---|
| Signature | |||
| Date | |||
| Name | |||
| Designation | |||
| Department |
11. Annexures
Annexure-1: kLa Determination Log
| Date | Bioreactor ID | Method | RPM | Airflow | kLa (1/hr) | Performed By |
|---|---|---|---|---|---|---|
| 04/05/2025 | BR-500L | Dynamic | 200 | 0.5 vvm | 45 | Ajay Verma |
Annexure-2: Sensor Calibration Certificate
| Sensor ID | Calibrated On | Calibrated By | Method | Certificate No. |
|---|---|---|---|---|
| DO-SN-038 | 03/05/2025 | Sunita Reddy | Two-Point | CAL/025/2025 |
Revision History:
| Revision Date | Revision No. | Revision Details | Reason for Revision | Approved By |
|---|---|---|---|---|
| 04/05/2025 | 2.0 | Added sodium sulfite method as alternative option | Flexibility for validation |