Standard Operating Procedure for DO Cascade Settings in Bioreactor Operations for Biosimilars

Department	Biosimilars
SOP No.	SOP/BS/077/2025
Supersedes	SOP/BS/077/2022
Page No.	Page 1 of 12
Issue Date	04/05/2025
Effective Date	06/05/2025
Review Date	04/05/2026

1. Purpose

To define the procedure for configuring, verifying, and managing Dissolved Oxygen (DO) cascade settings in bioreactor operations to ensure optimal cell growth conditions during biosimilar manufacturing.

2. Scope

This SOP is applicable to all bioreactor systems used for mammalian cell culture in upstream processes for biosimilar production where automated oxygen control via cascade logic is implemented.

3. Responsibilities

• Bioreactor Operators: Set and monitor DO cascade parameters during fermentation runs.
• Automation/Engineering Team: Maintain PLC control logic and validate DO cascade software configurations.
• QA Department: Review and approve DO cascade setting records and change controls.

4. Accountability

The Head of Upstream Manufacturing is accountable for ensuring correct implementation of DO cascade systems as per validated process requirements.

5. Procedure

5.1 Sensor Calibration

1. Calibrate DO sensors before every production batch using two-point calibration:
   • 0% (using nitrogen-purged solution)
   • 100% (using air-saturated water)
2. Document calibration results in the DO Sensor Calibration Log (Annexure-1).

5.2 Cascade Parameter Configuration

1. Access the control system HMI or SCADA interface and navigate to the DO cascade menu.
2. Set the DO setpoint to 30% or as per batch record.
3. Configure cascade steps in the following sequence:
   1. Step 1: Increase agitation from 100 to 300 RPM.
   2. Step 2: Increase air flow rate from 0.1 to 1.0 vvm.
   3. Step 3: Introduce pure oxygen blending if DO remains below setpoint.
4. Enable PID control loop and confirm tuning parameters are as validated.

5.3 Cascade Verification

1. Simulate low DO by manually increasing cell density in test mode or using nitrogen sparging.
2. Verify automatic activation of each cascade step in correct order.
3. Ensure system returns to baseline once DO reaches setpoint.
4. Record cascade test results in Annexure-2.

5.4 Monitoring During Batch Run

1. Monitor DO values continuously during the culture phase.
2. Ensure cascade is active and system is responding to DO fluctuations.
3. Log agitation, airflow, and oxygen blending trends every 4 hours (Annexure-3).

5.5 Deviation Handling

1. If DO fails to recover despite cascade, escalate to engineering and shift to manual control.
2. Log deviation and corrective actions in deviation report.

6. Abbreviations

• DO: Dissolved Oxygen
• HMI: Human Machine Interface
• SCADA: Supervisory Control and Data Acquisition
• vvm: Volume of air per volume of medium per minute

7. Documents

1. DO Sensor Calibration Log – Annexure-1
2. DO Cascade Verification Checklist – Annexure-2
3. DO Cascade Monitoring Log – Annexure-3

8. References

• ICH Q8 – Pharmaceutical Development
• WHO TRS 1010 – Bioprocess Equipment Control

9. SOP Version

Version: 2.0

10. Approval Section

	Prepared By	Checked By	Approved By
Signature
Date
Name
Designation
Department

11. Annexures

Annexure-1: DO Sensor Calibration Log

Date	Bioreactor ID	0% Calibration (mV)	100% Calibration (mV)	Calibrated By
04/05/2025	BR-12	12.6	198.4	Rajesh Kumar

Annexure-2: DO Cascade Verification Checklist

Step	Parameter	Status	Tested By	Date
1	Agitation Increase	Pass	Sunita Reddy	04/05/2025
2	Air Flow Increase	Pass	Sunita Reddy	04/05/2025
3	Oxygen Blending	Pass	Sunita Reddy	04/05/2025

Annexure-3: DO Cascade Monitoring Log

Date	Time	DO (%)	Agitation (RPM)	Air Flow (vvm)	O2 Blending (%)	Operator
04/05/2025	12:00	30.1	220	0.6	5	Ajay Verma

Revision History:

Revision Date	Revision No.	Revision Details	Reason for Revision	Approved By
04/05/2025	2.0	Updated control logic, cascade sequence, and annexures	Process Automation Enhancement