Standard Operating Procedure for Zeta Potential Determination in Analytical Method Development
| Department | Analytical Method Development |
|---|---|
| SOP No. | SOP/AMD/117/2025 |
| Supersedes | SOP/AMD/117/2022 |
| Page No. | Page 1 of 14 |
| Issue Date | 19/05/2025 |
| Effective Date | 20/05/2025 |
| Review Date | 19/05/2026 |
1. Purpose
This SOP defines the method for measuring zeta potential using electrophoretic light scattering (ELS) for pharmaceutical dispersions, nanoparticles, and colloidal systems in order to assess their physical stability and surface charge properties.
2. Scope
This procedure applies to the Analytical Method Development (AMD) laboratory for the characterization of formulations, APIs, and nanoformulations requiring evaluation of zeta potential in both research and product development stages.
3. Responsibilities
- Analytical Scientist: Prepares samples, conducts measurements, and analyzes data.
- Instrument Operator: Calibrates equipment and ensures maintenance logs are updated.
- QA Officer: Reviews and verifies the records for GMP compliance.
- Head – AMD: Approves method development and interpretation reports.
4. Accountability
The Head of Analytical Method Development is accountable for ensuring method integrity, accuracy, and traceability in zeta potential determination processes.
5. Procedure
5.1 Instrument Preparation
- Switch on the zeta potential analyzer (e.g., Malvern Zetasizer Nano ZS) and allow it to stabilize.
- Verify the cuvette type:
- Disposable folded capillary cells (DTS1070)
- Glass cuvettes (for non-aqueous systems)
- Calibrate instrument as per manufacturer’s instructions with 0.1 mM KCl or latex standard solution.
- Record in Annexure-1: Instrument Calibration Log.
5.2 Sample Preparation
- Prepare the formulation/sample at a concentration ensuring obscuration between 5–10%.
- Use freshly prepared or filtered dispersions to avoid particulate interference.
- If necessary, dilute with 1 mM electrolyte (e.g., KCl, NaCl) to maintain ionic strength.
- Avoid bubbles or precipitates; mix gently to ensure uniformity.
- Document preparation in Annexure-2: Sample Preparation Log.
5.3 Measurement Settings
- Set parameters:
- Measurement type: Zeta Potential
- Medium: Water or selected solvent
- Temperature: 25°C (unless otherwise justified)
- Voltage: As per instrument auto-adjust or manual setting (30–150 V)
- Run duration: 10–20 cycles per sample
- Load the sample into the cuvette and place in the holder.
- Start measurement and monitor electrophoretic mobility values.
- Software will auto-calculate zeta potential via Henry’s equation.
- Repeat triplicate readings and report average with standard deviation.
- Record results in Annexure-3: Zeta Potential Data Sheet.
5.4 Data Analysis and Interpretation
- Evaluate mean zeta potential and standard deviation across triplicates.
- Acceptable RSD: ≤ 10% across replicates.
- Typical interpretation:
- > +30 mV or < -30 mV: High stability
- ±20–30 mV: Moderate stability
- ±10–20 mV: Low stability
- < ±10 mV: Likely to aggregate
- Include summary in Annexure-4: Interpretation and Observation Sheet.
5.5 Cleaning and Post-Use Protocol
- Flush cuvettes with deionized water or ethanol (non-aqueous samples) after use.
- Store reusable cuvettes in dust-free containers.
- Log cleaning activities in Annexure-5: Post-Use Maintenance Record.
6. Abbreviations
- ELS: Electrophoretic Light Scattering
- RSD: Relative Standard Deviation
- mV: Millivolts
- SOP: Standard Operating Procedure
7. Documents
- Instrument Calibration Log – Annexure-1
- Sample Preparation Log – Annexure-2
- Zeta Potential Data Sheet – Annexure-3
- Interpretation and Observation Sheet – Annexure-4
- Post-Use Maintenance Record – Annexure-5
8. References
- USP General Chapter <790>: Zeta Potential Determination
- ICH Q6A – Specifications for New Drug Substances
- Instrument User Manual – Zetasizer Nano Series
9. SOP Version
Version: 2.0
10. Approval Section
| Prepared By | Checked By | Approved By | |
|---|---|---|---|
| Signature | |||
| Date | |||
| Name | |||
| Designation | |||
| Department |
11. Annexures
Annexure-1: Instrument Calibration Log
| Date | Standard Used | Expected Value | Measured Value | Status |
|---|---|---|---|---|
| 18/05/2025 | Latex standard | -42 mV | -41.8 mV | Pass |
Annexure-2: Sample Preparation Log
| Sample ID | Formulation | Diluent | Concentration | Prepared By |
|---|---|---|---|---|
| ZP-117-A | API-Nano Dispersion | 1 mM KCl | 0.5 mg/mL | Rajesh Kumar |
Annexure-3: Zeta Potential Data Sheet
| Trial | Zeta Potential (mV) | Remarks |
|---|---|---|
| 1 | -31.2 | Stable |
| 2 | -30.7 | Stable |
| 3 | -31.5 | Stable |
Annexure-4: Interpretation and Observation Sheet
Average Zeta Potential: -31.1 mV; RSD: 1.3%. Indicates good colloidal stability and electrostatic repulsion adequate to prevent aggregation.
Annexure-5: Post-Use Maintenance Record
| Date | Cuvette Type | Cleaning Agent | Cleaned By | Verified By |
|---|---|---|---|---|
| 18/05/2025 | Disposable (DTS1070) | DI Water | Sunita Reddy | QA |
Revision History:
| Revision Date | Revision No. | Details | Reason | Approved By |
|---|---|---|---|---|
| 04/05/2025 | 2.0 | Expanded interpretation ranges and added robustness in cleaning protocol | Annual Review |