SOP for Preparation of Aqueous Phase for Liposomal Formulations

Aqueous Phase Preparation for Liposomal Formulations

1) Purpose

This SOP provides guidelines for the preparation of the aqueous phase used in liposomal formulations. The aqueous phase is a critical component in liposome production, as it serves as the medium for the encapsulation of water-soluble drugs and other active pharmaceutical ingredients (APIs). The aqueous phase must be prepared with accuracy to ensure the efficacy and stability of the final liposome product.

2) Scope

This SOP applies to all personnel involved in the preparation of the aqueous phase for liposome formulations in research, development, and commercial production settings. It includes steps for selecting and preparing buffers, adjusting pH, and maintaining sterility during the process.

3) Responsibilities

Operators: Responsible for the accurate preparation of the aqueous phase according to this SOP. Operators must ensure that all equipment is properly cleaned, sterilized, and calibrated before use.
QA Team: Responsible for overseeing the preparation process, ensuring that all steps are carried out in compliance with SOP requirements, and verifying the accuracy of documentation.
QC Team: Responsible for testing the quality of the aqueous phase, including pH, osmolarity, and sterility, before it is used in the liposome formulation process.

4) Procedure

4.1 Equipment Setup

All equipment used in the preparation of the aqueous phase must be thoroughly cleaned, sterilized, and calibrated before starting the process. The following equipment is required:

4.1.1 Required Equipment

pH meter (calibrated)
Analytical balance
Autoclave (for sterilization)
Magnetic stirrer
Glass beakers and volumetric flasks
Filtration setup (for sterile filtration)
Thermometer

4.1.2 Equipment Calibration

Before preparing the aqueous phase, all instruments must be checked for calibration. Calibration must be performed as follows:

4.1.2.1 Calibrate the pH meter using standard pH buffer solutions (e.g., pH 4.0, 7.0, and 10.0).
4.1.2.2 Ensure the analytical balance is calibrated to an accuracy of 0.001 g.
4.1.2.3 Verify that the autoclave is functioning correctly, and perform a spore test if necessary to confirm sterilization efficacy.
4.1.2.4 Record all calibration results in the equipment logbook.

4.2 Preparation of Buffers

Buffers are typically used as the aqueous phase in liposome formulations to maintain the pH and stability of the drug encapsulation environment. Commonly used buffers include phosphate-buffered saline (PBS), citrate buffer, and acetate buffer.

4.2.1 Buffer Selection

4.2.1.1 Select the buffer based on the compatibility of the drug and lipid components. PBS is commonly used for neutral liposomes, while acetate or citrate buffers may be preferred for pH-sensitive formulations.
4.2.1.2 Consult the formulation protocol or batch manufacturing record (BMR) for buffer concentration and composition.

4.2.2 Buffer Preparation

The following steps must be followed to prepare the buffer solution:

4.2.2.1 Weigh the required amount of buffer salts (e.g., sodium chloride, potassium phosphate) using an analytical balance.
4.2.2.2 Dissolve the salts in distilled or deionized water, stirring with a magnetic stirrer to ensure complete dissolution.
4.2.2.3 Measure the volume of the buffer solution using a volumetric flask, ensuring that the final volume is accurate to within ±0.1%.
4.2.2.4 Adjust the pH of the buffer solution to the desired level using a pH meter. Add small amounts of acid (e.g., hydrochloric acid) or base (e.g., sodium hydroxide) to achieve the correct pH.
4.2.2.5 Filter the buffer solution through a 0.22 μm filter to ensure sterility.

4.3 pH Adjustment and Osmolarity Testing

The pH and osmolarity of the aqueous phase must be carefully controlled to ensure compatibility with the lipid components and drug being encapsulated. Deviations in pH or osmolarity can lead to instability of the liposomes or degradation of the active ingredients.

4.3.1 After the buffer is prepared, measure the pH using a calibrated pH meter. If necessary, make small adjustments by adding acid or base, as described in Section 4.2.2.4.
4.3.2 Test the osmolarity of the aqueous phase using an osmometer. The osmolarity should be within the specified range (e.g., 270-320 mOsm/kg) to match physiological conditions and prevent osmotic stress on the liposomes.
4.3.3 Record the pH and osmolarity measurements in the Batch Manufacturing Record (BMR, see Annexure 1).

4.4 Sterilization of the Aqueous Phase

To maintain the sterility of the liposome formulation, the aqueous phase must be sterilized before use. The following methods can be used for sterilization:

4.4.1 Sterile Filtration

4.4.1.1 Pass the prepared buffer solution through a 0.22 μm filter to remove any microorganisms or particulate matter.
4.4.1.2 Ensure that the filtration equipment is sterile and that the filtration process is performed in a laminar flow hood to prevent contamination.
4.4.1.3 Store the filtered buffer in sterilized containers, and label them with the buffer name, concentration, pH, preparation date, and expiration date.

4.4.2 Autoclaving

If sterile filtration is not feasible, autoclaving can be used to sterilize the aqueous phase:

4.4.2.1 Transfer the buffer solution into autoclavable glass containers.
4.4.2.2 Autoclave the buffer solution at 121°C for 20 minutes.
4.4.2.3 After autoclaving, allow the solution to cool to room temperature under sterile conditions.
4.4.2.4 Inspect the containers for cracks or contamination after the autoclaving process. If contamination is detected, discard the buffer and repeat the process.

4.5 Storage of Aqueous Phase

The prepared and sterilized aqueous phase can be stored for future use in liposome formulations. The following storage guidelines must be adhered to:

4.5.1 Store the sterilized buffer solution in a cool, dry place, preferably between 2°C and 8°C, unless otherwise specified in the formulation protocol.
4.5.2 Ensure that all storage containers are labeled with the buffer name, concentration, pH, preparation date, and expiration date.
4.5.3 Record the storage conditions in the Raw Material Storage Log (see Annexure 2).
4.5.4 Conduct regular inspections of the buffer solutions to ensure that there are no signs of contamination or degradation (e.g., color change, precipitation).

5) Abbreviations, if any

PBS: Phosphate-Buffered Saline
BMR: Batch Manufacturing Record
QC: Quality Control

6) Documents, if any

Batch Manufacturing Record (BMR, see Annexure 1)
Raw Material Storage Log (see Annexure 2)
pH Meter Calibration Log
Osmometer Calibration Log

7) References, if any

FDA Guidelines for Sterile Drug Products
ICH Q7: Good Manufacturing Practice Guide for Active Pharmaceutical Ingredients

8) SOP Version

Version 1.0

Annexure

Annexure 1: Batch Manufacturing Record Template


  
    Batch No.
    Buffer
    pH
    Osmolarity
    Filtration Method
    Operator Initials
    QA Signature
  
  
    Batch Number
    Buffer Name
    pH Value
    mOsm/kg
    0.22 μm Filter / Autoclave
    Operator Name
    QA Name

Batch No.	Buffer	pH	Osmolarity	Filtration Method	Operator Initials	QA Signature
Batch Number	Buffer Name	pH Value	mOsm/kg	0.22 μm Filter / Autoclave	Operator Name	QA Name

Annexure 2: Raw Material Storage Log Template


  
    Date
    Buffer Name
    Batch No.
    Storage Condition
    Location
    Operator Initials
  
  
    DD/MM/YYYY
    Buffer Name
    Batch Number
    2-8°C
    Storage Area
    Operator Initials

Date	Buffer Name	Batch No.	Storage Condition	Location	Operator Initials
DD/MM/YYYY	Buffer Name	Batch Number	2-8°C	Storage Area	Operator Initials