4.2 Selection of Lipid Components

The selection of lipids is critical for ensuring that the liposomes are stable at physiological temperatures and release their contents when exposed to hyperthermic conditions. Follow these steps to select the appropriate lipid components:

• 4.2.1 Select phospholipids with a phase transition temperature (Tm) close to the desired release temperature. Dipalmitoylphosphatidylcholine (DPPC), which has a Tm of 41°C, is commonly used in heat-sensitive liposomes.
• 4.2.2 Incorporate cholesterol to improve membrane stability and prevent premature leakage of the API at body temperature (37°C).
• 4.2.3 Choose APIs that remain stable during the preparation process and are effectively released at elevated temperatures.

4.3 Liposome Preparation Process

4.3.1 Preparation of the Lipid Film

• 4.3.1.1 Weigh the required amounts of DPPC, cholesterol, and other lipids as per the formulation protocol. Record the weights in the Batch Manufacturing Record (BMR).
• 4.3.1.2 Dissolve the lipids in an organic solvent such as chloroform in a round-bottom flask.
• 4.3.1.3 Use a rotary evaporator to remove the solvent under reduced pressure, forming a thin lipid film on the walls of the flask.
• 4.3.1.4 Dry the lipid film under vacuum for an additional 30 minutes to remove residual solvent.

4.3.2 Hydration of the Lipid Film

• 4.3.2.1 Prepare an aqueous solution containing the API or buffer solution and heat it to a temperature above the phase transition temperature of the lipids (e.g., 45°C).
• 4.3.2.2 Add the warm aqueous phase to the lipid film and stir gently for 30 minutes to form multilamellar vesicles (MLVs).

4.3.3 Size Reduction and Homogenization

• 4.3.3.1 Pass the MLV suspension through a high-pressure homogenizer to reduce vesicle size and form small unilamellar vesicles (SUVs).
• 4.3.3.2 Optionally, use ultrasonication to further reduce the vesicle size to the desired range (e.g., 100-200 nm).

4.4 Quality Control Testing

After preparing the heat-sensitive liposomes, perform the following quality control tests:

• 4.4.1 Measure the particle size using dynamic light scattering (DLS). The average particle size should typically range between 100-200 nm.
• 4.4.2 Perform a temperature-sensitive release assay by subjecting the liposomes to physiological (37°C) and hyperthermic conditions (42°C) and measuring the release of the API using analytical techniques such as HPLC.
• 4.4.3 Conduct stability testing by storing the liposomes at 4°C, room temperature, and accelerated conditions (e.g., 40°C) to monitor particle size changes and API retention over time.

4.5 Storage of Heat-Sensitive Liposomes

The prepared liposomes should be stored in sterilized, airtight containers. Label each container with the batch number, preparation date, and storage conditions. Store the liposomes at 4°C or as specified in the formulation protocol.

5) Abbreviations, if any

• DPPC: Dipalmitoylphosphatidylcholine
• Tm: Phase Transition Temperature
• MLV: Multilamellar Vesicles
• SUV: Small Unilamellar Vesicles
• QA: Quality Assurance
• QC: Quality Control

6) Documents, if any

• Batch Manufacturing Record (BMR)
• Particle Size Analysis Report
• Temperature-Sensitive Release Report
• Stability Test Report

7) References, if any

• ICH Q8: Pharmaceutical Development Guidelines
• FDA Guidelines for Liposomal Drug Products

8) SOP Version

Version 1.0

Annexure

Annexure 1: Batch Manufacturing Record Template

  

    
Batch No.
    
Lipid Type
    
Weight
    
API Name
    
API Concentration
    
Homogenization Time
    
Operator Initials
    
QA Signature
  
  

    
Batch Number
    
Lipid Name
    
Weight in grams
    
API Name
    
Concentration (mg/mL)
    
Minutes
    
Operator Name
    
QA Name