Category: Manufacturing and Bioprocessing
Purpose: A critical stage in the manufacture of a heat-labile parenteral drug is the sterilizing filtration step prior to filling of the drug product into the final dosage container or delivery system. Increasingly, biocompatible nanosuspensions are being considered and implemented in formulations to encapsulate drug molecules. They can be used to overcome solubility challenges, allowing for extended release and improved targeting of molecules at their intended sites of action.
The liposomal delivery method can be costly and presents filtration challenges, and consequently this presents opportunities to better understand the effects of filtering liposomes, including what test conditions, such as filtration pressure, will influence overall filtration behaviour. To evaluate the effect of the above conditions on filtration behaviour an investigation was performed where multiple sterilizing grade filters were challenged with fluorescent dye-encapsulated liposome solution.
Confocal microscopy was used to measure the impact of each test and filter morphology on liposome filterability. This was a key technique in this investigation, providing a method to track and differentiate between fluorescent particles and a method to analyse each challenged membrane for adsorption and penetration of liposome. This type of analysis can be used to provide useful insights when identifying a suitable filter for the sterilization of final drug product using liposomes.
Methods: Membranes were examined using the Leica SPE inverted Confocal Microscope (Leica Microsystems) and images captured using the xyz capabilities of the instrument.
Phosphatidylcholine lipoid was used in representing the liposome drug delivery system, and was used to encase the liposome tag TopFluor PC to provide fluorescence potential. The liposomes were incorporated with the fluorescent tag on the M-110EH Microfluidizer processor (Microfluidics International)
Liposome formulation properties are analysed prior to use as characteristics such as size or viscosity can affect filtration capacity.
Results: Images obtained on the Confocal Microscope, combined with normalised curves, concur with liposome throughput data obtained from filtration analysis across different membranes. Confocal Microscopy can be used to compare membrane layer differences of liposome capacity, demonstrating membranes with wider distribution of particles utilise more depth in filtration
Trends for liposome retention were similar across the membrane regardless of pressure used in processing. This suggests that Confocal Microscopy can be used to visualise the retention of liposome particles, the method is currently unable to differentiate between pressures to explain the increase in capacity
Conclusion: Characterisation results for DLS and TRPS of both liposome and fluorescent liposome solutions showed that fluorescent liposome was successfully manufactured using the Microfluidizer.
Using fluorescent liposome and confocal microscopy provides a method to assess liposome capacity, and can be used to explain why different membranes have higher performance than others
Further optimisation may be required to enable observation of the fine differences between different processing parameters to further understand the mechanisms behind the impact of pressure on filtration of liposome