Microbubbles manufactured using a tissue homogeniser: DoE-guided optimisation for a narrower size distribution and...

Area of Science:

• Biomaterials Science
• Ultrasound Technology
• Nanotechnology

Background:

• Microbubbles are clinically used as ultrasound contrast agents and are being explored for therapeutic uses.
• Predictable performance of microbubbles relies on a narrow size distribution, which is difficult to achieve with simple production methods.
• Phospholipid-coated microbubbles are essential for these applications.

Purpose of the Study:

• To evaluate a bead-type tissue homogeniser as an alternative to probe sonication for producing phospholipid-coated microbubbles.
• To identify influential parameters for microbubble generation using design of experiments (DoE).
• To compare the characteristics and production efficiency of microbubbles generated by homogenisation versus sonication.

Main Methods:

• A bead-type tissue homogeniser was used to generate DSPC-PEG40S (9:1), air-filled microbubbles.
• Design of Experiments (DoE) with a three-level full-factorial design assessed the effects of speed, time, and volume.
• Optical microscopy with ImageJ analysis quantified size and concentration; passive cavitation detection characterised acoustic response.

Main Results:

• Liquid volume significantly affected mean diameter, concentration, and polydispersity index (PDI); homogenisation speed primarily influenced PDI.
• Optimised settings (5 µm·s⁻¹, 45 s, 500 µl) yielded smaller, more uniform, and higher concentration microbubbles than sonication.
• Homogenisation produced less heat, had higher production rates, and resulted in improved short-term stability (above 10⁸ MB/ml at 6h at 37°C).
• Acoustic emissions were comparable to sonication when normalised to gas volume.

Conclusions:

• Bead homogenisation is a simple, less thermogenic, high-throughput method for reproducible phospholipid microbubble production.
• This method offers advantages over probe sonication in terms of efficiency, uniformity, and stability.
• The DoE approach effectively identified optimal parameters for microbubble generation.