Scientific Papers

Laouini, A, Charcosset, C, Fessi, H, Holdich, RG, Vladisavljevic, GT (2013) Preparation of liposomes: a novel application of microengineered membranes: From laboratory scale to large scale, Colloids and Surfaces B: Biointerfaces, 112, pp.272-278, DOI: 10.1016/j.colsurfb.2013.07.066.

A novel ethanol injection method using microengineered nickel membrane was employed to produce POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) and Lipoid® E80 liposomes at different production scales. A stirred cell device was used to produce 73 ml of the liposomal suspension and the product volume was then increased by a factor of 8 at the same transmembrane flux (140 l m−2 h−1), volume ratio of the aqueous to organic phase (4.5) and peak shear stress on the membrane surface (2.7 Pa). Two different strategies for shear control on the membrane surface have been used in the scaled-up versions of the process: a cross flow recirculation of the aqueous phase across the membrane surface and low frequency oscillation of the membrane surface (∼40 Hz) in a direction normal to the flow of the injected organic phase. Using the same membrane with a pore size of 5 μm and pore spacing of 200 μm in all devices, the size of the POPC liposomes produced in all three membrane systems was highly consistent (80–86 nm) and the coefficient of variation ranged between 26 and 36%. The smallest and most uniform liposomal nanoparticles were produced in a novel oscillating membrane system. The mean vesicle size increased with increasing the pore size of the membrane and the injection time. An increase in the vesicle size over time was caused by deposition of newly formed phospholipid fragments onto the surface of the vesicles already formed in the suspension and this increase was most pronounced for the cross flow system, due to long recirculation time. The final vesicle size in all membrane systems was suitable for their use as drug carriers in pharmaceutical formulations.

Dave Palmer, Alex Kerr, Sam Trotter & Dai Hayward​

Since their discovery in 1965, by Alec D. Bangham, liposomes have been recognised as the drug delivery vehicle of choice. Their biocompatibility results in minimal adverse reactions. Their amphiphilic structure allows encapsulation of both hydrophilic and hydrophobic active pharmaceutical ingredients (APIs). More recently the liposome’s analogous cousin, the lipid nanoparticle, has gained prominence because of its ability to deliver therapeutic payloads, including DNA and mRNA for vaccines. They can both deliver their payload very precisely through treating their surface with proteins allowing highly specific binding to a target cell type. This paper describes the advantages, over the most common commercial processes, that Micropore's membrane emulsification technology has in Liposomes and Lipid nanoparticle production.

Grace Walden, Xin Liao, Graham Riley, Simon Donell, Michael J. Raxworthy, and Aram Saeed

Dave Palmer, Alex Kerr, Sam Trotter & Dai Hayward​

Phase change materials (PCMs) allow the storage of large amounts of latent heat during phase transition. They have the potential to both increase the efficiency of renewable energies such as solar power through storage of excess energy, and to reduce overall energy demand through passive thermal regulation. NASA has identified more than a hundred of these materials. In addition to passive energy storage, they have application in thermo-regulated fabrics, high power electronics, telecommunication installations and microprocessors. PCMs are not suitable for use without prior encapsulation. Encapsulation in a shell material provides benefits including protection of the PCM from the external environment and increased specific surface area to improve heat transfer.

Joanna Wolska and Nasim Jalilnejad Falizi

The membrane emulsification process (ME) using a metallic membrane was the first stage for preparing a spherical and monodisperse thermoresponsive molecularly imprinted polymer (TSMIP). In the second step of the preparation, after the ME process, the emulsion of monomers was then polymerized. Additionally, the synthesized TSMIP was fabricated using as a functional monomer N-isopropylacrylamide, which is thermosensitive. This special type of polymer was obtained for the recognition and determination of trace bisphenol A (BPA) in aqueous media. Two types of molecularly imprinted polymers (MIPs) were synthesized using amounts of BPA of 5 wt.%(MIP-2) and 7 wt.% (MIP-1) in the reaction mixtures. Additionally, a non-imprinted polymer (NIP) was also synthesized. Polymer MIP-2 showed thermocontrolled recognition for imprinted molecules and a higher binding capacity than its corresponding non-imprinted polymer and higher than other molecularly imprinted polymer (MIP-1). The best condition for the sorption process was at a temperature of 35 ◦C, that is, at a temperature close to the phase transition value for poly(N-isopropylacrylamide). Under these conditions, the highest levels of BPA removal from water were achieved and the highest adsorption capacity of MIP-2 was about 0.5 mmol g−1 (about 114.1 mg g−1) and was approximately 20% higher than for MIP-1 and NIP. It was also observed that during the kinetic studies, under these temperature conditions, MIP-2 sorbed BPA faster and with greater efficiency than its non-imprinted analogue.

Jennifer Huen PhD, Beagle Scientific Inc.

When the first liposome-based drug, Doxil®, was approved in the mid-1990s, decades of research spurred the need for more suitable lipid formulations with faster, better controlled and more efficient manufacturing methods. Most recently, the urgency of the COVID19 pandemic pushed the scientific community for larger scale production of the mRNA vaccines with greater consistency—a need that is still evolving. Each of the current methods for industrial production of lipid nanoparticle (LNP) drugs have their unique set of pros and cons. Most liposome drugs are produced by lipid hydration and extrusion, a bulk method that is not suitable for consistent production of LNPs small enough to penetrate tissues for cell entry. From a formulation discovery perspective, microfluidic mixers can quickly produce large LNP libraries while minimizing waste. Yet these mixers cannot accommodate commercial-scale production volumes. Impingement jet mixing (IJM) is the most widespread manufacturing method as it can support high yields through unit parallelization but is less controllable and can potentially compromise LNP stability. Finally, advanced crossflow is a scalable, high efficiency method that has the potential to meet the demands of global disease emergencies. While IJM is currently the method of choice for mRNA vaccine manufacturers, other strategies must not yet be ruled out. As the development of new drug modalities and innovations accelerates, so too must strategies for faster, scalable drug manufacturing.

Karina C. Scheiner,† Roel F. Maas-Bakker,† Thanh T. Nguyen,‡ Ana M. Duarte,‡ Gert Hendriks,‡
Lídia Sequeira,‡ Garry P. Duffy,§ Rob Steendam,‡ Wim E. Hennink,† and Robbert J. Kok*,†
†Department of Pharmaceutics, Utrecht Institute of Pharmaceutical Sciences, Utrecht University, Universiteitsweg 99, 3584 CG
Utrecht, The Netherlands
‡InnoCore Pharmaceuticals B.V., L.J. Zielstraweg 1, 9713 GX Groningen, The Netherlands
§Discipline of Anatomy, School of Medicine, National University of Ireland Galway, University Road, H91 TK33 Galway, Ireland

Vascular endothelial growth factor (VEGF) is the major regulating factor for the formation of new blood vessels, also known as angiogenesis. VEGF is often incorporated in synthetic scaffolds to promote vascularization and to enhance the survival of cells that have been seeded in these devices. Such applications require sustained local delivery of VEGF of around 4 weeks for stable blood vessel formation. Most delivery systems for VEGF only provide short-term release for a couple of days, followed by a release phase with very low VEGF release. We now have developed VEGF-loaded polymeric microspheres that provide sustained release of bioactive VEGF for 4 weeks. Blends of two swellable poly(ε-caprolactone)−poly(ethylene glycol)−poly(ε-caprolactone)-b-poly(L-lactide) ([PCL−PEG−PCL]-b-[PLLA])-based multiblock copolymers with different PEG content and PEG molecular weight were used to prepare the microspheres.
Loading of the microspheres was established by a solvent evaporation-based membrane emulsification method. The resulting VEGF-loaded microspheres had average sizes of 40−50 μm and a narrow size distribution. Optimized formulations of a 50:50 blend of the two multiblock copolymers had an average VEGF loading of 0.79 ± 0.09%, representing a high average VEGF loading efficiency of 78 ± 16%. These microspheres released VEGF continuously over 4 weeks in phosphate-buffered saline pH
7.4 at 37 °C. This release profile was preserved after repeated and long-term storage at −20 °C for up to 9 months, thereby demonstrating excellent storage stability. VEGF release was governed by diffusion through the water-filled polymer matrix, depending on PEG molecular weight and PEG content of the polymers. The bioactivity of the released VEGF was retained within the experimental error in the 4-week release window, as demonstrated using a human umbilical vein endothelial cells proliferation assay. Thus, the microspheres prepared in this study are suitable for embedment in polymeric scaffolds with the aim of promoting their functional vascularization.

Hanga, MP and Holdich, RG (2014) Membrane emulsification for the production of uniform poly-N-isopropylacrylamide-coated alginate particles using internal gelation, Chemical Engineering Research and Design, 92(9), pp.1664-1673, DOI: 10.1016/j.cherd.2013.12.010.

Alginate particles, crosslinked by calcium ions, have a number of potential biopharmaceutical industry applications due to the biocompatibility of the materials used and formed. One such use is as microcarriers for cell attachment, growth and then detachment without the use of proteolytic enzymes. A straightforward and reproducible method for producing uniform calcium alginate particles with controllable median diameters which employs membrane emulsification and internal gelation (solid particles contained in the dispersed phase) is demonstrated, as well as functionalisation of the resulting beads with amine terminated poly N-isopropylacrylamide (pNIPAM) to form temperature responsive particles, by taking advantage of the electrostatic interaction between the carboxyl groups of the alginate and amino groups of the modified pNIPAM. Cell attachment, growth and detachment capabilities of these core–shell structures were assessed and successfully demonstrated by using phase contrast microscopy and fluorescent staining with calcein-AM and ethidium homodimer-1.

The formulation used for the alginate particles avoided non-GRAS chemicals by only using food grade and pharmaceutical grade reagents. The median particle size was controllable within the range between 55 μm and 690 μm and the size distributions produced were very narrow: ‘span’ values as low as 0.2. When using a membrane pore size of 20 μm no membrane blockage by the suspended calcium carbonate necessary for internal gelation of the alginate particles was observed. Membrane pore openings with diameters of 5 and 10 μm were also tested, but blocked with the 2.3 μm median diameter calcium carbonate solids.

Dave Palmer, Alex Kerr, Sam Trotter & Dai Hayward​

The term coacervation derives from the Latin verb “coacervare”, meaning “to crowd together”. The technique of coacervation was first characterised by Bungenberg de Jong in 1931, although the earliest reports of this technique go back to Tiebackx in 1911. Over the last 2-3 decades complex coacervation has been deployed in industries as diverse as food, cosmetics, agriculture and functional materials, as well as, more recently, generating an increasing interest in the pharmaceutical industry as a drug delivery mechanism. This White Paper focuses on drug delivery using complex coacervation. Achieving an accurate target capsule size in an industrial setting can be a challenge While the homogeniser is running, samples are taken and sized via electrozone sensing (Coulter principle) or laser diffraction. These techniques take time to run, and in the meantime the homogeniser continues to reduce the size of the emulsion droplets in the batch. This makes accurate sizing unpredictable.A preferred approach is a system where the desired size characteristics can be defined in advance and the emulsion produced in a single pass. Membrane emulsification makes this possible, by injecting the internal phase through the membrane pores and, by applying a known shear force, droplet sizes can be controlled precisely.