CHO immobilization in alginate/poly- l -lysine microcapsules: an understanding of potential and limitations

Microencapsulation offers a unique potential for high cell density, high productivity mammalian cell cultures. However, for successful exploitation there is the need for microcapsules of defined size, properties and mechanical stability. Four types of alginate/poly-l-Lysine microcapsules, containing recombinant CHO cells, have been investigated: (a) 800μm liquid core microcapsules, (b) 500μm liquid core microcapsules, (c) 880μm liquid core microcapsules with a double PLL membrane and (d) 740μm semi-liquid core microcapsules. With encapsulated cells a reduced growth rate was observed, however this was accompanied by a 2-3 fold higher specific production rate of the recombinant protein. Interestingly, the maximal intracapsular cell concentration was only 8.7×107cell mL-1, corresponding to a colonization of 20% of the microcapsule volume. The low level of colonization is unlikely to be due to diffusional limitations since reduction of microcapsule size had no effect. Measurement of cell leaching and mechanical properties showed that liquid core microcapsules are not suitable for continuous long-term cultures (>1month). By contrast semi-liquid core microcapsules were stable over long periods with a constant level of cell colonization (ϕ=3%). This indicates that the alginate in the core plays a predominant role in determining the level of microcapsule colonization. This was confirmed by experiments showing reduced growth rates of batch suspension cultures of CHO cells in medium containing dissolved alginate. Removal of this alginate would therefore be expected to increase microcapsule colonizatio

Alginate-based microcapsules for mammalian cell culture and other biotechnological applications

Since life began, nature has been using the envelopment of systems for their protection or for providing a particular reaction space, the enveloping wall displaying additional specific membrane functions. The abundance of examples is quite overwhelming and extends from arboreal fruits and plant seeds to plant seed spores, from the hen egg and its shell to the cell and its membrane/wall. The technique of microencapsulation aims at immobilizing gases, liquids or solids in an envelope. The result is a core contained in a capsule ranging from the nanometer to the millimeter scale. Nowadays, microencapsulation is being studied and developed in many different backgrounds throughout the chemical and life sciences, biotechnology, medicine and related industries. The aims of this thesis were to study the potentials and limitations of alginate-based microcapsules, for biotechnological applications, especially mammalian cell culture. It was shown that the limitations of alginate poly-L-lysine (PLL) microcapsules, commonly used for mammalian cell entrapment, arises from the poor mechanical stability of the polyelectrolytic complex constituting the membrane, and the intracapsular alginate concentration preventing optimal cell growth and core colonization. These limitations may be overcome by finding new techniques preventing the accumulation of alginate in the core, and by reinforcing the membrane through the formation of covalent bonds. To remove alginate from the core of the capsule, the use of an alginate degrading enzyme might be a possibility. However, alginate lyase degrades both gelled, dissolved and complexed forms of alginate. Therefore, this system should be more appropriated for cell release from microcapsules. Another alternative for the formation of alginate-free core microcapsules may be the hydrolysis of an organic-core capsule by a lipase. Aqueous-core capsules surrounded by an alginate matrix were successfully obtained, without accumulation of alginate in the core. However, this system is not suitable for mammalian cell encapsulation due to the toxicity of the process, especially the degradation product resulting from the enzymatic hydrolysis. The third solution to avoid the presence of alginate in the core was the use of the alginate/casein aqueous two-phase system. The combination of these compounds was suitable for the formation of aqueous-core microcapsules, using the jet break-up technique with a concentric nozzle. Casein was not toxic to cells, however precipitated in the presence of Ca2+ (dissolved in the buffer to induce alginate gelation), and hindered further cell growth. Membrane stability can be reinforced by the formation of covalent bonds. Acrylamide monomers polymerization and PLL / propylene-glycol-alginate / BSA transacylation reaction were the two systems studied. The formation of covalent bonds was successful for both systems, and produced reinforced capsules. However, acrylamide monomer, as well as the polymerization process, were toxic for the mammalian cells. Despite the harsh reaction conditions (pH 11), the transacylation reaction allowed cell growth and produced very resistant and stable capsules. As a perspective, the most promising technique seems to be the use of aqueous two-phase systems. The biocompatible nature of the molecules constituting the microcapsules is a great advantage, giving rise to promising issues. Moreover, the possibility to induce polymerization and cross-linking reactions in the membrane is also an advantage of this new type of microcapsules. Due to the harsh condition of reaction, acrylamide or transacylation reactions are probably not the optimal systems, however other covalent systems (genipin or poly(ethylene glycol)-based hydrogels) might be a promising alternative. Encapsulation applications are numerous, and are not only restricted to cell immobilization. The study of the immobilization of rapeseed press-cake in an alginate matrix is an example of a new potential application of alginate-based microcapsules / beads. It was shown to be suitable for removing organic pollutants from waste water, simplifying the process (i.e. phase separation), thus allowing to envisage large-scale industrial applications

Interdisciplinary projects applied to chemistry lessons ::example of chemistry linked to food science

An interdisciplinary project regarding the effect of ascorbic acid on bread dough’s physico-chemical properties was proposed to bachelor students in chemistry and food sciences. Such an approach was proposed to develop both scientific and soft skills, in order to prepare students for their future working environment. Together, students deepened their knowledge regarding food science and chemistry. They were then able to plan and design experiments demonstrating the impact of gluten network formation and ascorbic acid influence onto bread dough and finished product characteristics. This way of teaching was very appreciated by students, nevertheless it highlighted the fact that the professors’ investment was considerably high, and that good organization, alignment and preparation prior to the start of this project is key

125th anniversary of the School of Engineering and Architecture of Fribourg (HEIA-FR)

This year, on the 14th of January 2021 the School of Engineering and Architecture (HEIA-FR)[1] celebrated its 125th anniversary. To celebrate, many interactive events are organized from February to September 2021

125th Anniversary of the School of Engineering and Architecture of Fribourg (HEIA-FR): Chimia Report/Company News

This year, on the 14th of January 2021 the School of Engineering and Architecture (HEIA-FR) celebrated its 125th anniversary. To celebrate, many interactive events are organized from February to September 2021

Development of an integrated solution to prevent spring frost damage using an aqueous-based insulating foam

In recent years, agricultural crops experience unusually early onset of vegetation due to global warming, which can cause major frost damage with devastating effects on crop yields. To mitigate the risk of frost damage, an integrated solution was developed, consisting of an aqueous-based biocompatible foam and a portable foam applicator enabling wine cultivators to treat up to 1000 m2 of vineyards with one filling containing 10 L of foam. The foam is biocompatible, stable for several days and easily removed by rain. Foam application yielded an insulation efficiency of up to 1.5°C during spring frost nights for the buds covered by the foam when combined with an electrically heated wire. Moreover, it was observed that the foam also created a 'mini greenhouse' effect at positive temperatures during the days, which might be a positive side effect helping the plants to grow at this early stage of the year

Distance teaching in chemistry ::opportunities and limitations

Remote teaching in the tertiary education sector is a relatively common practice, and the implementation of digital solutions in chemistry teaching offers many new opportunities and tools. A survey was conducted after 3 months of emergency remote teaching linked to the COVID-19 pandemic and showed that half of the students estimated it was difficult to study remotely, and reported they had to invest more time compared to classroom teaching, which led to a drop in motivation. Professors also noted that the time necessary to invest in order to produce digital teaching content was enormous. Massive open online laboratories (MOOLs) and process simulators are interesting tools, but practical lab work and related know-how cannot fully be replaced by digital techniques. Finally, it appeared that the professor–student interaction is very important in the distance-learning process, and that a high level of pedagogical (inter)activity is mandatory to maintain motivation and better quality of teaching and learning

CHO immobilization in alginate/poly-l-lysine microcapsules: an understanding of potential and limitations

Microencapsulation offers a unique potential for high cell density, high productivity mammalian cell cultures. However, for successful exploitation there is the need for microcapsules of defined size, properties and mechanical stability. Four types of alginate/poly-l-Lysine microcapsules, containing recombinant CHO cells, have been investigated: (a) 800 μm liquid core microcapsules, (b) 500 μm liquid core microcapsules, (c) 880 μm liquid core microcapsules with a double PLL membrane and (d) 740 μm semi-liquid core microcapsules. With encapsulated cells a reduced growth rate was observed, however this was accompanied by a 2–3 fold higher specific production rate of the recombinant protein. Interestingly, the maximal intracapsular cell concentration was only 8.7 × 107 cell mL-1, corresponding to a colonization of 20% of the microcapsule volume. The low level of colonization is unlikely to be due to diffusional limitations since reduction of microcapsule size had no effect. Measurement of cell leaching and mechanical properties showed that liquid core microcapsules are not suitable for continuous long-term cultures (>1 month). By contrast semi-liquid core microcapsules were stable over long periods with a constant level of cell colonization (ϕ = 3%). This indicates that the alginate in the core plays a predominant role in determining the level of microcapsule colonization. This was confirmed by experiments showing reduced growth rates of batch suspension cultures of CHO cells in medium containing dissolved alginate. Removal of this alginate would therefore be expected to increase microcapsule colonization