Abstract:The purpose of this study was to introduce dielectric spectroscopy and biocalorimetry as monitoring methods to follow immobilised Chinese Hamster Ovary (CHO) cell culture development. The theory behind both monitoring techniques is explained and perfusion cultures are performed in a Reaction Calorimeter (eRC1 from Mettler Toledo) as an application example. The findings of this work show that dielectric spectroscopy gives highly reliable information upon the viable cell density throughout the entire culture. On the other hand, the RC1 could only provide accurate data from day 5, when the cell density exceeded 4 × 10 , a viable cell density commonly achieved in fed-batch and the early stages of a perfusion culture. This work suggests that biocalorimetry should be possible to implement at industrial scale to monitor CHO cell cultures.

Microcapsules for high cell density culture of mammalian cells have found an increasing interest, however, the poor stability of the microcapsules and the lack of characterisation methods led to few quantitative results. Alginate-poly-L-lysine (PLL) microcapsules have been studied in detail in order to form a basis for comparison of capsules made from different polymers. Since the microcapsules can be easily retained in the bioreactor without the need for a cell separation device, high cell densities were achieved with a maximum of 4 × 10(7) cell/ml(microcapsules), corresponding to a colonisation of 5% of the internal capsule volume. Measurement of microcapsule integrity and mechanical resistance showed that alginate-PLL microcapsules are not suitable for perfusion cultures since they are very sensitive to media composition, mainly the presence of non-gelling ions that have a higher affinity for alginate than PLL and Ca(2+), leading to the leakage of PLL and Ca(2+), and to microcapsule rupture.

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Given the limited number of materials available to design delivery platforms for nutrients, the rational combination of raw materials already approved as food ingredients and their processing through nano-micro technology can offer a unique tool for innovation. Here, we propose a nano-in-micro strategy to produce powders based on the hydrophobic protein zein, useful for the oral delivery of a hydrophilic iron source (iron bisglycinate) in anaemic patients. Iron-loaded powders were prepared through a two-step strategy consisting in the formation of a zein pseudolatex followed by a spray-drying step. To extend the manipulation space for zein and entrap iron bisglycinate, β-cyclodextrin (βCD) was selected as helping excipient. Addition of βCD allowed iron loading in the pseudolatex and greatly increased product yields after the drying process as compared to zein alone. Iron-loaded micro-sized powders were characterised by attenuated total reflectance–Fourier transform infrared (ATR-FTIR) spectra, thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC) to elucidate the role of βCD as a compatibilizer for the zein–iron system. Remarkably, micropowders released only 20% of FeBIS in a simulated gastric fluid, whereas release in a simulated intestinal fluid was almost completed in 7 h. In summary, βCD association to zein is a novel strategy to expand applications in the oral delivery of iron bisglycinate and, prospectively, to micronutrient chelates.

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