The purpose of the investigation was the design and development of an oxygen carrier system for oxygenation of liver cells (hepatocytes) in a bio-artificial liver support system. Acute liver failure is a devastating condition with higher than 80% mortality. Currently the only successful treatment is orthotopic liver transplantation. The high mortality rate could be reduced if a system could be developed that could bridge the patient either until recovery (due to the liver’s well-known regeneration ability) or until transplantation. Such a system requires a bioreactor with a high density of cultured cells. Sufficient oxygen delivery to the cells is critical to ensure efficient cell function. The CSIR and University of Pretoria (UP) have designed and developed a novel bio-artificial liver support system (BALSS) that utilizes perfluorooctyl bromide (PFOB) as artificial oxygen carrier. As the PFOB is not miscible with water, it needs to be emulsified. To enable the use of the PFOB emulsion in the UP-CSIR BALSS, a study was carried out to investigate relevant aspects relating to the PFOB emulsion, i.e. the formulation, manufacturing procedure, stability, rheology and mass transfer characteristics. The study results are reported in this dissertation, including a proposed mass transfer model for describing oxygen mass transfer to and from the PFOB emulsions. Emulsion stability can be improved through control of the droplet size and size distribution, limiting Ostwald ripening, and control of zeta potential of the dispersed phase droplets. PFOB emulsions with dispersed phase (PFOB) volume fractions between 0.4 and 0.5 and Sauter mean droplet diameter between 100 and 200 nm were found to be optimal for oxygen mass transfer in cell culture systems. The PFOB emulsion in the UP-CSIR BALSS can be concentrated and recirculated using ultrafiltration. Quantitative recovery of PFOB from its emulsions can be carried out using distillation with orthophosphoric acid. Experimental overall mass transfer coefficients for membrane oxygenators obtained without PFOB compared well with literature reported values of 2.5x10-5 m/s by Goerke et al. (2002) and 1 – 3x10-5 m/s by Schneider et al. (1995) for similar systems. The addition of 0.2 v/v PFOB leads to an increase in the membrane oxygenator mass transfer coefficient by a factor of about 30, and an increase in oxygen carrying capacity by a factor of about 4.5. It was also shown that suitable PFOB emulsions can have a significant impact on the growth and function of hepatocytes in a BALSS. / Thesis (PhD (Chemical Engineering))--University of Pretoria, 2005. / Chemical Engineering / unrestricted

Introduction The development of an appropriate animal model of ALF is paramount for the understanding of the disease pathogenesis and evaluation of potential therapeutic approaches. Acute liver failure (ALF) is a severe, usually rapidly progressive disease characterized by high mortality (60 - 90 %). Besides acute liver transplantation which faces a shortage of donors, the only possible therapeutic alternative is applying biological or non-biological liver support systems. To confirm the effectiveness of these methods, clinically relevant model of ALF on a large laboratory animal is essential. Surgically induced ALF models seem to be more reliable than models based on chemical intoxication. Ideal model of ALF has not yet been published. Surgical models are usually performed with devascularisation, large liver resection or hepatectomy. The aim of this work was to introduce three surgical models of ALF and evaluate their usefulness for testing biological and non-biological liver support systems. Materials and Methods Female laboratory pig weighing 35 - 45 kg was used for the experimental study. After induction of general anesthesia the thermodilution catheter was introduced via jugular vein. Femoral artery and vein were cannulated for invasive blood pressure monitoring and for infusions and...

Die Arbeit befasst sich mit Methoden der Erfassung metabolischer Funktion einer dynamischen Zellkultur in Bioreaktoren zur klinischen Verwendung als Leberunterstützungssystem. Der Fokus liegt auf der Etablierung von Tests zur Qualitätssicherung als effiziente In- Prozess Kontrollen während der Bereitstellungsphase der Zellkulturen. Weitere Ziele sind ein Beitrag zu Wirksamkeitsnachweis und zur Charakterisierung des Systems im Rahmen von Zulassungen als "Biologic". Bei der experimentellen Untersuchung unter Verwendung von Leberzellkulturen aus porcinen Organen wurden der Lidocain- MEGX Test, die Elimination von Galaktose und Sorbitol sowie die Elimination von Ammoniak und die Synthese von Harnstoff, Lactat und Albumin berücksichtigt. Dies erfolgte mittels Bolus- und Fliessgleichgewichtsuntersuchungen sowie Konzentrationsbestimmungen. Die Ergebnisse zeigten für alle Parameter einen hochsignifikanten Unterschied zu Zellfreien Bioreaktoren, sowie einen typischen Verlauf, der in eine initiale Adhäsionsphase, eine stabile Kulturphase und eine darauf folgende erweiterte Phase mit langsamer Abschwächung der Zellleistungen gegliedert werden kann. Die Parameter erwiesen sich prinzipiell alle geeignet für den Einsatz in der Qualitätssicherung der dynamischen Zellkultur. Ein kombiniertes Untersuchungsschema bestehend aus Parametern die die Integrität der Zellmembranen reflektieren, wie die Freisetzung von Enzymen, sowie metabolischen Parametern wie Lidocain, Galaktose, Sorbitol als auch Syntheseparameter, für Proteine Albumin, als Hepatozyten typische Leistungen die Synthese von Harnstoff und der Abbau von Lactat ist in der Lage einen ausreichend umfassenden Einblick in den Status der dynamischen Zellkultur zu liefern. / Objectives of this work were to find easy to handle every day quality assessment procedure for bioreactors that are intended for clinical trials, further more usefulness of these parameters in characterization and standardization of the Liver Support System. Parameters for evaluation had been: Lidocaine, Megx, Sorbitol, Galactose, Urea, Albumin and Lactate. Bolus and Continuous metabolic liver functions tests had been done, for synthesis and detoxification parameters liberation was measured by concentration. Highly significant difference between bioreactor with primary porcine liver cells and cell-free devices was shown by t-tests. Differences to other groups like infected bioreactors had been demonstrated with ANOVA. Results show typical course over the culture period that can be categorized into a initial phase of cell reorganization, a stable culture phase and an extended phase with slow decay. All parameter proved to be suitable for daily routine quality assessment in dynamic cell culture systems. The combination of parameters reflecting different specific cellular function is able to give more comprehensive insights in the status of the cell culture.