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Culture Volume (culture + volume)

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Life Sciences100%

Selected Abstracts

Bioreactor for cultivation of red beet hairy roots and in situ recovery of primary and secondary metabolites

ENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 3 2009
Bhagyalakshmi Neelwarne
Abstract To arrive at an appropriate bioreactor design and in situ recovery of the products, red beet hairy roots were used as a model system where the levels of betalain pigments (betacyanins and betaxanthins) were followed as secondary metabolite and the peroxidase enzyme as primary metabolite. Medium volume and other kinetic parameters were found to play significant roles by way of directly affecting the biomass yield rather than a specific metabolite. The hydrodynamic stress created on the roots by large culture volume could be minimized by pulse-feeding of medium in shake-flasks; and by separating the biomass chamber from the aerated medium reservoir in circulatory fed-batch bioreactor. Accordingly the bioreactor was modified to provide anchorage and air-enrichment chamber which resulted in higher formation of both the metabolites than in shake-flasks. Various down-stream processing aspects such as in situ release of pigments by non-destructive methods, followed by adsorption through a column and recovery by desorption were optimized for betalains. A strategy for simultaneous recovery of pigment and peroxidase was worked out using aqueous two phase extraction (ATPE). [source]

Noninvasive measurement of dissolved oxygen in shake flasks

BIOTECHNOLOGY & BIOENGINEERING, Issue 5 2002
Leah Tolosa
Abstract Shake flasks are ubiquitous in cell culture and fermentation. However, conventional devices for measuring oxygen concentrations are impractical in these systems. Thus, there is no definitive information on the oxygen supply of growing cells. Here we report the noninvasive, nonintrusive monitoring of dissolved oxygen (DO) in shake flasks using a low-cost optical sensor. The oxygen-sensitive element is a thin, luminescent patch affixed to the inside bottom of the flask. The sensitivity and accuracy of this device is maximal up to 60% DO, within the range that is critical to cell culture applications. By measuring actual oxygen levels every 1 or 5 min throughout the course of yeast and E. coli fermentations, we found that a modest increase in shaker speed and a decrease in culture volume slowed the onset of oxygen limitation and reduced its duration. This is the first time that in situ oxygen limitation is reported in shake flasks. The same data is unattainable with a Clark type electrode because the presence of the intrusive probe itself changes the actual conditions. Available fiber optic oxygen sensors require cumbersome external connections and recalibration when autoclaved. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 80: 594,597, 2002. [source]

Feed development for fed-batch CHO production process by semisteady state analysis

BIOTECHNOLOGY PROGRESS, Issue 3 2010
Sarwat F. Khattak
Abstract Semisteady state cultures are useful for studying cell physiology and facilitating media development. Two semisteady states with a viable cell density of 5.5 million cells/mL were obtained in CHO cell cultures and compared with a fed-batch mode control. In the first semisteady state, the culture was maintained at 5 mM glucose and 0.5 mM glutamine. The second condition had threefold higher concentrations of both nutrients, which led to a 10% increase in lactate production, a 78% increase in ammonia production, and a 30% reduction in cell growth rate. The differences between the two semisteady states indicate that maintaining relatively low levels of glucose and glutamine can reduce the production of lactate and ammonia. Specific amino acid production and consumption indicated further metabolic differences between the two semisteady states and fed-batch mode. The results from this experiment shed light in the feeding strategy for a fed-batch process and feed medium enhancement. The fed-batch process utilizes a feeding strategy whereby the feed added was based on glucose levels in the bioreactor. To evaluate if a fixed feed strategy would improve robustness and process consistency, two alternative feeding strategies were implemented. A constant volume feed of 30% or 40% of the initial culture volume fed over the course of cell culture was evaluated. The results indicate that a constant volumetric-based feed can be more beneficial than a glucose-based feeding strategy. This study demonstrated the applicability of analyzing CHO cultures in semisteady state for feed enhancement and continuous process improvement. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010 [source]

293 cell cycle synchronisation adenovirus vector production

BIOTECHNOLOGY PROGRESS, Issue 1 2009
Tiago B. Ferreira
Abstract As the market requirements for adenovirus vectors (AdV) increase, the maximisation of the virus titer per culture volume per unit time is a key requirement. However, despite the fact that 293 cells can grow up to 8 × 106 cell/mL in simple batch mode operations, for optimal AdV infection a maximum cell density of 1 × 106 cell/mL at infection time has usually been utilized due to the so called "cell density effect". In addition, AdV titer appears to be dependent upon cell cycle phase at the time of infection. To evaluate the dependence of AdV production upon cell cycle phase, 293 cells were chemically synchronised at each phase of the cell cycle; a 2.6-fold increase on AdV cell specific titer was obtained when the percentage of cells at the S phase of the cell cycle was increased from 36 to 47%; a mathematical equation was used to relate AdV cell specific productivities with cell synchronisation at the S phase using this data. To avoid the use of chemical inhibitors, a temperature shift strategy was also used for synchronisation at the S phase. S phase synchronisation was obtained by decreasing the culture temperature to 31°C during 67 h and restoring it to 37°C during 72 h. By using this strategy we were able to synchronise 57% of the population in the S phase of the cell cycle obtaining an increase of 7.3-fold on AdV cell specific titer after infection. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source]