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This study was aimed to determine the effects of different concentrations of L-phenylalanine (0.25, 0.50 and 1.00 mM L-Phe) applied to adventitious root cultures for different culture durations on root growth and production of tropane alkaloids and phenolics in Hyoscyamus niger. For this purpose, adventitious roots obtained from petiole explants of in vitro seedlings were used as plant materials. After adventitious roots were cultured in liquid Murashige and Skoog (MS) medium, L-Phe was applied to the adventitious roots. Then the roots were harvested on the 1st, 3rd, and 7th day of the applications, fresh and dry root weights, root growth indexes were analysed. It was also examined the contents of tropane alkaloids (hyoscyamine and scopolamine) and phenolics (gallic acid, catechin, epicatechin, vanillin, cinnamic acid, rosmarinic acid, p-coumaric, o-coumaric acid, rutin, caffeic acid, ferulic acid, chlorogenic acid, and quercetin) by HPLC in both adventitious roots and culture medium. As a result, according to root growth parameters data, day*application interaction was not found statistically significant. The maximum scopolamine and hyoscyamine amounts were detected in the cultures with the application of 0.50 mM L-Phe for 3 days. In terms of phenolics, 0.50 mM and 1.00 mM L-Phe application with 1st and 3rd days were selected as the most appropriate L-Phe applications and culture duration. Consequently, it has been determined that in vitro secondary metabolite production may be increased in H. niger root cultures with the selection of appropriate L-Phe concentrations and culture durations. Key Message: L-phenylalanine is an effective and inexpensive application for the production of secondary metabolites in adventitious root cultures of Hyoscyamus niger with the correct concentration and appropriate culture duration. [ABSTRACT FROM AUTHOR]

ABSTRACT In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity and quality. Titer and N-glycosylation of mAbs, as well as proteomic signature and metabolic status of the production cells in the culture were assessed. We found that the impact of glucose starvation on the titer and N-glycosylation of mAbs was dependent on the degree of starvation during early stationary phase of the fed-batch culture. Higher degree of glucose starvation reduced intracellular concentrations of UDP-GlcNAc and UDP-GalNAc, but increased the levels of UDP-Glc and UDP-Gal. Increased GlcNAc and Gal occupancy correlated well with increased degree of glucose starvation, which can be attributed to the interplay between the dilution effect associated with change in specific productivity of mAbs and the changed nucleotide sugar metabolism. Herein, we also show and discuss that increased cell culture duration negatively affect the maturation of glycans. In addition, comparative proteomics analysis of cells was conducted to observe differences in protein abundance between early growth and early stationary phases. Generally higher expression of proteins involved in regulating cellular metabolism, extracellular matrix, apoptosis, protein secretion and glycosylation was found in early stationary phase. These analyses offered a systematic view of the intrinsic properties of these cells and allowed us to explore the root causes correlating culture duration with variations in the productivity and glycosylation quality of monoclonal antibodies produced with CHO cells. Biotechnol. Bioeng. 2015;112: 2172-2184. © 2015 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

BACKGROUND Currently used mathematical models for animal cell growth and metabolism cannot simulate changing culture pH and its effect on metabolism, and hence cannot accurately model cultures lacking pH control. Animal cell cultures are however widely cultured at a small scale without pH control, e.g. for clone screening during cell line development. A mathematical model for animal cell growth without pH control is developed and applied to design optimal conditions for clone screening without pH control. RESULTS The mathematical model successfully simulates published data for suspension CHO culture in shake flasks. It is further used to simulate clonal heterogeneity and predict relative clone performance in different culture conditions. Predictions of correlation coefficients between clone titers in batch and fed batch screening conditions and pH controlled fed batch agree with published values. Fed batch mode is predicted to give better screening outcomes than batch, but only for clones able to consume lactate. Simulations reveal that culture duration of the screening assay has a large effect on the screening outcome and optimal durations differ based on the ability or inability of clones to consume lactate. CONCLUSION This study presents a tool for optimal design of screening assay for clone selection without pH control. © 2014 Society of Chemical Industry [ABSTRACT FROM AUTHOR]