Intracellular pH based controlled cultivation of yeast cells: I. Measurement methodology

Sureshkumar, G. K.; Mutharasan, Raj

doi:10.1002/bit.260410116

Cited by 12 publications

(3 citation statements)

References 28 publications

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“…in specific buffers prior to intracellular pH measurement. Therefore, several authors suggest that these studies do not necessarily reflect what is happening inside of a bioreactor culture . To evaluate if extracellular pH inhomogeneities might affect the intracellular pH of the cells, one‐compartment batch fermentations with controlled pH shifts were conducted.…”

Section: Introductionmentioning

confidence: 99%

The impact of pH inhomogeneities on CHO cell physiology and fed‐batch process performance – two‐compartment scale‐down modelling and intracellular pH excursion

Brunner

Braun

Doppler

et al. 2017

Biotechnology Journal

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Due to high mixing times and base addition from top of the vessel, pH inhomogeneities are most likely to occur during large-scale mammalian processes. The goal of this study was to set-up a scale-down model of a 10-12 m stirred tank bioreactor and to investigate the effect of pH perturbations on CHO cell physiology and process performance. Short-term changes in extracellular pH are hypothesized to affect intracellular pH and thus cell physiology. Therefore, batch fermentations, including pH shifts to 9.0 and 7.8, in regular one-compartment systems are conducted. The short-term adaption of the cells intracellular pH are showed an immediate increase due to elevated extracellular pH. With this basis of fundamental knowledge, a two-compartment system is established which is capable of simulating defined pH inhomogeneities. In contrast to state-of-the-art literature, the scale-down model is included parameters (e.g. volume of the inhomogeneous zone) as they might occur during large-scale processes. pH inhomogeneity studies in the two-compartment system are performed with simulation of temporary pH zones of pH 9.0. The specific growth rate especially during the exponential growth phase is strongly affected resulting in a decreased maximum viable cell density and final product titer. The gathered results indicate that even short-term exposure of cells to elevated pH values during large-scale processes can affect cell physiology and overall process performance. In particular, it could be shown for the first time that pH perturbations, which might occur during the early process phase, have to be considered in scale-down models of mammalian processes.

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Section: Introductionmentioning

confidence: 99%

The impact of pH inhomogeneities on CHO cell physiology and fed‐batch process performance – two‐compartment scale‐down modelling and intracellular pH excursion

Brunner

Braun

Doppler

et al. 2017

Biotechnology Journal

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“…9-Acridinamine (the name generally used for the compound), one of the simplest representatives of the family of nitrogen organic bases, is a molecule interesting from the cognitive point of view owing to its relatively simple constitution and interesting physicochemical features, which are reflected, among other things, in its ability to interact with biomolecules. , For these reasons, 9-acridinamine has often been used as a model compound in studying the general properties of chemical entities. − Its biological relevance is exhibited by its mutagenic activity, − as well as its ability to interact with DNA ,− and other biologically important molecules. ,− Of some interest is the photodynamic activity of the compound. − 9-Acridinamine is also a convenient fluorescent probe for investigating pH and various physical features of biological systems. − …”

Section: Introductionmentioning

confidence: 99%

Hartree−Fock and Density Functional Methods and IR and NMR Spectroscopies in the Examination of Tautomerism and Features of Neutral 9-Acridinamine in Gaseous and Condensed Media

et al. 1997

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Two tautomeric molecules, 9-acridinamine (9-AA) and 9(10H)-acridinimine (9-AI), were examined at the ab initio Hartree−Fock (HF) and density functional (DFT) levels of theory with the 6-31G** basis sets. Solvent (hexane, CH3CN, H2O) effects were included in ab initio HF optimizations through the self-consistent reaction field (SCRF) technique. Subsequent Hessian calculations followed by the normal-mode analyses revealed all harmonic frequencies to be positive, thus confirming the validity of the geometry optimizations. The energies of the molecules at stationary points corresponding to ab initio HF geometries were supplemented with the second-order Møller−Plesset (MP2) electron correlation correction. Standard routines utilizing relationships of statistical thermodynamics enabled determination of enthalpies of formation (supplemented further with corrections arising from isogyric, hydrogenation, and isodesmic processes) and entropies (heat capacities) at selected temperatures, as well as constants revealing equilibrium between two tautomeric forms. Other physicochemical characteristics, such as bond orders, dipole moments, and energies of the lowest unoccupied (LUMO) and highest occupied (HOMO) molecular orbitals were also obtained from theoretical calculations. Thermochemical data indicate that 9-AA and 9-AI should coexist at ambient temperature. This is also confirmed by a comparison of experimental IR and Raman spectra with harmonic frequencies derived theoretically. 1H and 13C chemical shifts obtained at the GIAO level of theory correlate only qualitatively with relevant experimental NMR data and do not exclude the existence of tautomeric phenomena. The distributions of atomic partial charges and electrostatic potential around the molecules differ noticeably, which implies that 9-AA and 9-AI may behave differently with respect to biomolecules.

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“…The F 1 F 0 -ATPase uses the proton motive force generated by the electron transport chain across the inner mitochondrial membrane to drive phosphorylation of ADP to ATP. Consequently, intracellular pH and pH gradients are directly linked to cellular energy levels and metabolism [8].…”

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confidence: 99%

Intracellular pH homeostasis in the filamentous fungus Aspergillus niger

Hesse

Ruijter

Dijkema

et al. 2002

European Journal of Biochemistry

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Intracellular pH homeostasis in the filamentous fungus Aspergillus niger was measured in real time by 31 P NMR during perfusion in the NMR tube of fungal biomass immobilized in Ca 2+ -alginate beads. The fungus maintained constant cytoplasmic pH (pH cyt ) and vacuolar pH (pH vac ) values of 7.6 and 6.2, respectively, when the extracellular pH (pH ex ) was varied between 1.5 and 7.0 in the presence of citrate. Intracellular metabolism did not collapse until a DpH over the cytoplasmic membrane of 6.6-6.7 was reached (pH ex 0.7-0.8). Maintenance of these large pH differences was possible without increased respiration compared to pH ex 5.8. Perfusion in the presence of various hexoses and pentoses (pH ex 5.8) revealed that the magnitude of DpH values over the cytoplasmic and vacuolar membrane could be linked to the carbon catabolite repressing properties of the carbon source. Also, larger DpH values coincided with a higher degree of respiration and increased accumulation of polyphosphate. Addition of protonophore (carbonyl cyanide m-chlorophenylhydrazone, CCCP) to the perfusion buffer led to decreased ATP levels, increased respiration and a partial (1 lM CCCP), transient (2 lM CCCP) or permanent (10 lM CCCP) collapse of the vacuolar membrane DpH. Nonlethal levels of the metabolic inhibitor azide (N 3 -, 0.1 mM) caused a transient decrease in pH cyt that was closely paralleled by a transient vacuolar acidification. Vacuolar H + influx in response to cytoplasmic acidification, also observed during extreme medium acidification, indicates a role in pH homeostasis for this organelle. Finally, 31 P NMR spectra of citric acid producing A. niger mycelium showed that despite a combination of low pH ex (1.8) and a high acidsecreting capacity, pH cyt and pH vac values were still well maintained (pH 7.5 and 6.4, respectively).

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Intracellular pH based controlled cultivation of yeast cells: I. Measurement methodology

Cited by 12 publications

References 28 publications

The impact of pH inhomogeneities on CHO cell physiology and fed‐batch process performance – two‐compartment scale‐down modelling and intracellular pH excursion

The impact of pH inhomogeneities on CHO cell physiology and fed‐batch process performance – two‐compartment scale‐down modelling and intracellular pH excursion

Hartree−Fock and Density Functional Methods and IR and NMR Spectroscopies in the Examination of Tautomerism and Features of Neutral 9-Acridinamine in Gaseous and Condensed Media

Intracellular pH homeostasis in the filamentous fungus Aspergillus niger

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