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General Information
Manuscript title Generating short-term kinetic responses of primary metabolism of Penicillium chrysogenum through glucose perturbation in the bioscope mini reactor.
PubMed ID 16807032
Journal Metabolic Engineering
Year 2006
Authors U. Nasution, W.M. Van Gulik, A. Proell, W.A. van Winden, J.J. Heijnen
Affiliations Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, Netherlands
Keywords glucose pulse, in vivo kinetics, metabolome, Penicillium chrysogenum
Full text article Nasution_2006.pdf
Project name not specified

Experiment Description
Organism Penicillium chrysogenum
Strain DS12975
Data type time-series data of metabolites
Data units (µmol/gDW)
Execution date not specified

Experimental Details
Temperature (°C) 25.0
pH 6.5
Carbon source glucose
Culture mode chemostat
Process condition aerobic
Dilution rate (h⁻¹) 0.05
Working volume (L) 4.0
Biomass concentration (g/L) 6.21 ± 0.16
Medium composition

16.5 g/L glucose·H2O, 1 g/L KH2PO4, 5 g/L (NH4)2SO4, 0.5 g/L MgSO4·7H2O, and 10 ml/L of trace element solution. The trace element solution contained 15 g/L Na2-EDTA·2H2O, 0.5 g/L CuSO4·5H2O, 2 g/L ZnSO4·7H2O, 2 g/L MnSO4·H2O, 4 g/L FeSO4·7H2O and 0.5 g/L CaCl2·2H2O. For penicillin-G production the side chain precursor phenylacetic acid (PAA) was added at a concentration of 0.94 g/L.

General protocol information Sampling method: rapid sampling of broth for measurement of intracellular metabolites was carried out at 5, 10, 15, 20, 25, 35, 50, 70 and 90 s after injection of the glucose solution.

Quenching procedure: 60 w/w % analytical grade methanol (ARCOS, Geel, Belgium) and water purified in a milli-QUFplus system (Millipore, Bedford, MA), buffered with 10 mM HEPES (Merck, Darmstadt, Germany) and adjusted to pH 7.5 with 3M KOA (Baker, Deventer, The Netherlands).

Extraction technique: hot ethanol

Sample analyzing method: HPLC-UV/RI, LC-ESI-MS

Methods description - Notes

Chemostat pulse experiments - for the pulse response experiment carried out in the chemostat 16 ml of a 125 g/L glucose solution was injected into the reactor within 1 s. This increased the residual glucose concentration to approximately 0.5 g/L. Subsequently, rapid sampling ...

[1] Lange, H.C., Heijnen, J.J., 2001. Improved rapid sampling for in vivo kinetics of intracellular metabolites in S. cerevisiae. Biotechnol. Bioeng. 75, 406–415.
[2] Mashego, M.R., van Gulik, W.M., Vinke, J.L., Heijnen, J.J., 2003. Critical evaluation of sampling techniques for residual glucose determination in carbon-limited chemostat culture of Saccharomyces cerevisiae. Biotechnol. Bioeng. 83, 395–399.
[3] Gonzalez, B., Francois, J., Renaud,M., 1997. A rapid and reliable method for metabolite extraction in yeast using boiling buffered ethanol. Yeast 13, 1347–1355.
[4] Nasution, U., van Gulik, W., Kleijin, R., Heijnen, J.J., 2006. Measurement of intracellular metabolites of primary metabolism and adenine nucleotides in chemostat cultivated P. chrysogenum. Biotechnol. Bioeng. 94, 159–166.
[5] Wu, L., Mashego, M.R., Van Dam, J.C., Proell, A.M., Vinke, J.L., Ras, C., van Winden, W.A., van Gulik, W.M., Heijnen, J.J., 2005. Quantitative analysis of the microbial metabolome by isotope dilution mass spectrometry using uniformly C-13-labeled cell extracts as internal standards. Anal. Biochem. 336, 164–171.
[6] Christensen, L.H., Mandrup, G., Nielsen, J., Villadsen, J., 1994. A robust liquid-chromatographic method for measurement of medium components during penicillin fermentations. Anal. Chim. Acta 296, 51–62.

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Entered by: Administrator KiMoSysFirst name: Administrator
Affiliation: INESC-ID/IST
Interests: mathematical modeling, accessible data, use of data

Created: 2013-06-21 17:03:42 UTC

Updated: 2020-04-24 16:10:34 UTC

Version: 0

Status: (reviewed) 2013-12-06 17:17:38 UTC

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