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Respirometric 13C flux analysis, Part I: Design, construction and validation of a novel multiple reactor system using on-line membrane inlet mass spectrometry

Paper ID Volume ID Publish Year Pages File Format Full-Text
31977 44864 2006 15 PDF Available
Title
Respirometric 13C flux analysis, Part I: Design, construction and validation of a novel multiple reactor system using on-line membrane inlet mass spectrometry
Abstract

A novel method for 13C flux analysis based on on-line CO2 labeling measurements is presented. This so-called respirometric 13C flux analysis requires multiple parallel 13C labeling experiments using differently labeled tracer substrates. In Part I of the work, a membrane-inlet mass spectrometry-based measurement system with 6 parallel reactors with each 12 ml liquid volume and associated experimental and computational methods for the respirometric 13C data acquisition and evaluation are described. Signal dynamics after switching between membrane probes follow exactly first-order allowing extrapolation to steady state. Each measurement cycle involving 3 reactors takes about 2 min. After development of a dynamic calibration method, the suitability and reliability of the analysis was examined with a lysine-producing mutant of Corynebacterium glutamicum using [1-13C1], [6-13C1], [1,6-13C2] glucose. Specific rates of oxygen uptake and CO2 production were estimated with an error less than ±0.3 mmol g−1 h−1 and had ±3% to ±10% deviations between parallel reactors which is primarily caused by inaccuracies in initial biomass concentration. The respiratory quotient could be determined with an uncertainty less than ±0.02 and varied only ±3% between reactors. Fractional labeling of CO2 was estimated with much higher precision of about ±0.001 to ±0.005. The detailed statistical analysis suggested that these data should be of sufficient quality to allow physiological interpretation and metabolic flux estimation. The obtained data were applied for the respirometric 13C metabolic flux analysis in Part II.

Keywords
CO2 labeling; Respiration; Mass spectrometry; Multi-channeled membrane-inlet; Miniaturized reactor; Multiple 13C labeling experiments; Respirometric 13C flux analysisMCS, Monte Carlo simulation; MS, mass spectrometer; OSDM, on-site short-term dynamic meas
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Respirometric 13C flux analysis, Part I: Design, construction and validation of a novel multiple reactor system using on-line membrane inlet mass spectrometry
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Publisher
Database: Elsevier - ScienceDirect
Journal: Metabolic Engineering - Volume 8, Issue 5, September 2006, Pages 417–431
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
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