Application of multi-parameter flow cytometry using fluorescent probes to study substrate toxicity in the indene bioconversion

The bioconversion of indene to cis‐(1S,2R) indandiol, a potential key intermediate in the synthesis of Merck's HIV protease inhibitor, CRIXIVAN™, can be achieved using a Rhodococcus strain. This study using Rhodococcus I24 reports on the application of multiparameter flow cytometry for the meas...

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Bibliographic Details
Published in:Biotechnology and bioengineering 2002-11, Vol.80 (3), p.239-249
Main Authors: Amanullah, A., Hewitt, C. J., Nienow, A. W., Lee, C., Chartrain, M., Buckland, B. C., Drew, S. W., Woodley, J. M.
Format: Article
Language:English
Subjects:
Adenosine Triphosphate - metabolism
Biological and medical sciences
Bioreactors
Biotechnology
Biotransformation - physiology
Catalysis
Cell Membrane - metabolism
Fermentation
Flow Cytometry - instrumentation
Flow Cytometry - methods
Fluorescent Dyes
Fundamental and applied biological sciences. Psychology
indene bioconversion
indene toxicity
Indenes - pharmacology
Methods. Procedures. Technologies
multi-parameter flow cytometry
Others
Oxygen - metabolism
Reference Values
Reproducibility of Results
Rhodococcus
Rhodococcus - cytology
Rhodococcus - drug effects
Rhodococcus - growth & development
Rhodococcus - metabolism
Sensitivity and Specificity
Staining and Labeling - methods
Various methods and equipments
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Summary:The bioconversion of indene to cis‐(1S,2R) indandiol, a potential key intermediate in the synthesis of Merck's HIV protease inhibitor, CRIXIVAN™, can be achieved using a Rhodococcus strain. This study using Rhodococcus I24 reports on the application of multiparameter flow cytometry for the measurement of cell physiological properties based on cytoplasmic membrane (CM) integrity and membrane depolarization as indicators of toxic effects of the substrate, indene. Quantification of intact polarized CM, intact depolarized CM and permeabilized CM of a large population of bacterial cells has been conducted using specific intracellular and membrane‐binding fluorescent stains. Measurements of oxygen uptake rate (OUR) and optical density (OD) as indicators of metabolic activity and biomass growth, respectively, were also made. Indene concentrations of up to 0.25 g/L (0.037 g indene/g dry cell weight) did not significantly (
ISSN:0006-3592
1097-0290
DOI:10.1002/bit.10477