Applied Microbiology and Biotechnology

, Volume 51, Issue 6, pp 765–772

Quantification of bacterial polyhydroxyalkanoic acids by Nile red staining

  • V. Gorenflo
  • A. Steinbüchel
  • S. Marose
  • M. Rieseberg
  • T. Scheper
ORIGINAL PAPER

DOI: 10.1007/s002530051460

Cite this article as:
Gorenflo, V., Steinbüchel, A., Marose, S. et al. Appl Microbiol Biotechnol (1999) 51: 765. doi:10.1007/s002530051460

Abstract

The fluorescence properties of one chemically and seven biologically produced polyhydroxyalkanoic acids were investigated as film castings and in living cells respectively after staining with Nile red. All these polyesters show a similar fluorescence behaviour, revealing a clear fluorescence maximum at an excitation wavelength between 540 nm and 560 nm and an emission wavelength between 570 nm and 605 nm. This could be shown by the use of two-dimensional fluorescence spectroscopy and flow cytometry. The examination of native poly(3-hydroxybutyric acid), poly(3HB), granules isolated from cells of Ralstonia eutropha H16 showed that the addition of 6.0 μg Nile red is necessary for total staining of 1.0 mg granules. The fluorescence intensity at an excitation wavelength of 550 nm and an emission wavelength of 600 nm showed high correlation to the poly(3HB) concentration of grana suspensions at different grana concentrations. These results and the staining of cell suspensions during cultivation experiments revealed that Nile red has a high potential for the quantitative determination of hydrophobic bacterial polyhydroxyalkanoic acids.

Copyright information

© Springer-Verlag Berlin Heidelberg 1999

Authors and Affiliations

  • V. Gorenflo
    • 1
  • A. Steinbüchel
    • 1
  • S. Marose
    • 2
  • M. Rieseberg
    • 2
  • T. Scheper
    • 2
  1. 1.Institut für Mikrobiologie, Westfälische Wilhelms-Universität Münster, Corrensstraße 3, D-48149 Münster, Germany e-mail: steinbu@uni-muenster.de Tel.: +49-251-8339821 Fax: +49-251-8338388DE
  2. 2.Institut für Technische Chemie, Universität Hannover, Callinstraße 3, D-30167 Hannover, GermanyDE