TY - JOUR

T1 - Quantification of bacterial polyhydroxyalkanoic acids by Nile red staining

AU - Gorenflo, V.

AU - Steinbüchel, A.

AU - Marose, S.

AU - Rieseberg, M.

AU - Scheper, T.

N1 - Funding information: This study was partially supported by a grant provided by the European Commission (FAIR programme) and by a BriteEuram project of the European Commission (BE project BRE2-CT94-1005). We thank Professor Y. Doi (RIKEN, Saitama, Japan) for providing a sample of synthetic poly(R,S-3HB).

PY - 1999/6

Y1 - 1999/6

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=0032973638&partnerID=8YFLogxK

U2 - 10.1007/s002530051460

DO - 10.1007/s002530051460

M3 - Article

C2 - 10422224

AN - SCOPUS:0032973638

VL - 51

SP - 765

EP - 772

JO - Applied Microbiology and Biotechnology

JF - Applied Microbiology and Biotechnology

SN - 0175-7598

IS - 6

ER -