Abstract
The development of an efficient analytical method for the reliable detection and identification of the biosynthetic intermediates found in microbial cultures, which usually produce complex intermediates of the metabolites of interest, is essential for further biosynthetic investigations. This study developed a simple and highly selective method for detecting the biosynthetic intermediates involved in the FK506 pathway of Streptomyces clavuligerus KCTC 10561BP involving a cleanup procedure using a solid-phase extraction technique to provide reliable extraction of FK506-related compounds from a cell culture broth and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) to separate and detect the FK506-related intermediates at concentrations as low as 0.2 μg/L in the broth. This method enabled the analytical profiling of the intermediates formed during the biosynthesis of FK506 in this S. clavuligerus strain, which produced FK506 as a main product. Eight FK506 intermediates-FK520, 37,38-dihydroFK506, prolylFK506, 9-decarbonyl-9-hydroxylFK506, 9-deoxoFK506, desmethylFK520, prolylFK520, and 9-deoxoFK520-were identified. This is the first report of the LC-ESI-MS/MS characterization of a wide range of FK506 analogs from a bacterial fermentation broth. The protocol employed in this study may be useful for estimating the structure of the metabolites without the need for a time-consuming isolation process and nuclear magnetic resonance (NMR) spectroscopy.
| Original language | English |
|---|---|
| Pages (from-to) | 1-7 |
| Number of pages | 7 |
| Journal | Analytical Biochemistry |
| Volume | 393 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - 2009 Oct 1 |
| Externally published | Yes |
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Keywords
- FK506 profiling
- Liquid chromatography-mass spectrometry (LC-MS)
- Tacrolimus
ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Molecular Biology
- Cell Biology
Cite this
Liquid chromatography-mass spectrometry characterization of FK506 biosynthetic intermediates in Streptomyces clavuligerus KCTC 10561BP. / Park, Je Won; Mo, Sang Joon; Park, Sung Ryeol; Ban, Yeon Hee; Yoo, Young Ji; Yoon, Yeo Joon.
In: Analytical Biochemistry, Vol. 393, No. 1, 01.10.2009, p. 1-7.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Liquid chromatography-mass spectrometry characterization of FK506 biosynthetic intermediates in Streptomyces clavuligerus KCTC 10561BP
AU - Park, Je Won
AU - Mo, Sang Joon
AU - Park, Sung Ryeol
AU - Ban, Yeon Hee
AU - Yoo, Young Ji
AU - Yoon, Yeo Joon
PY - 2009/10/1
Y1 - 2009/10/1
N2 - The development of an efficient analytical method for the reliable detection and identification of the biosynthetic intermediates found in microbial cultures, which usually produce complex intermediates of the metabolites of interest, is essential for further biosynthetic investigations. This study developed a simple and highly selective method for detecting the biosynthetic intermediates involved in the FK506 pathway of Streptomyces clavuligerus KCTC 10561BP involving a cleanup procedure using a solid-phase extraction technique to provide reliable extraction of FK506-related compounds from a cell culture broth and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) to separate and detect the FK506-related intermediates at concentrations as low as 0.2 μg/L in the broth. This method enabled the analytical profiling of the intermediates formed during the biosynthesis of FK506 in this S. clavuligerus strain, which produced FK506 as a main product. Eight FK506 intermediates-FK520, 37,38-dihydroFK506, prolylFK506, 9-decarbonyl-9-hydroxylFK506, 9-deoxoFK506, desmethylFK520, prolylFK520, and 9-deoxoFK520-were identified. This is the first report of the LC-ESI-MS/MS characterization of a wide range of FK506 analogs from a bacterial fermentation broth. The protocol employed in this study may be useful for estimating the structure of the metabolites without the need for a time-consuming isolation process and nuclear magnetic resonance (NMR) spectroscopy.
AB - The development of an efficient analytical method for the reliable detection and identification of the biosynthetic intermediates found in microbial cultures, which usually produce complex intermediates of the metabolites of interest, is essential for further biosynthetic investigations. This study developed a simple and highly selective method for detecting the biosynthetic intermediates involved in the FK506 pathway of Streptomyces clavuligerus KCTC 10561BP involving a cleanup procedure using a solid-phase extraction technique to provide reliable extraction of FK506-related compounds from a cell culture broth and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) to separate and detect the FK506-related intermediates at concentrations as low as 0.2 μg/L in the broth. This method enabled the analytical profiling of the intermediates formed during the biosynthesis of FK506 in this S. clavuligerus strain, which produced FK506 as a main product. Eight FK506 intermediates-FK520, 37,38-dihydroFK506, prolylFK506, 9-decarbonyl-9-hydroxylFK506, 9-deoxoFK506, desmethylFK520, prolylFK520, and 9-deoxoFK520-were identified. This is the first report of the LC-ESI-MS/MS characterization of a wide range of FK506 analogs from a bacterial fermentation broth. The protocol employed in this study may be useful for estimating the structure of the metabolites without the need for a time-consuming isolation process and nuclear magnetic resonance (NMR) spectroscopy.
KW - FK506 profiling
KW - Liquid chromatography-mass spectrometry (LC-MS)
KW - Tacrolimus
UR - http://www.scopus.com/inward/record.url?scp=67651004256&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=67651004256&partnerID=8YFLogxK
U2 - 10.1016/j.ab.2009.06.021
DO - 10.1016/j.ab.2009.06.021
M3 - Article
C2 - 19539594
AN - SCOPUS:67651004256
VL - 393
SP - 1
EP - 7
JO - Analytical Biochemistry
JF - Analytical Biochemistry
SN - 0003-2697
IS - 1
ER -