PMID- 23726243
OWN - NLM
STAT- MEDLINE
DCOM- 20131022
LR  - 20131121
IS  - 1873-3778 (Electronic)
IS  - 0021-9673 (Linking)
VI  - 1298
DP  - 2013 Jul 12
TI  - Direct quantification of bacterial molybdenum and iron metallophores with 
      ultra-high-performance liquid chromatography coupled to time-of-flight mass 
      spectrometry.
PG  - 50-60
LID - S0021-9673(13)00740-1 [pii]
LID - 10.1016/j.chroma.2013.05.008 [doi]
AB  - Metallophores are a unique class of organic ligands released, for example, by 
      nitrogen fixing bacteria in their environment in order to recruit the 
      micronutrients molybdenum (Mo) and iron (Fe). Mo and Fe are essential cofactors 
      of nitrogenase that reduces atmospheric nitrogen into bioavailable ammonium. Upon 
      release, these bacterial metallophores bind to both metal cations and oxo-anions 
      in the extracellular medium increasing the bioavailability of the metals to the 
      nitrogen fixers, which can subsequently recruit the complexes. The efficient 
      quantification of those metal complexes is crucial for the understanding of the 
      homeostasis of the metal cofactors of nitrogenase (e.g., Fe and Mo), the dynamics 
      of nitrogen fixation and the nitrogen cycle. A novel direct 
      ultra-high-performance liquid chromatography coupled to a time-of-flight mass 
      spectrometer (UHPLC-ToF-MS) was developed to quantify and monitor the production 
      of Fe and Mo complexes of the catecholate metallophores protochelin (Prot) and 
      azotochelin (Azo) in the growth medium of the nitrogen fixer and model organism 
      Azotobacter vinelandii. Chromatographic separations were carried on a reversed 
      C18-phase with a mobile phase ramped from water to acetonitrile spiked with 1 
      mmol/L ammonium acetate (pH 6.6) to achieve stability of the metal complexes. 
      Linearity for Mo-protochelin and Fe-protochelin was found at the concentration 
      range between 5.0x10(-8) and 9.0x10(-7) mol/L with a limit of detection of 
      2.0x10(-8) and 3.0x10(-8) mol/L, respectively. The coefficient of variation of 
      the procedure is in the range from 1.5 to 3.4%. The validation has hence 
      demonstrated that the UHPLC-ToF-MS methodology is a fast, precise, specific, 
      robust, and sensitive approach allowing the direct measurement of metallophores 
      in growth medium without any sample preparation. The UHPLC-ToF-MS methodology was 
      applied to the analysis of metallophores in our laboratory. Under lower Mo 
      concentration, the Mo-protochelin concentration peaks in the middle lag phase, 
      while the Fe-protochelin concentration rises to two maxima at the beginning of 
      the exponential phase and during the stationary phase. The results indicate that 
      the production of metallophores is highly dynamic throughout the growth and has 
      to be monitored with high sensitivity and temporal resolution.
CI  - Copyright (c) 2013 Elsevier B.V. All rights reserved.
FAU - Deicke, Michael
AU  - Deicke M
AD  - Friedrich Schiller University Jena, Institute of Inorganic and Analytical 
      Chemistry, Jena School for Microbial Communication, Lessingstr. 8, 07743 Jena, 
      Germany.
FAU - Bellenger, Jean-Philippe
AU  - Bellenger JP
FAU - Wichard, Thomas
AU  - Wichard T
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20130510
PL  - Netherlands
TA  - J Chromatogr A
JT  - Journal of chromatography. A
JID - 9318488
RN  - 81AH48963U (Molybdenum)
RN  - E1UOL152H7 (Iron)
SB  - IM
MH  - Chromatography, High Pressure Liquid/*methods
MH  - Iron/*chemistry
MH  - Mass Spectrometry/*methods
MH  - Molybdenum/*chemistry
EDAT- 2013/06/04 06:00
MHDA- 2013/10/23 06:00
CRDT- 2013/06/04 06:00
PHST- 2013/02/23 00:00 [received]
PHST- 2013/04/30 00:00 [revised]
PHST- 2013/05/02 00:00 [accepted]
PHST- 2013/06/04 06:00 [entrez]
PHST- 2013/06/04 06:00 [pubmed]
PHST- 2013/10/23 06:00 [medline]
AID - S0021-9673(13)00740-1 [pii]
AID - 10.1016/j.chroma.2013.05.008 [doi]
PST - ppublish
SO  - J Chromatogr A. 2013 Jul 12;1298:50-60. doi: 10.1016/j.chroma.2013.05.008. Epub 
      2013 May 10.