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Determination of heme in microorganisms using HPLC-MS/MS and cobalt(III) protoporphyrin IX inhibition of heme acquisition in Escherichia coli
Stockholm University, Faculty of Science, Department of Environmental Science and Analytical Chemistry.
Stockholm University, Faculty of Science, Department of Environmental Science and Analytical Chemistry.
2017 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650Article in journal (Refereed) Epub ahead of print
Abstract [en]

One of the main threats to the achievements in modern medicine is antimicrobial resistance. Molecular targeting of bacterial acquisition mechanisms of heme has been suggested to be an alternative to antibiotics. In the present study, HPLC-MS/MS combined with a simple clean-up based on liquid-liquid extraction has been developed and evaluated for simultaneous determination of heme and porphyrin heme precursors in microorganisms. Experimental design was used to optimize the extraction parameters, to obtain a method with high recovery, low matrix effects, and high precision. The effects of additives in the culture medium on the biosynthesis of heme were studied using Escherichia coli as a model microorganism. 5-Aminolaevulinic acid and hemin increased the heme concentration in E. coli by a factor of 1.5 and 4.5, respectively. Addition of 5-aminolaevulinic acid bypassed the E. coli negative feedback control of heme biosynthesis, which led to high amounts of intracellular porphyrins. The high heme concentration obtained when hemin was used as a culture additive shows that E. coli has an uptake of heme from its surroundings. In contrast, addition of cobalt protoporphyrin IX to the growth medium reduced the amount of heme in E. coli, demonstrating this compound’s ability to mimic real heme and inhibit the heme acquisition mechanisms.

Place, publisher, year, edition, pages
2017.
Keyword [en]
Heme acquisition, Heme analysis, Escherichia coli, HPLC-MS/MS, Antimicrobial resistance, Porphyrins
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
Identifiers
URN: urn:nbn:se:su:diva-149378DOI: 10.1007/s00216-017-0610-5PubMedID: 29043383OAI: oai:DiVA.org:su-149378DiVA: diva2:1161248
Available from: 2017-11-29 Created: 2017-11-29 Last updated: 2017-12-04
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