Rapid structural elucidation of composite bacterial hopanoids by atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry

Abstract

Bacteriohopanepolyols (BHPs) are membrane lipids produced by a wide range of eubacteria. Their use, however, as molecular markers of bacterial populations and processes has until recently been hampered by the lack of a suitable rapid method for fingerprinting their composition in complex environmental matrices. New analytical procedures employing ion trap mass spectrometry now allow us to investigate the occurrence of BHPs in diverse biological and environmental samples including bacterial cultures, soils, and recent and ancient sediments. Here, we describe the structural characterisation using atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry (APCI-LC/MS(n)) of a number of previously identified but less commonly occurring BHPs such as adenosylhopane and ribonylhopane. Many of the structures described here have previously only been reported in one or just a small number of cultured organisms having been isolated from large amounts of cellular mass (4-26 g) and identified by nuclear magnetic resonance (NMR) techniques after purification of individual compounds. Now, having established characteristic APCI fragmentation patterns, it is possible to rapidly screen many more bacterial cultures using only small amounts of material (<50 mg) as well as environmental samples for these atypical structures and a rapidly growing suite of novel structures.

Composite Bacterial Hopanoids and Their Microbial Producers across Oxygen Gradients in the Water Column of the California Current

“…BHPs exhibit considerable structural diversity, differing in the number, position, and nature of the functional groups in the side chain (1). The most common BHPs identified are tetrafunctionalized, usually with hydroxyl groups at C-32, C-33, and C-34, with the C-35 position (see Fig.…”

Biosynthesis of 2-methylbacteriohopanepolyols by an anoxygenic phototroph

“…PCC 6803 at a concentration below that which would be expected to produce detectable BHP based on prior experiments with this organism (data not shown), resulting in a PCR product of the expected length under the same PCR conditions. Given that we have not used methyl- 13 C, 2 H 3 -labeled compounds in any prior experiments, contamination by cyanobacteria can confidently be excluded.…”

“…Until now, BHPs methylated at C-2 ( Fig. 1a) had been found in abundance only in cyanobacteria (9)(10)(11), and in ''trace'' quantities in several other bacteria cultured aerobically (12,13). The accumulation of substantial quantities of 2-methylhopanes in sediments has thus been proposed as a marker for cyanobacteria (10), and the discovery of these biomarkers in 2.7-billion-year-old shales has been used as one line of evidence that cyanobacteria had evolved and were conducting oxygenic photosynthesis by that time (14)(15)(16).…”

Anaerobic ammonium-oxidising bacteria: A biological source of the bacteriohopanetetrol stereoisomer in marine sediments

“…BHPs were analysed by high performance liquid chromatography coupled to positive ion atmospheric pressure chemical ionization mass spectrometry (HPLC/APCI-MS), using a data-dependent scan mode (3 events) on an HPLC system equipped with an ion trap MS, as described in Talbot et al (2007) and van Winden et al (2012). BHT and BHT isomer ( …”