Quantification and Carbon and Nitrogen Isotopic Measurements of Heme B in Environmental Samples

Abstract

Heme B is an iron-coordinated tetrapyrrole molecule that acts as a cofactor in hemoproteins. It is expected to be ubiquitous in the environment, as b-type hemoproteins catalyze a variety of essential biochemical reactions. In this study, we developed an analytical method to quantify heme B in biological and environmental samples using high-performance liquid chromatography (HPLC) coupled to a photodiode array detector. The applicability of our method was further extended by the use of liquid chromatography/mass spectrometry (LC/MS; detection limit: ∼1 fmol), which enabled the quantification of a trace amount of dissolved heme B in filtered seawater and sedimentary heme B coexisting with an abundant interfering organic matrix. For compound-specific carbon and nitrogen isotopic measurements, heme B was successfully isolated and purified from biological and environmental samples by a combination of anion-exchange column chromatography, methyl esterification, and dual-step HPLC. While carbon and nitrogen isotopic compositions of heme B in phototrophs were mostly comparable to those of chlorophyll a, heme B in suspended particulate materials in coastal water and an intertidal sediment was 13C-depleted and 15N-enriched relative to chlorophyll a, suggesting that nonphototrophic microorganisms are also a significant source of heme B in natural environments.

Novel Aggregation-Enhanced PEC Photosensitizer Based on Electrostatic Linkage of Ionic Liquid with Protoporphyrin IX for Ultrasensitive Detection of Molt-4 Cells

“…Specifically, a weak ion peak is located at m/z = 561.2, corresponding to PPIX. 44 In addition, a stronger ion peak is centered at m/z = 844.9, which is well assigned to molecular weight of PPIX with two [BMIm] + groups, owing to the PPIX containing double side-chain carboxyl groups.…”

Cultivating the uncultured: Harnessing the “sandwich agar plate” approach to isolate heme‐dependent bacteria from marine sediment

“…We found that bacteria in natural environments could adapt to lower heme concentrations than host‐associated bacteria, and bacteria in sediment required a higher hemin concentration than aquatic bacteria (Figure S7 and Table S7). The concentrations of total heme in sheep blood, sediment, and water were ~7 mM, 0.14–1.8 nmol/g, and 73.9–174.7 pM, respectively 30 . Bacteria in natural habitats have a low heme requirement, which may be an adaptation to oligotrophic environments.…”

A new era of isotope ecology: Nitrogen isotope ratio of amino acids as an approach for unraveling modern and ancient food web

“…162) A recent analysis of heme B, the most abundant heme species in the natural environment, suggested that the TDF of this compound may be relatively small. 163) The gap between the two approaches, amino acids and bulk, should narrow as more such data become available.…”