Isotope effect in gas—liquid chromatography of labelled compounds

Matucha, M.; Jockisch, W.; Verner, P; Anders, G

doi:10.1016/0021-9673(91)85030-j

Cited by 91 publications

(73 citation statements)

References 20 publications

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“…3). It should be noted that the retention behavior of per-deuterated fatty acids and hydrocarbons, with elution long before corresponding unlabeled compounds, was coherent with previous observations on per-deuterated alkanes (Matucha et al, 1991). Overall, labeling experiments therefore showed that C. reinhardtii and C. variabilis had enzyme(s) that catalyzed the conversion of long-chain fatty acids into alka(e)nes by a formal decarboxylation.…”

Section: Reinhardtii and C Variabilis Synthesize Longchain Alka(esupporting

confidence: 87%

Microalgae Synthesize Hydrocarbons from Long-Chain Fatty Acids via a Light-Dependent Pathway

Sorigué

Légeret²,

Cuiné³

et al. 2016

Plant Physiol.

119

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Microalgae are considered a promising platform for the production of lipid-based biofuels. While oil accumulation pathways are intensively researched, the possible existence of a microalgal pathways converting fatty acids into alka(e)nes has received little attention. Here, we provide evidence that such a pathway occurs in several microalgal species from the green and the red lineages. In Chlamydomonas reinhardtii (Chlorophyceae), a C17 alkene, n-heptadecene, was detected in the cell pellet and the headspace of liquid cultures. The Chlamydomonas alkene was identified as 7-heptadecene, an isomer likely formed by decarboxylation of cis-vaccenic acid. Accordingly, incubation of intact Chlamydomonas cells with per-deuterated D 31 -16:0 (palmitic) acid yielded D 31 -18:0 (stearic) acid, D 29 -18:1 (oleic and cis-vaccenic) acids, and D 29 -heptadecene. These findings showed that loss of the carboxyl group of a C 18 monounsaturated fatty acid lead to heptadecene formation. Amount of 7-heptadecene varied with growth phase and temperature and was strictly dependent on light but was not affected by an inhibitor of photosystem II. Cell fractionation showed that approximately 80% of the alkene is localized in the chloroplast. Heptadecane, pentadecane, as well as 7-and 8-heptadecene were detected in Chlorella variabilis NC64A (Trebouxiophyceae) and several Nannochloropsis species (Eustigmatophyceae). In contrast, Ostreococcus tauri (Mamiellophyceae) and the diatom Phaeodactylum tricornutum produced C 21 hexaene, without detectable C 15 -C 19 hydrocarbons. Interestingly, no homologs of known hydrocarbon biosynthesis genes were found in the Nannochloropsis, Chlorella, or Chlamydomonas genomes. This work thus demonstrates that microalgae have the ability to convert C 16 and C 18 fatty acids into alka(e)nes by a new, lightdependent pathway.

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Section: Reinhardtii and C Variabilis Synthesize Longchain Alka(esupporting

confidence: 87%

Microalgae Synthesize Hydrocarbons from Long-Chain Fatty Acids via a Light-Dependent Pathway

Sorigué

Légeret²,

Cuiné³

et al. 2016

Plant Physiol.

119

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“…Our analyses showed a strong correlation between the deuteration rate and a decrease in retention time. This chromatographic effect has been described previously for deuterated isotopomeres of several other molecules, including caffeine (5), n-alkanes (23), and fatty acid pentafluorobenzyl esters (26). This isotope effect is primarily attributed to the shorter COD covalent bond (instead of the longer COH bond), which modifies several physical properties of the deuterated FAMEs, such as hydrophobicity, which in turn affect the chromatographic properties of the molecules.…”

Section: Discussionsupporting

confidence: 63%

Stable-Isotope-Based Labeling of Styrene-Degrading Microorganisms in Biofilters

Alexandrino

Knief

Lipski

2001

Appl Environ Microbiol

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Deuterated styrene ([2 H 8 ]styrene) was used as a tracer in combination with phospholipid fatty acid (PLFA) analysis for characterization of styrene-degrading microbial populations of biofilters used for treatment of waste gases. Deuterated fatty acids were detected and quantified by gas chromatography-mass spectrometry. The method was evaluated with pure cultures of styrene-degrading bacteria and defined mixed cultures of styrene degraders and non-styrene-degrading organisms. Incubation of styrene degraders for 3 days with [ 2 H 8 ]styrene led to fatty acids consisting of up to 90% deuterated molecules. Mixed-culture experiments showed that specific labeling of styrene-degrading strains and only weak labeling of fatty acids of non-styrenedegrading organisms occurred after incubation with [ 2 H 8 ]styrene for up to 7 days. Analysis of actively degrading filter material from an experimental biofilter and a full-scale biofilter by this method showed that there were differences in the patterns of labeled fatty acids. For the experimental biofilter the fatty acids with largest amounts of labeled molecules were palmitic acid (16:0), 9,10-methylenehexadecanoic acid (17:0 cyclo9-10), and vaccenic acid (18:1 cis11). These lipid markers indicated that styrene was degraded by organisms with a Pseudomonas-like fatty acid profile. In contrast, the most intensively labeled fatty acids of the full-scale biofilter sample were palmitic acid and cis-11-hexadecenoic acid (16:1 cis11), indicating that an unknown styrene-degrading taxon was present. Iso-, anteiso-, and 10-methyl-branched fatty acids showed no or weak labeling. Therefore, we found no indication that styrene was degraded by organisms with methyl-branched fatty fatty acids, such as Xanthomonas, Bacillus, Streptomyces, or Gordonia spp.Extraction and analysis of chemotaxonomically important lipid markers from environmental samples constitute a wellestablished method for characterizing microbial communities, detecting community changes through time, or obtaining information about the metabolic status of a community (39). Recently, phospholipid fatty acid (PLFA) analyses were combined with carbon isotope labeling techniques to link degradation activities with specific microbial populations (2, 12, 25). 14 C tracers were successfully used for this approach (28). However, the main disadvantage of the procedure was the low efficiency of separation of the radiolabeled fatty acid methyl esters (FAMEs) caused by the discontinuous collection of fractions prior to scintillation counting. This was avoided by using substrates labeled with the stable 13 C isotope, which facilitated continuous detection of FAMEs by gas chromatography and on-line-combustion isotope ratio mass spectrometry (27).We studied the use of a deuterated substrate as an alternative to 13 C isotopes for characterization of actively degrading microbial populations in complex communities. The use of deuterated substrates with subsequent gas chromatographymass spectrometry analysis of the products is a well-establ...

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“…As can be seen in the selected-ion monitoring (SIM) chromatograms of the TMS ethers of the reaction product and authentic samples ( Figure 8B), the t R of the enzymatically formed methylation product was almost the same as, but slightly larger than, that of the deuterium-labeled standard, (±)-[4′-OC 2 H 3 ] arctigenins. This difference was ascribed to the well-known isotope effect on GC t R (Matucha et al 1991). On the other hand, the t R was very different from that of (±)-[4-OC 2 H 3 ]-isoarctigenins.…”

Section: Substrate Specificity Of Recombinant Ctomt2 and Ctomt3mentioning

confidence: 85%

“…Figure 3B) and the mass spectra (TMS ether) ( Figure 3A) with those of unlabeled authentic samples (TMS ethers) [(±)-arctigenins: t R , 14.74 min ( Figure 3B) and the mass spectrum: Figure 3A; (±)-isoarctigenins: t R , 15.20 min ( Figure 3B) and the mass spectrum: Figure 3A]. The slightly shorter t R of deuterium-labeled [OC 2 H 3 ] arctigenin compared with that of the unlabeled standard was due to an isotope effect in GC (Matucha et al 1991). Control experiments were conducted using the cell-free extracts from 12, 14, and 15 DAF seeds (Table 1).…”

Section: Omt Assay and Reaction Kinetics Of Recombinant C Tinctoriusmentioning

confidence: 99%

A lignan O-methyltransferase catalyzing the regioselective methylation of matairesinol in Carthamus tinctorius

Umezawa

Ragamustari

Nakatsubo

et al. 2013

Plant Biotechnology

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Lignans are a group of plant phenolic compounds with various biological activities, including antitumor and antioxidant properties. O-Methylation is a critical step in biosynthesis of these compounds. However, little is known about the O-methyltransferase (OMT) enzymes that catalyze lignan O-methylation. We discovered a highly regioselective OMT activity in safflower (Carthamus tinctorius) seeds that catalyzed the methylation of matairesinol, a dibenzylbutyrolactone lignan, into 4′-O-methylmatairesinol (arctigenin) but not 4-O-methylmatairesinol (isoarctigenin). By examining such OMT activity in correlation with OMT transcript abundances during seed development, we cloned a few putative OMT cDNAs and produced their recombinant proteins in Escherichia coli. Among them, one protein exhibited O-methylation activity for matairesinol with the regioselectivity identical to that of the plant protein, and was named C. tinctorius matairesinol OMT (CtMROMT). CtMROMT did not show any detectable OMT activities towards phenylpropanoid monomers under the reaction conditions tested, while it methylated flavonoid apigenin efficiently into 4′-O-methylapigenin (acacetin). However, quantitative real-time polymerase chain reaction analysis demonstrated that expression of the CtMROMT gene was synchronized with the CtMROMT activity profile and arctigenin accumulation in the plant. These results demonstrated that CtMROMT is a novel plant OMT for lignan methylation.

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Isotope effect in gas—liquid chromatography of labelled compounds

Cited by 91 publications

References 20 publications

Microalgae Synthesize Hydrocarbons from Long-Chain Fatty Acids via a Light-Dependent Pathway

Microalgae Synthesize Hydrocarbons from Long-Chain Fatty Acids via a Light-Dependent Pathway

Stable-Isotope-Based Labeling of Styrene-Degrading Microorganisms in Biofilters

A lignan O-methyltransferase catalyzing the regioselective methylation of matairesinol in Carthamus tinctorius

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