Insights Into the Known 13C Depletion of Methane—Contribution of the Kinetic Isotope Effects on the Serine Hydroxymethyltransferase Reaction

We determined the kinetic isotope effect on the serine hydroxymethyltransferase reaction (SHMT), which provides important C1 metabolites that are essential for the biosynthesis of DNA bases, O-methyl groups of lignin and methane. An isotope effect on the SHMT reaction was suggested being responsible for the well-known isotopic depletion of methane. Using the cytosolic SHMT from pig liver, we measured the natural carbon isotope ratios of both atoms involved in the bond splitting by chemical degradation of the remaining serine before and after partial turnover. The kinetic isotope effect 13(VMax/Km) was 0.994 0.006 and 0.995 0.007 on position C-3 and C-2, respectively. The results indicated that the SHMT reaction does not contribute to the 13C depletion observed for methyl groups in natural products and methane. However, from the isotopic pattern of caffeine, isotope effects on the methionine synthetase reaction and on reactions forming Grignard compounds, the involved formation and fission of metal organic bonds are likely responsible for the observed general depletion of “activated” methyl groups. As metal organic bond formations in methyl transferases are also rate limiting in the formation of methane, they may likely be the origin of the known 13C depletion in methane.

Deep neural network for detecting arbitrary precision peptide features through attention based segmentation

A promising technique of discovering disease biomarkers is to measure the relative protein abundance in multiple biofluid samples through liquid chromatography with tandem mass spectrometry (LC-MS/MS) based quantitative proteomics. The key step involves peptide feature detection in the LC-MS map, along with its charge and intensity. Existing heuristic algorithms suffer from inaccurate parameters and human errors. As a solution, we propose PointIso, the first point cloud based arbitrary-precision deep learning network to address this problem. It consists of attention based scanning step for segmenting the multi-isotopic pattern of 3D peptide features along with the charge, and a sequence classification step for grouping those isotopes into potential peptide features. PointIso achieves 98% detection of high-quality MS/MS identified peptide features in a benchmark dataset. Next, the model is adapted for handling the additional ‘ion mobility’ dimension and achieves 4% higher detection than existing algorithms on the human proteome dataset. Besides contributing to the proteomics study, our novel segmentation technique should serve the general object detection domain as well.

Towards Aldehydomics: Untargeted Trapping and Analysis of Reactive Diet-Related Carbonyl Compounds Formed in the Intestinal Lumen

Lipid peroxidation and subsequent formation of toxic aldehydes, such as 4-hydroxynonenal, is known to be involved in numerous pathophysiological processes, possibly including the development of colorectal cancer. This work aimed at the development of an untargeted approach using high-performance liquid chromatography coupled with high-resolution mass spectrometry (HPLC–HRMS) for tracking aldehydes in both suspect screening and untargeted methods in fecal water, representing the aqueous environment of colon epithelial cells. This original approach is based on the introduction of a characteristic isotopic labeling by selective derivatization of the carbonyl function using a brominated reagent. Following a metabolomics workflow, the developed methodology was applied to the characterization of aldehyde compounds formed by lipid peroxidation in rats fed two different diets differentially prone to lipoperoxidation. Derivatized aldehydes were first selectively detected on the basis of their isotopic pattern, then annotated and finally identified by tandem mass spectrometry. This original approach allowed us to evidence the occurrence of expected aldehydes according to their fatty acid precursors in the diet, and to characterize other aldehydes differentiating the different diets.

Enzyme specific isotope effects of the Nap and Nar nitrate reductases

Dissimilatory nitrate reduction (DNR) to nitrite is the first step in denitrification, the main process through which bioavailable nitrogen is removed from ecosystems. DNR fractionates the stable isotopes of nitrogen (14N, 15N) and oxygen (16O, 18O) and thus imparts an isotopic signature on residual pools of nitrate in many environments. Data on the relationship between the resulting isotopic pattern in oxygen versus nitrogen isotopes (18ε / 15ε) suggests systematic differences exist between marine and terrestrial ecosystems that are not fully understood. DNR can be catalyzed by both cytosolic (Nar) and periplasmic (Nap) nitrate reductases, and previous work has revealed differences in their 18ε / 15ε isotopic signatures. In this study, we thus examine the 18ε / 15ε of six different nitrate-reducing microorganisms that encode Nar, Nap or both enzymes, as well gene deletion mutants of the enzymes’ catalytic subunits (NarG and NapA) to test the hypothesis that enzymatic differences alone could explain the environmental observations. We find that the distribution of the 18ε / 15ε fractionation ratios of all examined nitrate reductases form two distinct, non-overlapping peaks centered around a 18ε / 15ε proportionality of 0.55 and a 18ε / 15ε proportionality of 0.91, respectively. All Nap reductases studied to date cluster around the lower proportionality (0.55) and none exceed a 18ε / 15ε proportionality of 0.68. Almost all Nar reductases, on the contrary, cluster tightly around the higher proportionality (0.91) with no values below a 18ε / 15ε proportionality of 0.84 with the notable exception of the Nar reductases from the genus Bacillus which fall around 0.62 and thus closely resemble the isotopic fingerprints of the Nap reductases. Our findings confirm the existence of two remarkably distinct isotopic end-members in the dissimilatory nitrate reductases that could indeed explain differences in coupled N and O isotope fractionation between marine and terrestrial systems, and almost but not fully match reductase phylogeny.

Development and Application of High-Precision Algorithm for Non-Target Identification of Organohalogens Based on Ultrahigh-Resolution Mass Spectrometry

The brominated and/or chlorinated organic compounds (referred to as organohalogens) are frequently detected in natural and engineered environments. However, the ultrahigh resolution mass spectrometry (UHR-MS)-based non-target identification of the organohalogens remains challenging due to the presence of vast number of halogenated and non-halogenated organic molecules in the same aqueous sample. In this study, a new algorithm, namely NOMDBP Code, was developed, based on natural organic matter (NOM) chemistry, to simultaneously identify organohalogens and non-organohalogens from the UHR-MS spectra of natural and engineered waters. In addition to isotopic pattern extraction, for the first time, three optional filter rules (namely selection of minimum non oxygen heteroatoms, inspection of newly formed halogenated disinfection byproducts [X23 DBPs] and precursors) were incorporated in our code, which can accurately identify DBPs associated peaks and further elucidate the X-DBPs generation and transformation mechanisms. The formulae assignment rate against previously reported 2,815 unique organohalogens and their 11,583 isotopologues was determined to be >97%. Application of our algorithm to disinfected NOM indicated that oxygen-containing X-DBPs species accounted for a majority of X-DBPs. Further, brominated X-DBPs (Br-DBPs) during disinfection process were characterized by higher degree of unsaturation compared to chlorinated X-DBPs (Cl-DBPs). Our algorithm also suggested that, in addition to electrophilic substitution and electrophilic addition reactions, the decomposition/transformation is another important mechanism in Br-DBPs formation. Results of this study highlight the superior potential of this code to efficiently detect yet- unknown organohalogens (including organohalogens with non-oxygen heteroatoms) in a non-target manner and identify their generation mechanism during the disinfection process

Development and Application of High-Precision Algorithm for Non-Target Identification of Organohalogens Based on Ultrahigh-Resolution Mass Spectrometry

The brominated and/or chlorinated organic compounds (referred to as organohalogens) are frequently detected in natural and engineered environments. However, the ultrahigh resolution mass spectrometry (UHR-MS)-based non-target identification of the organohalogens remains challenging due to the presence of vast number of halogenated and non-halogenated organic molecules in the same aqueous sample. In this study, a new algorithm, namely NOMDBP Code, was developed, based on natural organic matter (NOM) chemistry, to simultaneously identify organohalogens and non-organohalogens from the UHR-MS spectra of natural and engineered waters. In addition to isotopic pattern extraction, for the first time, three optional filter rules (namely selection of minimum non oxygen heteroatoms, inspection of newly formed halogenated disinfection byproducts [X23 DBPs] and precursors) were incorporated in our code, which can accurately identify DBPs associated peaks and further elucidate the X-DBPs generation and transformation mechanisms. The formulae assignment rate against previously reported 2,815 unique organohalogens and their 11,583 isotopologues was determined to be >97%. Application of our algorithm to disinfected NOM indicated that oxygen-containing X-DBPs species accounted for a majority of X-DBPs. Further, brominated X-DBPs (Br-DBPs) during disinfection process were characterized by higher degree of unsaturation compared to chlorinated X-DBPs (Cl-DBPs). Our algorithm also suggested that, in addition to electrophilic substitution and electrophilic addition reactions, the decomposition/transformation is another important mechanism in Br-DBPs formation. Results of this study highlight the superior potential of this code to efficiently detect yet- unknown organohalogens (including organohalogens with non-oxygen heteroatoms) in a non-target manner and identify their generation mechanism during the disinfection process

Isotopic Composition of Abiogenic Gas Produced in Closed-System Fischer-Tropsch Synthesis: Implications for the Origins of the Deep Songliao Basin Gases in China

In this study, closed-system Fischer-Tropsch synthesis was conducted at 380°C and 30 MPa for 72 h with magnetite as a catalyst. The isotopic composition of the closed-system Fischer-Tropsch synthesis gas and the composition of known abiogenic gas were systematically studied, and the deep Songliao Basin gas was also investigated. The results show that closed-system Fischer-Tropsch synthesis of gaseous hydrocarbon isotopes exhibits a partial reverse order, which includes the reverse order of methane and ethane such as δ13C-C1>δ13C-C2<δ13C-C3 and δ2H-C1>δ2H-C2<δ2H-C3. Furthermore, experimental data on the control of NaBH4 content indicates that the carbon isotopes demonstrate a reverse order on condition that the H2/CO2 (mole ratio) is equal to or greater than 4.0; meanwhile, the hydrogen isotopes show a normal order. The deep Songliao Basin hydrocarbon gas component is similar to thermogenic gas and has a trend of a transition to oceanic hydrothermal system abiogenic gas. In addition, the deep Songliao Basin gas isotopic pattern is different from both Lost City and Kidd Creek where the deep Basin gas carbon isotopic pattern has a reverse order, and the hydrogen isotopic pattern has a normal order. Therefore, the deep Basin gas might be a mixture of the oil-type gas and the coal-formed gas, which could be the cause of the isotopic reverse.

GC-MS fragmentation patterns of sprayed endosulfan and its sulphate metabolite in samples of Theobroma cacao L from a field kinetic study

Most environmental analytical methods for the determination of organochlorine pesticides (OCPs) are multi-residual with other organic compounds co-extracted and co-eluted. This has been observed in GC spectra using classical detectors like electron-capture detector (ECD) even after appropriate clean-up. This limitation could be resolved by using GC-MS methods which are more specific and selective. Thus, a commercial-grade endosulfan treated Theobroma cacao plantation was sampled. Representative samples comprising leaves, stem bark and pulp were obtained between 0.5 h and 60 days after treatment. Samples were analyzed for residual parent endosulfan ( α- and β-isomers) as well as the metabolite endosulfan sulphate using an ion trap GC-MS. The retention times and chromatogram peaks obtained for various endosulfan were identified and compared with reference standards, and confirmed with National Institute of Standards and Technology library. Results showed that the molecular ion at m/z 407 was exhibited by α- and β-endosulfan, representing the parent molecular ion M+• ([C9H6Cl6SO3]+•). The α-isomer was more thermally stable, hence exhibited more relative abundance. Other predominant peaks were 339, 307, 277, 265, 243, 241, 207, 195, 160, 159, 99 and 75 m/z. The peak at m/z 159 was the base molecular ion. For endosulfan sulphate, the peak at m/z 422 corresponded to parent molecular ion (M+•), while m/z 424 was due to isotopic pattern characteristic of the chlorine atom. The peaks at 387, 357, 289, 272, 229, 206, 170, and 120 m/z were characteristic for the sulphate metabolite. The m/z peak at 272 was the base molecular ion, while m/z 143 may be due to metabolite diol and lactone. These results showed that the various endosulfan species can be identified and confirmed simultaneously using a GC-MS.

New Photodegradation Products of the Fungicide Fluopyram: Structural Elucidation and Mechanism Identification

Identifying the fate of agrochemicals is important to understand their potential risk for living organisms. We report here new photodegradation products (PPs) of the fungicide fluopyram. The PPs were produced by irradiating a fluopyram standard in 0.1% acetonitrile aqueous media by a 150-W medium pressure Hg-lamp that emits wavelengths between 200–280 nm. The structural elucidation of PPs was achieved by combining the retention time, isotopic pattern, targeted fragmentation, and accurate mass measurements using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and high resolution-MS (HRMS). In addition to previously known PPs, seven new PPs of fluopyram were identified in this work: mainly dihydroxyl and hydroxylimide fluopyram as well as mono, di, and trihydroxyl lactam. Additionally, two PPs were found to be formed by rearrangement after the loss of H2C=CH2. Hence, the results of the work contribute to extending the current knowledge regarding the photoinduced fate of agrochemicals, and fluopyram in particular.