Search Results (627)

Functional foods like flax (Linum usitatissimum L.) are rich sources of specialized metabolites that contribute to their nutritional and health-promoting properties. Understanding the biosynthesis of these compounds is essential for improving their quality and potential applications. However, dissecting complex metabolic networks in plants remains challenging due to the dynamic nature and interconnectedness of biosynthetic pathways. In this study, we present a synergistic approach combining stable isotopic labeling (SIL), Candidate Substrate–Product Pair (CSPP) networks, and a time-course study with high temporal resolution to reveal the biosynthetic fluxes shaping phenylpropanoid metabolism in young flax seedlings. By feeding the seedlings with ¹³C₃-p-coumaric acid and isolating isotopically labeled metabolization products prior to the construction of CSPP networks, the biochemical validity of the connections in the network was supported by SIL, independent of spectral similarity or abundance correlation. This method, in combination with multistage mass spectrometry (MSⁿ), allowed confident structural proposals of lignans, neolignans, and hydroxycinnamic acid conjugates, including the presence of newly identified chicoric acid and related tartaric acid esters in flax. High-resolution time-course analyses revealed successive waves of metabolite formation, providing insights into distinct biosynthetic fluxes toward lignans and early lignification intermediates. No evidence was found here for the involvement of chlorogenic or caftaric acid intermediates in chicoric acid biosynthesis in flax, as has been described in other species. Instead, our findings suggest that in flax seedlings, chicoric acid is synthesized through successive hydroxylation steps of p-coumaroyl tartaric acid esters. This work demonstrates the power of combining SIL and CSPP strategies to uncover novel metabolic routes and highlights the nutritional potential of flax sprouts rich in chicoric acid. Full article

Ammonium (NH₄⁺) toxicity impairs plant growth, but nitrate (NO₃⁻) can mitigate this effect through unresolved mechanisms. Using leaf δ¹³C values (photosynthetic capacity) and a bidirectional ¹⁵N tracer (NH₄⁺ assimilation efficiency and source utilization), this study investigated these mechanisms in 35-day-old Orychophragmus violaceus plantlets grown in modified Murashige and Skoog media under varying NH₄⁺:NO₃⁻ ratios. ¹⁵N isotope fractionation during NH₄⁺ (same fixed 20 mM NH₄Cl) assimilation decreased with increasing NO₃⁻ supply (10, 20, and 40 mM NaNO₃). Under 20 mM NH₄⁺ (δ¹⁵N = −2.64‰) at two ¹⁵NO₃⁻-labels (δ¹⁵N-NO₃⁻ = 8.08‰, low ¹⁵N, L) and (δ¹⁵N-NO₃⁻ = 22.67‰, high ¹⁵N, H), increasing NO₃⁻ concentrations enhanced NO₃⁻ assimilation, alleviating acidic stress from NH₄⁺ and improving photosynthesis. Higher NO₃⁻ levels also increased NH₄⁺ utilization efficiency, reducing futile NH₄⁺ cycling and decreasing associated ¹⁵N fractionation during assimilation. Our results demonstrate that NO₃⁻ alleviates NH₄⁺ toxicity primarily by enhancing photosynthetic performance and optimizing NH₄⁺ utilization efficiency. Full article

The priming effect (PE), a microbially mediated process, critically regulates the balance between carbon sequestration and mineralization. This study used soils from different soil depths (0–20 cm, 20–40 cm, and 40–60 cm) under Picea schrenkiana forest in the Tianshan Mountains as the research object. An indoor incubation experiment was conducted by adding three concentrations (1% SOC, 2% SOC, and 3% SOC) of ¹³C-labelled glucose. We applied ¹³C isotope probe-phospholipid fatty acid (PLFA-SIP) technology to investigate the influence of readily labile organic carbon inputs on soil priming effect (PE), microbial community shifts at various depths, and the mechanisms underlying soil PE. The results indicated that the addition of ¹³C-labeled glucose accelerated the mineralization of soil organic carbon (SOC); CO₂ emissions were highest in the 0–20 cm soil layer and decreased trend with increasing soil depth, with significant differences observed across different soil layers (p < 0.05). Soil depth had a positive direct effect on the cumulative priming effect (CPE); however, it showed negative indirect effects through physico-chemical properties and microbial biomass. The CPE of the 0–20 cm soil layer was significantly positively correlated with ¹³C-Gram-positive bacteria, ¹³C-Gram-negative bacteria, and ¹³C-actinomycetes. The CPE of the 20–40 cm and 40–60 cm soil layers exhibited a significant positive correlation with cumulative mineralization (CM) and microbial biomass carbon (MBC). Glucose addition had the largest and most significant positive effect on the CPE. Glucose addition positively affected PLFAs and particularly microbial biomass. This study provides valuable insights into the dynamics of soil carbon pools at varying depths following glucose application, advancing the understanding of forest soil carbon sequestration. Full article

Background: Liquid chromatography-mass spectrometry (LC-MS), widely used in metabolomics, is often limited by low ionization efficiency and ion suppression, which reduce overall metabolite detectability and quantification accuracy. To address these challenges, chemical isotope labeling (CIL) LC-MS has emerged as a powerful approach, offering high sensitivity, accurate quantification, and broad metabolome coverage. This method enables comprehensive profiling by targeting multiple submetabolomes. Specifically, amine-/phenol- and hydroxyl-containing metabolites are labeled using dansyl chloride under distinct reaction conditions. While this strategy provides extensive coverage, the sequential analysis of each submetabolome reduces throughput. To overcome this limitation, we propose a two-channel mixing strategy to improve analytical efficiency. Methods: In this approach, samples labeled separately for the amine/phenol and hydroxyl submetabolomes are combined prior to LC-MS analysis, leveraging the common use of dansyl chloride as the labeling reagent. This integration effectively doubles throughput by reducing LC-MS runtime and associated costs. The method was evaluated using human urine and serum samples, focusing on peak pair detectability and metabolite identification. A proof-of-concept study was also conducted to assess the approach’s applicability in putative biomarker discovery. Results: Results demonstrate that the two-channel mixing strategy enhances throughput while maintaining analytical robustness. Conclusions: This method is particularly suitable for large-scale studies that require rapid sample processing, where high efficiency is essential. Full article

Alkaline earth elements have emerged as crucial electronic modifiers for regulating active sites in catalytic systems, yet the influence of metal–support interactions (MSIs) between alkaline earth compounds and active metals remains insufficiently understood. This study systematically investigated Pt nanoparticles supported on alkaline earth carbonates (Pt/MCO₃, M = Mg, Ca, Ba) for low-temperature propylene combustion. The Pt/BaCO₃ catalyst exhibited outstanding performance, achieving complete propylene conversion at 192 °C, significantly lower than Pt/MgCO₃ (247 °C) and Pt/CaCO₃ (282 °C). The enhanced activity stemmed from distinct MSI effects among the supports, with Pt/BaCO₃ showing the poorest electron enrichment and lowest propylene adsorption energy. Through kinetic analyses, ¹⁸O₂ isotope labeling, and comprehensive characterization, the reaction was confirmed to follow the Mars–van Krevelen (MvK) mechanism. Pt/BaCO₃ achieves an optimal balance between propylene and oxygen adsorption, a critical factor underlying its superior activity. Full article

Spirocyclic architectures, which feature two rings sharing a single atom, are common in natural products and exhibit beneficial biological and material properties. Due to the significance of these architectures, biocatalytic dearomative spirocyclization has recently emerged as a powerful approach for constructing three-dimensional spirocyclic frameworks under mild, sustainable conditions and with exquisite stereocontrol. This review surveys the latest advances in biocatalyzed spirocyclization of all-carbon arenes (phenols and benzenes), aza-aromatics (indoles and pyrroles), and oxa-aromatics (furans). We highlight cytochrome P450s, flavin-dependent monooxygenases, multicopper oxidases, and novel metalloenzyme platforms that effect regio- and stereoselective oxidative coupling, epoxidation/semi-pinacol rearrangement, and radical-mediated cyclization to produce diverse spirocycles. Mechanistic insights gleaned from structural, computational, and isotope-labeling studies are discussed where necessary to help the readers further understand the reported reactions. Collectively, these examples demonstrate the transformative potential of biocatalysis to streamline access to spirocyclic scaffolds that are challenging to prepare through traditional methods, underscoring biocatalysis as a transformative tool for synthesizing pharmaceutically relevant spiroscaffolds while adhering to green chemistry paradigms to ultimately contribute to a cleaner and more sustainable future. Full article

The widespread environmental dissemination of imidazole compounds necessitates robust analytical monitoring tools. This study developed a novel isotope-labeled surrogate-based high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) method for the simultaneous determination of 21 imidazoles in water, sediment, and soil. The optimized SPE protocol using Oasis HLB cartridges achieved high recoveries, with chromatographic separation completed in 25 min. Six isotope-labeled standards effectively corrected matrix effects (−57% to 8%), yielding MQLs < 1.0 ng·L⁻¹ (water) and <1.0 μg·kg⁻¹ (sediment/soil). Validation confirmed linearity (R² > 0.995), accuracy (60–120% recovery for 20/21 analytes), and precision (relative standard deviation, RSD < 15%). Its application in Jiulong River revealed significant contamination, detecting eight imidazoles in both water (up to 49.29 ng·L⁻¹) and sediment (up to 24.01 μg·kg⁻¹). This standardized tool enables routine monitoring of pharmaceutical residues across environmental compartments, supporting regulatory frameworks. Full article

Background: Hydroxyl and amino compounds play a significant role in defining the flavor and quality of sauce-flavor Baijiu, yet their comprehensive analysis remains challenging due to limitations in detection sensitivity. In this study, we developed a novel bromine isotope labeling approach combined with ultra-high performance liquid chromatography–high-resolution mass spectrometry (UHPLC-HRMS) to achieve high-coverage profiling of these compounds in sauce-flavor Baijiu. Methods: The method employs 5-bromonicotinoyl chloride (BrNC) for rapid (30 s) and mild (room temperature) labeling of hydroxyl and amino functional groups, utilizing bromine’s natural isotopic pattern (Δm/z = 1.998 Da) for efficient screening. Annotation was performed hierarchically at five confidence levels by integrating retention time, accurate mass, and MS/MS spectra. Results: A total of 309 hydroxyl and amino compounds, including flavor substances (e.g., tyrosol and phenethyl alcohol) and bioactive compounds (e.g., 3-phenyllactic acid), were identified in sauce-flavor Baijiu. The method exhibited excellent analytical performance, with wide linearity (1–4 orders of magnitude), precision (RSD < 18.3%), and stability (RSD < 15% over 48 h). When applied to sauce-flavor Baijiu samples of different grades, distinct compositional patterns were observed: premium-grade products showed greater metabolite diversity and higher contents of bioactive compounds, whereas lower-grade samples exhibited elevated concentrations of acidic flavor compounds. Conclusions: These results demonstrate that the established method is efficient for the comprehensive analysis of hydroxyl and amino compounds in complex food matrices. The findings provide valuable insights for quality control and flavor modulation in sauce-flavor Baijiu production. Full article

Calpain-5/CAPN5 is a calcium-activated, non-lysosomal cysteine (thiol) protease. The substrate repertoire of CAPN5 is not known. Calpains catalyze limited proteolysis of their substrates, generating neo-N-termini that correspond to internal residues of their nascent substrate proteins. To identify such neo-N-termini generated by CAPN5, we employed an N-terminomics approach called TAILS (Terminal amine isotopic labeling of substrates) to quantitatively compare the N-terminal peptides detected in parental and CAPN5-deficient SH-SY5Y neuroblastoma cells. Thirty neo-N-termini corresponding to 29 protein groups and 24 unique proteins were detected to be depleted in the CAPN5^−/− cells. A subset of the identified putative substrates was further studied with CAPN5 co-immunoprecipitation, in vitro calcium-induced CAPN5 proteolysis assay, and their cellular fragmentation patterns were compared in parental and CAPN5-deficient SH-SY5Y cells. Here, we provide evidence for CAPN5-mediated proteolysis of the synaptic proteins DLGAP4, IQSEC1 and MPDZ, the neurodegeneration-related EWS, hnRNPU, TFG and UGP2, the DNA replication regulator MCM3, and the neuronal differentiation regulator LMTK1. Our data provide new relevance for neovascular inflammatory vitreoretinopathy (NIV), a progressive eye disease caused by pathogenic mutations in CAPN5. Data are available via ProteomeXchange with identifier PXD064313. Full article

Positron emission tomography (PET) imaging targeting glypican-3 (GPC3) holds promise for improving the detection and characterization of hepatocellular carcinoma (HCC). Preclinical and early clinical studies have largely utilized high-molecular-weight antibodies radiolabeled with isotopes such as ⁸⁹Zr and ¹²⁴I, demonstrating high affinity and tumor uptake but suffering from prolonged circulation times and suboptimal signal-to-background ratios. To address these limitations, interest has shifted toward low-molecular-weight vectors—synthetic peptides and small antibody fragments—labeled with shorter-lived radionuclides (e.g., ⁶⁸Ga and ¹⁸F) to enable rapid pharmacokinetics and same-day imaging protocols. Emerging platforms such as affibodies and aptamers offer further advantages in target affinity and reduced immunogenicity. However, clinical translation requires rigorous validation: larger, histologically confirmed cohorts, head-to-head comparison with CT/MRI, and correlation with hard clinical endpoints. Moreover, leveraging GPC3 expression as a biomarker could guarantee a deeper knowledge of tumor biology—differentiation grade and vascular invasion risk—and guide theranostic strategies. While β-emitters (⁹⁰Y, ¹⁷⁷Lu) have been explored for GPC3-directed therapy, their efficacy is influenced by oxygenation and cell-cycle status, whereas α-emitters (²²⁵Ac) may overcome these constraints, albeit with challenges in radionuclide selection and daughter nuclide management. Finally, dual-targeting probes combining GPC3 and prostate-specific membrane antigen (PSMA) have demonstrated superior uptake and retention in murine models, suggesting a versatile approach for future clinical diagnostics and therapy planning. Full article

Nitrogen deposition is a critical driver of plant-soil interactions in forest ecosystems. However, the species-specific coordination of nitrogen uptake and carbon assimilation—traced using ¹⁵N- and ¹³C-labeled compounds—under varying nitrogen forms, depths, and time points remains poorly understood. We conducted a dual-isotope (¹⁵NH₄Cl, K¹⁵NO₃, and Na₂¹³CO₃) labeling experiment in a temperate secondary forest to investigate nutrient uptake and carbon assimilation in three understory species—Carex siderosticta, Maianthemum bifolium, and Oxalis acetosella—across three nitrogen treatments (control, low N, and high N), two soil depths (0–5 cm and 5–15 cm), and two post-labeling time points (24 h and 72 h). We quantified ¹⁵N uptake and ¹³C assimilation in above- and belowground plant tissues, as well as ¹⁵N and ¹³C retention in soils. C. siderosticta exhibited the highest total ¹⁵N uptake (2.2–6.9 μg N m⁻² aboveground; 1.4–4.1 μg N m⁻² belowground) and ¹³C assimilation (58.4–111.2 mg C m⁻² aboveground; 17.6–39.2 mg C m⁻² belowground) under high ammonium at 72 h. High nitrogen input significantly enhanced the coupling between plant biomass and nutrient assimilation (R² > 0.9), and increased ¹⁵N-TN and ¹³C-SOC retention in the surface soil layer (13,200–17,400 μg N kg⁻¹; 30,000–44,000 μg C kg⁻¹). Multifactorial analysis revealed significant interactions among nitrogen treatment, form, depth, and time. These findings demonstrate that ammonium-based enrichment promotes nutrient acquisition and carbon assimilation in responsive species and enhances surface soil C—N retention, highlighting the integrative effects of nitrogen form, species traits, and spatial–temporal dynamics on forest biogeochemistry. Full article

Background: Radiolabeled compounds can serve as diagnostic or therapeutic agents depending on the characteristics of the isotopes used. IMPY (6-iodo-2-(4′-dimethylamino)-phenyl-imidazo[1,2-a]pyridine) is a lipophilic derivative of thioflavin-T, designed to function as a tracer when labeled with radioactive iodine. While it has been primarily studied for imaging applications, its potential therapeutic effects when labeled with iodine-125 (¹²⁵I) remain to be explored. Methods: In this study, IMPY was synthesized and labeled with ¹²⁵I for therapeutic purposes. Three different labeling methods were employed: isotope exchange reaction, redox reaction, and the Iodogen technique. The radiochemical yield of each method was determined to identify the most effective approach. Additionally, the effects of ¹²⁵I-IMPY on neuroblastoma cells were evaluated by assessing its toxicity and cellular uptake. Results: The radiochemical yields for the isotope exchange reaction, redox reaction, and Iodogen technique were found to be 0.96%, 10.74%, and 96.52%, respectively. The Iodogen technique exhibited the highest yield, exceeding 90% even after 48 h, making it the most efficient method. Furthermore, the impact of ¹²⁵I-IMPY on neuroblastoma cells was analyzed, revealing significant cellular uptake and potential therapeutic effects. Conclusions: This study demonstrated that the Iodogen technique is the most effective method for labeling IMPY with ¹²⁵I. The high labeling efficiency and observed cellular effects suggest that ¹²⁵I-IMPY could be considered not only as a tracer but also as a potential therapeutic agent for neuroblastoma. Further studies are needed to explore its full therapeutic potential and mechanism of action. Full article

In this study, we use coal as a carbon source from Zhangjiamao and doped with different nitrogen sources for co-pyrolysis. Nitrogen-rich tar was successfully prepared, and the content and variety were also increased. From the elemental analysis, the nitrogen content of all the tars was significantly enhanced, among which the nitrogen content of the tars after co-pyrolysis with melamine was enhanced by 5.21%, and the nitrogen content of coke was enhanced by 10.87%. According to the GC/MS results, it was found that the nitrogen compounds in the tar after full pyrolysis were richer and more abundant than those in Py-GC/MS. For the free radical reaction, the reaction process is extremely rapid, and the ¹⁵N substitution product after isotope labeling was successfully captured by adding ¹⁵NH₄Cl for isotope labeling, which can be more intuitively and accurately illustrated from the m/z change. Among them, 26 nitrogen-containing compounds were screened out, which accounted for 66.28% of the content, and they were categorized. It was found that the five-membered nitrogen heterocycles were the most abundant, accounting for 34.88%. In addition, five other nitrogen-containing compounds containing different functional groups and the tar from the co-pyrolysis of tar-rich coal were also analyzed by GC/MS, among which the tar from melamine had the highest content of nitrogen-containing compounds, with 70.48%. Finally, the functional groups of nitrogen-containing compounds were further analyzed by XPS and FTIR, and the results were consistent with those of GC/MS analysis. In this paper, nitrogen-rich tar was prepared by co-pyrolysis of tar-rich coal and nitrogen compounds. This achievement provides a valuable reference for the high-value utilization of coal tar. Full article

Vineyard soils are frequently contaminated with copper due to the use of Cu fungicides to prevent downy mildew. This study investigated the effects of an aerated compost tea (ACT) made from grape pomace and animal manure on the downward transfer of Cu and on the accumulation of Cu in plants in a sandy loam vineyard soil. Crimson clover and pot marigold were grown in a 40 cm soil column with Cu supplied to the surface at loadings representative of those applied in European vineyards, plus additions of ACT. A source of Cu enriched in the stable isotope ⁶⁵Cu was used to distinguish freshly added Cu (fresh Cu) from Cu already present in the soil (aged Cu). ACT increased the concentration of soluble humic substances (SHS) in pore water in the top 7.5 cm of the column, and increased the concentration of Cu, Al, and Fe in pore water in proportion to the concentration of SHS. The transfer of fresh Cu to deeper soil was limited to the top 5 cm, even after the addition of ACT, although fresh Cu reacted slightly more to ACT than aged Cu. ACT had no effect on Cu phytoextraction but increased the concentration of Cu in roots by almost twofold. Relatively more fresh Cu was transferred to plants than aged Cu, primarily due to its preferential accumulation on the surface. The risk associated with the use of ACT on vineyard soils is not that of promoting the downward transfer of Cu, but rather of increasing Cu availability to plants and likely to other living organisms in the topsoil. Full article

Polysaccharides are important biomolecules, which play a key role in biological, medical, and industrial processes due to their diverse structures and important functions. This paper looks into the significant progress made in the structural and functional analysis of polysaccharides by nuclear magnetic resonance (NMR) spectroscopy. NMR spectroscopy, including solution-state and solid-state technology, has revolutionized this field through detailed molecular insights into the structure, conformation, and dynamics of polysaccharides at the molecular level. There have been some important historical breakthroughs in 1D and 2D NMR, which have led to modern methods like multidimensional NMR and nuclear dynamic polarization (DNP). These modern methods offer a high level of resolution and sensitivity and have made it easier to come up with innovative applications. The applications range from the structural elucidation of microbial and plant structures of polysaccharides to improving food texture, developing therapies, and creating sustainable materials. Despite challenges such as signal overlap and limited sensitivity, emerging solutions are making significant progress. Computational modeling, isotope labeling, and integrated methods that combine complementary technologies are driving the boundaries of polysaccharide research. This review demonstrates the transformative role of NMR in revealing the complexity of polysaccharides and its potential to promote future discoveries and new ideas in the dynamic field. Full article