2D NMR Research Articles

1H NMR Trajectories for Analyzing the Growth and Purification of 2D Polyaramids.

The recent synthesis of two-dimensional (2D) polyaramid (PA) polymers has shown that they exhibit remarkable mechanical and gas barrier properties. However, new methods are necessary to enable the efficient measurement of discoidal sizes of 2D polyaramids and tracking of their growth over time during reaction. Herein, we employ 1H NMR peak analysis of the aromatic and proton end group regions to characterize 2D-PA growth from monomeric precursors, alongside purification using two-stage filtration and washing steps. The ratio of aromatic to end group protons yields the molecular weight and discoidal size, while the skewness of the NMR peaks in the aromatic region provides a relative weighting between dendritic, small, and larger polycyclic domains. These two metrics delineate a distinct two-dimensional 1H NMR trajectory that we show to be valuable for analyzing the results of differing synthetic and processing conditions. Theoretical bounds corresponding to dendrimer and polycyclic limits as a function of repeat unit size are derived as an aid in trajectory analysis. These results establish an analytical framework for evaluating 2D-PA and its variants, facilitating the exploration of the synthetic landscape.

Bisabolane Sesquiterpenes with Anti-Chlamydial Activity Isolated from Ligularia narynensis

Chlamydia are obligate intracellular bacterial pathogens affecting humans and animals, causing miscarriage, stillbirth, or weak fetuses in the late stages of pregnancy of goats and sheep. Because there is no commercial vaccine for chlamydia in animals, drug treatment has become the most effective curative method. Natural products, also known as secondary metabolites, are becoming one of the main sources used in new drug development because of their structural diversity and biodiversity. In natural products, plant sources play a major role in the development process of new drugs. In this study, five undescribed highly oxygenated bisabolane sesquiterpenes (Pararubin W, Pararubin X, Pararubin Y., Pararubin Z, and Pararubin AA) were isolated from whole plants of Ligularia narynensis. Their chemical structures were determined via analyses of HRESIMS, IR, 1D, and 2D NMR data, along with the assignment of their relative configurations. These compounds were tested for their anti-chlamydial activity. The results show that compounds 1 and 5 inhibited the growth of Chlamydia abortus in host cells in a dose-dependent manner.

Deuterated oxazines are bright near-infrared fluorophores for mitochondrial imaging and single molecule spectroscopy.

Bright near-infrared fluorophores are in demand for microscopy. We showcase a deuterated oxazine being 23% brighter vs. ATTO700. With a longer lifetime of 1.85 nanoseconds, we find the best-in-class SulfoOxazine700-d10 to stain mitochondria for confocal microscopy, and demonstrate unaffected diffusion properties in single molecule fluorescence correlation spectroscopy.

Water-Soluble Curcumin Derivatives Including Aza-Crown Ether Macrocycles as Enhancers of Their Cytotoxic Activity.

The synthesis of three novel curcumin derivative compounds, featuring aza-crown ether macrocycles of various sizes (aza-12-crown-4, aza-15-crown-5, and aza-18-crown-6), is described. The incorporation of these aza-crown macrocycles significantly enhances their water solubility, positioning them as groundbreaking instances of curcumin derivatives that are fully soluble in aqueous environments. These curcumin ligands (L1, L2, and L3) were then reacted with zinc acetate to afford the coordination metal complexes (L1-Zn, L2-Zn, and L3-Zn). Comprehensive characterization of all compounds was achieved using various analytical techniques, including 1D and 2D NMR spectroscopy, ATR-FTIR spectroscopy, mass spectrometry (ESI+), elemental analysis and UV-Vis spectroscopy. The in vitro cytotoxic activity of both, ligands and complexes were evaluated on three human cancer cell lines (U-251, MCF-7, and SK-LU-1). Compared to conventional curcumin, these compounds demonstrated improved antiproliferative potential. Additionally, a wound healing assay was conducted to assess their antimigration properties. The obtained results suggest that these modifications to the curcumin structure represent a promising approach for developing therapeutic agents with enhanced cytotoxic properties.

Gossypin-Loaded Ethosome Gel for Cutaneous Administration: A Preliminary Study on Melanoma Cells

A preformulative study was conducted to produce and characterize ethosomes for the transdermal delivery of gossypin. This plant-derived compound possesses many pharmacological properties, including antitumoral potential. Ethosome dispersions were designed as transdermal delivery systems for gossypin, employing two different production procedures. The evaluation of vesicle size distribution by photon correlation spectroscopy, morphology by cryogenic transmission electron microscopy, and gossypin entrapment capacity, as well as in vitro release and permeation by vertical diffusion cells, enabled us to select a production strategy based on the injection of a phosphatidylcholine ethanolic solution in water. Indeed, vesicles prepared by this method were almost unilamellar and measured roughly 150 nm mean diameter while displaying an entrapment capacity higher than 94%. Moreover, vesicles prepared by the ethanol injection method enabled us to control gossypin release and to improve its permeation with respect to the solution of the drug. To obtain semi-solid forms suitable for cutaneous gossypin administration, ethosome dispersions were thickened with 0.5% w/w xanthan gum, selected by a spreadability test. These ethosome gels were then further characterized by small- and wide-angle X-ray scattering, while their antioxidant activity was demonstrated in vitro by a radical scavenging assay. Finally, in vitro biological studies were conducted on A375 melanoma cell lines. Namely, wound healing and cell migration assays confirmed the potential antitumoral effect of gossypin, especially when loaded in the selected ethosomal gel. The promising results suggest further investigation of the potential of gossypin-loaded ethosomal gel in the treatment of melanoma.

Jejupeptins A and B: Polyketide-Cyclic Peptide Hybrids with Anti-Corticosterone Activity from Streptomyces sp. KCB15JA151.

Two new polyketide-cyclic peptide hybrids jejupeptins A (1) and B (2), together with a known compound eurystatin B (3), were isolated from a culture of Streptomyces sp. KCB15JA151. The chemical structures of the compounds were elucidated using a combination of 1D and 2D NMR spectroscopy and DP4+ probability analyses. Comprehensive spectroscopic analysis revealed that 1 and 2 are the first examples of hybrid peptide-polyketides possessing an unprecedented 4-amino-3-hydroxymethyl-3-methyl-2-oxopentanoic acid moiety. A plausible biosynthetic pathway for these compounds was proposed. Biological evaluation demonstrated that compounds 1 and 2 exhibit protective effects against corticosterone-induced apoptosis and cellular oxidative stress without any associated cytotoxicity.

Intraparticle Diffusion Behavior of Rhodamine 6G in Single Silica Particle Revealed by Fluorescence Correlation Spectroscopy.

Understanding the intraparticle diffusion mechanism within single particles is crucial for processes such as chromatographic separation, drug delivery, and solid extraction. However, when the time required to reach distribution equilibrium is extremely short (on the order of several seconds), conventional kinetic detection methods pose significant challenges in observing intraparticle diffusion behavior. In this study, we employed fluorescence correlation spectroscopy (FCS)─a technique capable of detecting diffusion behavior at equilibrium─to investigate the intraparticle diffusion of rhodamine 6G (Rh6G) within single porous silica particles of varying pore sizes. The autocorrelation coefficients of Rh6G were fitted using two-component analysis, revealing faster and slower diffusion components associated with the pore diffusion, without and with adsorption/desorption, respectively-behaviors that were not observed by the kinetic method described by our previous study. Further analysis of the slower diffusion component was conducted using pore and surface diffusion models. Our findings indicate that pore diffusion is the primary diffusion mechanism for Rh6G within silica particles. Thus, we demonstrated that the intraparticle diffusion mechanism of Rh6G in silica particles can be elucidated using FCS measurements.

Derailment of the Biosynthesis via an Acid-Mediated Intramolecular Cyclo-rearrangement Leads to a Novel Cytochalasin Skeleton.

Cytochalasins are notable for their structural diversity and broad range of biological activities. The gene cluster responsible for the biosynthesis of cytochalasins bearing diverse polycycles was identified in Phomopsis sp. DHS-48. Characterization of the cluster indicated that only the 5/6/11-tricycle can be biosynthetically produced. The chemical space of cytochalasins was expanded by acid-mediated intramolecular cyclo-rearrangement of the 5/6/11-tricycle, and three new cytochalasins, phomoparagins D-F (13, 17, and 18, respectively), with diversified polycycles were obtained, among which compound 13 featured an unprecedented 5/6/5/7/6-pentacycle. Their structures and absolute configurations were established by spectroscopic analysis (1D, 2D NMR), electronic circular dichroism calculations, and a single-crystal X-ray diffraction experiment. The compounds inhibited the growth of HeLa and RKO cell lines with IC50 values ranging from 0.8 to 47.3 μM. Cytoskeleton staining experiments and molecular docking models revealed that compound 13 showed cytotoxicity by targeting F-actin.

Effect of Halogen Substitution on the Regioselective Deiodination of Thyroid Hormone Analogues by Deiodinase Mimics.

The deiodination of L-thyroxine (T4) by the three isoforms of selenium-containing iodothyronine deiodinases (DIOs) is an important process in the human body for the maintenance of the thyroid hormone homeostasis. Selenium compounds are known to mimic the function of DIO3 by catalyzing the 5-deiodination of T4 and a co-operative halogen and chalcogen bonding is responsible for their activity. The substitution of 4'-OH group of T4 by an electron withdrawing group not only alters the reactivity but also changes the regioselectivity of deiodination. However, the effect of other halogen atoms (F, Cl or Br) in place of iodine in T4 on the regioselectivity has not been explored. In this paper, we describe simple synthetic methodologies toward various halogen analogues of T4. We also show that 2D NMR spectroscopy can be used as a powerful tool to identify the location of various halogen substitutions on the phenolic or tyrosyl rings. The deiodination experiments with peri-substituted naphthalene diselenol reveal that the regioselectivity or specificity towards iodine atom is not altered upon introduction of more electronegative halogen atoms (F, Cl and Br), reinforcing the concept of halogen bonding in the deiodination reactions.

MDN-0057 to MDN-0060, a Family of Broad-Spectrum Antibiotics against Gram-Negative Pathogens Produced by Ophiosphaerella korrae.

A novel family of antibiotics, MDN-0057 to MDN-0060 (1-4), was isolated from liquid cultures of the fungus Ophiosphaerella korrae. These compounds incorporate two isocyanide groups in their complex structures that were elucidated by extensive spectroscopic analyses, including HRESIMS, and 1D and 2D NMR experiments. The relative configurations were determined by using J-based configuration analyses and the interpretation of key NOESY correlations consistent with the existence of a major conformation in solution. Mosher ester derivatization analysis allowed the establishment of their absolute configurations. All four compounds displayed in vitro antibacterial activity with a broad spectrum against Gram-negative pathogens.

Development of morphology-dependent nanoselenium carriers for enhancing biological activity and reducing plant stress.

Owing to their small size, morphology and release modification properties, nanopesticides are considered promising alternative strategies for enhancing biological activity and minimizing pesticide losses. In this study, we used a colloidal self-assembly method to develop a morphology-stable, regularly rod-shaped nanoselenium pesticide carrier (NSer), which was further modified with chitosan. After loading penthiopyrad (PEN), the biological activity of NSer@PEN and its impact on the physiological and biochemical processes of plants were further compared with those of spherical nanoselenium pesticides (NSes@PEN) and commercial materials (20 % PEN SC). The biological activities were quantified through the EC50 values, which revealed that NSer@PEN (0.71 mg/L) and NSes@PEN (1.09 mg/L) exhibited significantly greater activity against Colletotrichum orbiculare Arx compared to 20 % PEN SC (2.70 mg/L). Moreover, through further investigation into the impact of nanopesticides on plant root exudates, Fourier transform infrared spectroscopy (FTIR) and two-dimensional correlation spectroscopy (2D-COS) analysis revealed that the ketone CO bond exhibited the strongest binding affinity, and the CO bond of phenols contributed significantly to the binding of cucumber root exudates induced by NSer@PEN, resulting in a mild response of the plant. The morphology-dependent nanoselenium carriers developed in this work are expected to enhance biological activity and reduce plant stress caused by pesticides, tackling one of the application challenges of pesticides.

Synthesis of MnFe2O4-biochar with surficial grafting hydroxyl for the removal of Cd(II)-Pb(II)-Cu(II) pollutants: Competitive adsorption, application prospects and binding orders of functional groups.

A novel biochar material with magnetic modification by MnFe2O4 and surficial hydroxyl grafting (h-MFO-BC) was synthesized for capturing HMs (Cd, Pb and Cu) and their competition in composite systems was investigated. The modification of hydroxyl considerably improved the adsorption capacity of HMs. Chemisorption and monolayer and homogeneous reaction dominated adsorption processes. Moreover, a pronounced competitive adsorption effect between HMs was observed in composite systems. The order of selectivity by h-MFO-BC was Pb>Cu≫Cd. The distinction in the adsorption of HMs was related to different adsorption pathways and binding sequences of functional groups. Two-dimensional correlation spectroscopy revealed that Pb and Cu preferred to bind to the active sites (Mn/Fe-OH) on h-MFO-BC surface. Moreover, they could generate hydroxide precipitation more easily, which prevented further adsorption of Cd due to the occupation or coverage of binding sites and electrostatic repulsion. Furthermore, h-MFO-BC could be effectively regenerated and recycled and possessed fascinating performance in HMs removal from real water, indicating its potential for widespread applicability. This work provided a novel composite material for the treatment of HMs in wastewater or selective recovery of Pb and Cu and gave a new perspective on understanding the competition mechanisms between HMs on adsorbents.

The impact of the structural transformation mechanism of fulvic acid on redox capacity during composting with different biowastes.

Fulvic acid (FA) derived from composting functions can act as electron shuttle, facilitating and expediting the redox reaction during the composting process. However, limited research has been conducted on the redox capacity and structural transformation of FA during composting with different biowastes. The Fe (II) production quantity of the single S. oneidensis MR-1 (MR-1), MR-1 with FA derived from lignocellulose-rich and lignin-rich composting after 300h inoculation were up to 2.28, 3.67 and 2.52mmol/L, indicating the redox capacity of FA in lignocellulose-rich composts was stronger than that in lignin-rich composting. Furthermore, two-dimensional correlation spectroscopy revealed that FA derived from lignocellulose-rich composting exhibited a layer-by-layer structure, characterized by aromatic functional groups and other groups. In contrast, FA obtained from lignin-rich composting displayed an arrangement where the inner aromatic functional groups were obstructed by the methyl group. The structural equation model revealed that the high relative abundance and aromatic functional groups of FA derived from lignocellulose-rich composting process exert a direct influence on the reduction of Fe (III)-citrate, and the methyl groups of FA obtained from lignin-rich composting process also directly linked to the reduction of Fe (III)-citrate. The present study thus posited that the quantity of aromatic functional groups within outer of FA structure during composting constituted a crucial factor influencing its redox capacity. The findings offer novel insights into the transformation mechanism of FA derives from diverse biowastes and its redox characteristics, thereby providing significant guidance for the application of FA in mitigating environment pollutants.

Sesquiterpene lactones from Artemisia porrecta Krasch.ex Poljakov and their anti-inflammatory, cytotoxic and anti-vitiligo activities.

This experiment focused on investigating the chemical constituents of the plant Artemisia porrecta Krasch.ex Poljakov, resulting in the isolation and characterization of twenty-seven sesquiterpene lactones belonging to the eudesmanolides, germacronolides, melampolides, and heliangolide types. These compounds include ten undescribed sesquiterpene lactones, Porrectolide A-I, 11-epi-Deacetylherbolide D and seventeen known ones. All known compounds were isolated from this plant for the first time. Their structures were established by 1D, 2D NMR spectroscopic and HR-ESI-MS data, electronic circular dichroism, and single crystal X-ray diffraction analysis. Evaluations of the anti-inflammatory activity revealed that 6 had moderate anti-inflammatory activity on lipopolysaccharide-induced nitric oxide production in RAW264.7 cells with the IC50 value of 14.57±0.24 μM.

Investigation of All Disease-Relevant Lysine Acetylation Sites in α-Synuclein Enabled by Non-canonical Amino Acid Mutagenesis.

Aggregates of α-synuclein (αS) are hallmarks of synucleinopathies, including Parkinson's Disease (PD) and Multiple System Atrophy (MSA). We have recently shown that αS lysine acetylation in the soluble monomer pool varies between healthy controls, PD, and MSA patients. To study the effects of lysine acetylation at all disease-relevant sites of αS, we first compared production of acetylated αS through either native chemical ligation or non-canonical amino acid (ncAA) mutagenesis. Since yields were comparable, ncAA mutagenesis was deemed superior for scanning many acetylation sites. We expressed and purified 12 disease-relevant variants and studied their binding to membranes as well as their aggregation propensities, and found that acetylation of lysine 12, 43, and 80 had particularly strong effects. To understand the implications for acetylation of monomeric αS found in healthy cells, we performed NMR experiments to study protein conformation and fluorescence correlation spectroscopy experiments to quantify lipid binding. We also investigated the effects of acetylation at lysine 12, 43, and 80 on fibril seeding in neurons. Collectively, our biochemical and cell biological investigations indicated that acetylation of K 80 could inhibit aggregation without conferring negative effects on monomer function in healthy cells. Therefore, we studied the structures of fibrils with K 80 acetylation through cryo-electron microscopy to uncover the structural basis for these effects. Finally, we identified inhibition of HDAC8 as a way of potentially increasing acetylation at K 80 and other inhibitory sites for therapeutic benefit.

Theobroma mariae: Bioactive Compound-Rich Flowers

Edible flowers have gained attention as unconventional food sources due to their nutritional and functional properties. This study provides novel information on the chemical composition, cytotoxicity and antiproliferative effects of Theobroma mariae flowers. The objective of this paper was to identify bioactive compounds in its flowers using one- and two-dimensional nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS). The phenolic fraction of the flowers revealed bioactive compounds such as hyperoside, guaijaverin, astragalin, juglalin, and kaempferol. The results confirmed the potential of T. mariae flowers as a source of phenolic compounds, emphasizing their feasibility for possible applications in the development of functional foods. Moreover, the antiproliferative assay demonstrated that the phenolic fraction inhibits cell proliferation (GI50) while presenting low cytotoxicity in both cancer and normal cells.

The activity of indigo carmine against bacteriophages: an edible antiphage agent

Bacteriophage infections in bacterial cultures pose a significant challenge to industrial bioprocesses, necessitating the development of innovative antiphage solutions. This study explores the antiphage potential of indigo carmine (IC), a common FDA-approved food additive. IC demonstrated selective inactivation of DNA phages (P001, T4, T1, T7, λ) with the EC50 values ranging from 0.105 to 0.006 mg/mL while showing no activity against the RNA phage MS2. Fluorescence correlation spectroscopy (FCS) revealed that IC selectively binds to dsDNA, demonstrated by a significant reduction in the diffusion coefficient, whereas no binding was observed with ssDNA or RNA. Mechanistically, IC permeates the phage capsid, leading to genome ejection and capsid deformation, as confirmed by TEM imaging. Under optimal conditions (50 °C, 220 rpm), IC achieved up to a 7-log reduction in phage titer, with kinetic theory supporting the enhanced collision frequency induced by agitation. Additionally, IC protected E. coli cultures from phage-induced lysis without affecting bacterial growth or protein production, as demonstrated by GFP expression assays. IC’s effectiveness and environmental safety, combined with its FDA approval and cost-effectiveness, make it a promising antiphage agent for industrial applications.Key points• Indigo carmine effectively inactivates a broad spectrum of bacteriophages, offering protection to bacteria in industrial cultures.• A novel application of indigo carmine as a food-grade, environmentally safe, and FDA-approved antiphage agent protecting bacterial cultures.• Antiphage activity arises from indigo carmine’s interaction with DNA within the phage capsid without harming bacterial cells or compromising protein production in bacterial cultures.Graphical abstract

Genome-informed Discovery of Monchicamides A-K: Cyanobactins from the Microcoleaceae Cyanobacterium LEGE 16532.

Genome mining has emerged as an important tool for the discovery of natural products and is particularly effective for the swift identification of ribosomally synthesized and post-translationally modified peptides (RiPPs). Among RiPPs, cyanobactins have gained attention due to their diverse structures and bioactive properties. Here, we explored the Microcoleaceae cyanobacterium LEGE 16532 strain and identified the mon biosynthetic gene cluster (BGC), which was predicted to encode cyanobactin-like molecules. This led to the detection of 11 macrocyclic cyanobactins, the monchicamides, some of which feature mono- or diprenylation. One of the compounds was isolated, monchicamides I (9), and its planar structure was established by LC-HRESIMS/MS data as well as 1D and 2D NMR spectroscopy, confirming forward O-prenylation in Tyr. In addition, the absolute configuration of compound 9 was determined by Marfey's method and chiral-phase HPLC. The structures of the additional cyanobactins were proposed from MS/MS data analysis. The bioactivity profile of the isolated compound was also evaluated, but no cytotoxic, antimicrobial, or antiamoebic activity was observed.

Discovery of Undescribed Flavonoid and Rotenoid Derivatives from the Leaves of Derris trifoliata with Their Cytotoxic Activity.

Phytochemical study on the leaves of Derris trifoliata led to the isolation of five rotenoids (3-7) and ten flavonoid derivatives (1, 2, 8-15). Three of them, (2S)-5,7-dihydroxy-6-(2-hydroxyethyl)-2-(4-hydroxyphenyl)-8-(3-methylbut-2-en-1-yl)chroman-4-one (1), keampferol 3-O-[6-O-(E)-feruloyl-β-D-glucopyranosyl]-(1→2)-O-[α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside (2), and (+)-13-homo-13-oxa-6a,12a-dehydrorotenone (3) were verified by Scifinder as previously undescribed compounds. Their chemical structures were elucidated by using IR, ECD, HR-ESI-MS, 1D and 2D NMR methods. Compounds 1-15 were tested for their cytotoxic activities against SK-LU-1 and HepG2 cell lines. Compounds 4, 8, 14, and 15 showed moderate cytotoxic activities against SK-LU-1 cells with IC50 values ranging from 8.52 to 28.89 µM, and against HepG2 cells with IC50 values ranging from 7.11 to 34.85 µM.

Rapid-Scan Fourier Transform Infrared Difference Spectroscopy with Two-Dimensional Correlation Analysis to Show the Build-Up of Light-Adapted States in Bacterial Photosynthetic Reaction Centers.

Time-resolved, rapid-scan Fourier transform infrared (FT-IR) difference spectra have been recorded upon illumination on photosynthetic reaction centers (RCs) from Rhodobacter sphaeroides under fixed hydration conditions (relative humidity = 76%). Two different illumination schemes were adopted. Whereas the use of a laser flash (duration: 7 ns) made it possible to follow the kinetics of recombination of the light-induced state P+QA- to the neutral state PQA, the use of a 20.5 s continuous light from a lamp made it possible to follow both the build-up of a steady-state P+QA- population and its decay to PQA. Comparison between P+QA-/PQA FT-IR difference spectra obtained under (or 650 ms after) continuous illumination and obtained after one laser flash show small but meaningful differences, reflecting structural changes in the light-adapted state produced by the 20.5 s period of illumination. These differences are strikingly similar to those observed when comparing FT-IR difference spectra reflecting charge separation in photosystem II in light-adapted states and non-light-adapted states (c.f. Sipka et al., "Light-Adapted Charge-Separated State of Photosystem II: Structural and Functional Dynamics of the Closed Reaction Center". Plant Cell. 2021. 33(4): 1286-1302). Two-dimensional correlation spectroscopy analysis revealed that in all the observed series of time-resolved FT-IR difference spectra (under illumination, after illumination, and after a laser flash), marker bands at 1749, 1716, and 1668 cm-1 all evolve synchronously, demonstrating that electron transfer reactions and protein backbone response (at least the one reflected by the 1668 cm-1 band) are strongly correlated. Conversely, for spectra under and after continuous illumination, many asynchronicities are observed for (still unassigned) bands throughout the whole 1740-1200 cm-1 region, reflecting a more complicated molecular scenario in the RC upon build-up of the light-adapted state and during its relaxation to the resting neutral state.