Abstract This study proposes a chemical analysis-based framework for wood species discrimination by integrating Fourier-transform infrared spectroscopy (FT-IR), gas chromatography–mass spectrometry (GC–MS), and chemometric analysis. After data standardization and principal component analysis (PCA) dimensionality reduction, the leading principal components were used to construct classification models. Several multivariate approaches, including k-nearest neighbors (KNN), linear discriminant analysis (LDA), and artificial neural networks (ANN), were evaluated to examine classification behavior under limited sample conditions. While ANN and KNN showed reasonable classification performance, ANN results varied across repeated trials, reflecting sensitivity to small datasets. In contrast, LDA provided more stable and interpretable discrimination, particularly when FT-IR and GC–MS data were combined. The combined LDA model consistently outperformed single-technique models in cross-validation, demonstrating the complementary value of structural information captured by FT-IR and extractive-related chemical features revealed by GC–MS. The influence of FT-IR spectral range selection was also examined, and restricting analysis to chemically relevant regions improved model stability as the number of species increased. Overall, the results demonstrate that integrating multiple chemical descriptors enables effective species-level discrimination of hardwoods under practical sampling constraints and supports future expansion to broader taxonomic datasets.

To elucidate site-specific lignin in Ginkgo biloba leaves, the chemical architecture of lignin in leaf veins (LV) and petioles (LP) was analyzed by combining thioacidolysis and 2D Heteronuclear Single Quantum Correlation (HSQC) NMR. LV and LP were separated from the leaves, and polysaccharides were enzymatically digested to give residual lignin (EL). Acetylated EL (ELAc) and milled-wood lignin (MWL) were also prepared. As a result of thioacidolysis, guaiacyl (G) units were the predominant monomers in all samples. In LV and LP samples, the p-hydroxyphenyl (H) unit (approx. 1–2

We investigated how lignin precursors are stored and used during secondary-wall formation in Pinus thunbergii stems, addressing the current lack of quantitative, spatially resolved information, especially on monolignols and their glucosides. Two complementary cryogenic workflows, microdissection (CryoMD) for µm-scale fractionation and secondary ion mass spectrometry with gas cluster ion beam (Cryo-GCIB-SIMS) for sub-µm imaging, were developed to quantify and visualize monolignols and their glucosides across defined cell-wall formation stages in compression and opposite woods of P. thunbergii. Coniferin (CF) and p-glucocoumaryl alcohol were the dominant storage forms, while free coniferyl alcohol was in trace amounts, and p-coumaryl alcohol was detected mainly in the mature xylem region; CF concentration peaked near the cambial zone and declined with cell wall thickening. GCIB-SIMS visualized CF in domains adjacent to the cell wall at the cell expansion-cell wall thickening boundary and revealed a continuous distribution over 10-20 μm along the cell axis direction. These findings indicate that monolignol glucosides reside in spatially restricted, possibly mobile storage domains that supply precursors precisely where and when lignification proceeds, refining current models of monolignol transport. The integrated cryo approach provides a framework for linking intracellular metabolism to cell and tissue-level cell wall architecture.

The chirality of amino acids in extraterrestrial materials may provide an insight into the origin of the essential l-enantiopure amino acids in the terrestrial biosphere. In 2020, the Hayabusa2 mission succeeded in bringing back surface materials from the C-type asteroid (162173) Ryugu to the Earth. Amino acids were one of the targeted organic molecules to be studied in the Ryugu samples. To analyze the various structural isomers of amino acids, which were expected to be present, from the limited amount of the returned samples, the development of a highly-sensitive and selective analytical method was necessary. In the present study, a three-dimensional high-performance liquid chromatography (3D-HPLC) system has been developed for the enantioselective determination of five proteinogenic and three non-proteinogenic amino acids in the Ryugu samples, in which amino acids in the sample were separated by reversed-phase, anion-exchange and enantioselective columns after the fluorescence derivatization with 4-fluoro-7-nitro-2,1,3-benzoxadiozole. The applicability of the analytical system to the extraterrestrial samples was evaluated by analyzing several types of carbonaceous meteorites before applying the system to the Ryugu samples. In the analysis of the Ryugu samples, all of the target amino acids were successfully determined quantitatively. Non-proteinogenic amino acids including 2-amino-n-butyric acid, isovaline and norvaline, rarely present in the terrestrial environment, were found as almost racemic mixtures with 47.1 to 55.2%l.

Lignin is an aromatic polymer that constitutes plant cell walls. The polymerization of lignin proceeds by radical coupling, and this process requires radicalization of the phenolic end of lignin by enzymes. However, due to the steric hindrance between enzymes, lignin, and polysaccharides, the direct oxidation of the phenolic end of lignin by the enzyme would be difficult, and the details of the growth of lignin are still unknown. In this study, enzymatic dehydrogenative polymerization experiments were conducted using coniferyl alcohol (CA) and the deuterium-labeled lignin model compound (D-LM) under a noncontact condition in which horseradish peroxidase cannot directly oxidize D-LM due to separation by a dialysis membrane. Analysis of deuterium-labeled degraded compounds obtained by a combination of methylation and thioacidolysis revealed the formation of the bond between the phenolic end of D-LM and CA, suggesting that membrane-permeable, low-molecular-weight lignols functioned as a redox shuttle mediator.

A lignin model oligomer with only beta-O-4 linkages and a selectively deuterium-labelled methoxy group at the phenolic terminal units was synthesised to clarify the behaviour of the phenolic end of oligolignols. First, t-butoxycarbonylmethyl vanillin was synthesised and oligomerised by nucleophilic addition, a known method. The terminal of the oligomers was then subjected to nucleophilic addition to [3-OCD3]benzyl vanillin to achieve selective labelling of the terminal units. A deuterium-labelled lignin model oligomer (D-LM) was obtained through debenzylation and subsequent reduction. The results of thioacidolysis after methylation revealed that the degree of polymerisation was about five, and the deuterium-labelled phenylpropane unit was located only at the phenolic terminal moiety.

IntroductionTaxus species contain the anticancer alkaloid paclitaxel, as well as other taxanes similar in structure and potentially in effect to paclitaxel. Tissue-specific distribution patterns and seasonal variations of taxanes in some Taxus species have been reported; however, it is still under-presented for the taxanes in Taxus cuspidata.MethodsThe radial distributions of eight taxanes in the transverse surface of freeze-fixed T. cuspidata stems from the late summer and the spring seasons were investigated by cryo-time-of-flight secondary ion mass spectrometry and scanning electron microscopy (cryo-TOF-SIMS/SEM) visualization and liquid chromatography-mass spectrometry (LC-MS) quantitative analysis. By optical microscopic observation, seasonal differences in the amounts and distribution patterns of target taxanes were further characterized in specific tissues.Results and DiscussionThe overall amount of taxanes was higher in the late summer than in the spring. Also, taxanes’ radial distribution was generally found at higher concentration in the phloem, the cambium and lower level in the periderm, the latest-forming xylem, with different taxanes showing several patterns with distinction between seasons, which were considered related to seasonal plant physiological behaviors. In addition, the distribution of baccatin III (BAC) was investigated at the cellular level, which was regarded in specific cells suggesting its transport in the radial and axial directions in the T. cuspidata stem. Characterizing the microscopic distribution of taxanes in the T. cuspidata stem is expected to play a role in the further study of their biosynthesis and in planta behaviors.

The distributions of monolignol glucosides (MLGs) in compression and opposite woods of Pinus thunbergii were assessed using cryo-time-of-flight secondary ion mass spectrometry to investigate their involvement in lignification. p-Glucocoumaryl alcohol (PG) was identified in the region of the differentiating xylem adjacent to the cambial zone only in compression wood, whereas coniferin (CF) was similarly localized in both compression and opposite woods. Their distribution from the phloem to the xylem was evaluated by high-performance liquid chromatography (HPLC) using serial tangential sections. Variations in storage amounts of CF and PG in the stem of P. thunbergii agreed with lignification stages of the tracheid, supporting the idea that MLGs act as a storage and transportation form of lignin precursors. The imaging of monolignol (ML)-dependent active lignification sites using fluorescence-tagged MLs supported distinct distribution patterns of MLGs for lignification in compression and opposite woods. Methylation-thioacidolysis was applied to compression and opposite wood samples to examine the structural difference between the guaiacyl (G) and p-hydroxyphenyl (H) units in lignin. Most of the H units in compression wood were detected as lignin end groups via thioacidolysis. PG was detected in opposite wood by HPLC; however, the H unit was not detected by thioacidolysis. The differences in ML and MLG distributions, enzyme activity, and resultant lignin structures between the G and H units suggest the possibility of individual mechanisms regulating the heterogeneous structures of G and H unit in lignin.

The authors' series of studies aimed to explore the potential of terahertz time-domain spectroscopy (THz-TDS) in cellulose crystallographic studies, since THz radiation can detect most intermolecular vibrations and respond to lattice phonons. In this study, we tracked changes in four types of cellulose after ball milling. As the planetary ball milling time increases, it is observed through electron microscopy that the four types of cellulose particles are gradually destroyed into finer particles, while gel permeation chromatography can prove that the molecular weight gradually decreases after ball milling and the dispersity gradually approaches one, which indicates that the dispersion of the material was reduced. The most fascinating observation was made by THz-TDS, that is we have confirmed that after ball milling, the absorption characteristics of cellulose I and II in cellulose I treated with 10% NaOH (crystalline partial transition from cellulose I to II) exhibited an opposite trend. Specifically, the absorption of cellulose II at 2.40THz and 2.77THz increased, while the absorption of cellulose I at 2.11THz and 3.04THz decreased after ball milling, which suggests an increased conversion rate of cellulose I to cellulose II post-milling. Cellulose with different crystalline allomorphs shows different characteristic absorption in the THz region, and the peak position will not change even after the ball milling, only the absorption intensity changes. Although it can be observed through the most traditional X-ray diffraction method that the crystallinity index of all cellulose samples gradually decreases after ball milling. However, different from the THz results, the change after ball milling of cellulose I treated with 10% NaOH is only reflected in very subtle pattern changes, that is, the peak close to the 200 crystalline plane position is slightly shifted after ball milling.

Lignin dimeric units are characterized by various inter-unit linkage types such as beta-O-4, beta-5, beta-beta, and beta-1. Spirodienones are the native form of the beta-1 structures, but the content in lignin has not been clarified. In this study, the ring-1-C-13 labeled coniferin was synthesized and administered to Ginkgo biloba shoots, obtaining ring-1 selectively labeled xylem samples. Enzymatically saccharified lignin (EL) samples were prepared from the xylem sample (400-600 mu m distant region from the cambial zone), and solution-state quantitative C-13 NMR and solid-state CP/MAS NMR measurements were conducted. Acetylated EL (ELAc) was also prepared from the xylem sample (600-800 mu m distant region from the cambial zone), and solution-state quantitative C-13 NMR and H-1-C-13 2D NMR measurements were conducted. Difference spectra obtained by subtracting the unlabeled spectra from the ring-1 labeled spectra showed that the ring-1 was responsible for broad signals at 134 ppm and signals of the spirodienone structure at 56.11 ppm (in solid-state), 54.70 ppm (EL in solution-state), and 54.72 ppm (ELAc in solution-state). The ratio of spirodienone structure was evaluated as 0.68 % (EL) and 0.72 % (ELAc) by the solution-state quantitative C-13 difference spectra, and 2.3 % (ELAc) by HSQC volume ratio of C alpha '-H to G2-H.

This study demonstrated the antibacterial and antiviral potential of condensed tannins and tannic acid when incorporated into fiber networks tested for functional material purposes. Condensed tannins were extracted from industrial bark of Norway spruce by using pressurized hot water extraction (PHWE), followed by purification of extracts by using XADHP7 treatment to obtain sugar-free extract. The chemical composition of the extracts was analyzed by using HPLC, GC‒MS and UHPLC after thiolytic degradation. The test matrices, i.e., lignocellulosic handsheets, were produced and impregnated with tannin-rich extracts, and tannic acid was used as a commercial reference. The antibacterial and antiviral efficacy of the handsheets were analyzed by using bioluminescent bacterial strains (Staphylococcus aureus RN4220+pAT19 and Escherichia coli K12+pCGLS11) and Enterovirus coxsackievirus B3. Potential bonding of the tannin-rich extract and tannic acid within the fiber matrices was studied by using FTIR-ATR spectroscopy. The deposition characteristics (distribution and accumulation patterns) of tannin compounds and extracts within fiber networks were measured and visualized by direct chemical mapping using time-of-flight secondary ion mass spectrometry (ToF-SIMS) and digital microscopy. Our results demonstrated for the first time, how tannin-rich extracts obtained from spruce bark side streams with green chemistry possess antiviral and antibacterial properties when immobilized into fiber matrices to create substitutes for plastic hygienic products, personal protection materials such as surgical face masks, or food packaging materials to prolong the shelf life of foodstuffs and prevent the spread of infections. However, more research is needed to further develop this proof-of-concept to ensure stable chemical bonding in product prototypes with specific chemistry.

The Hayabusa2 spacecraft collected samples from the surface of the carbonaceous near-Earth asteroid (162173) Ryugu and brought them to Earth. The samples were expected to contain organic molecules, which record processes that occurred in the early Solar System. We analyzed organic molecules extracted from the Ryugu surface samples. We identified a variety of molecules containing the atoms CHNOS, formed by methylation, hydration, hydroxylation, and sulfurization reactions. Amino acids, aliphatic amines, carboxylic acids, polycyclic aromatic hydrocarbons, and nitrogen-heterocyclic compounds were detected, which had properties consistent with an abiotic origin. These compounds likely arose from an aqueous reaction on Ryugu's parent body and are similar to the organics in Ivuna-type meteorites. These molecules can survive on the surfaces of asteroids and be transported throughout the Solar System.

Abstract Ginkgo biloba L. saplings were cultivated in an airtight growth chamber with 13CO2 for two months. The 13C ratio of the newly developed xylem region was ca. 85%, evaluated by high lateral resolution secondary ion mass spectrometry and thioacidolysis/GC-MS. Quantitative solid-state 13C direct polarisation/magic angle spinning (DP/MAS) NMR measurements with high-speed MAS of 70.0 kHz were conducted for cutting-milled wood (CMW), ball-milled wood (BMW), and enzymatically saccharified lignin (EL) samples. In addition, quantitative liquid-state 13C NMR measurements were carried out for EL in DMSO-d 6. Major lignin aromatic signals were classified into three groups of aromatic carbons of C–H, C–C, and C–O, and their area ratio was compared within these measurements. EL samples in solid- and liquid-state showed nearly the same results. However, the results for CMW and BMW in solid-state NMR suggest the structural alteration of lignin within the sample preparation procedure, including ball milling.

The lignocellulosic biorefinery industry can be an important contributor to achieving global carbon net zero goals. However, low valorization of the waste lignin severely limits the sustainability of biorefineries. Using a hydrothermal reaction, we have converted sulfuric acid lignin (SAL) into a water-soluble hydrothermal SAL (HSAL). Here, we show the improvement of HSAL on plant nutrient bioavailability and growth through its metal chelating capacity. We characterize HSAL's high ratio of phenolic hydroxyl groups to methoxy groups and its capacity to chelate metal ions. Application of HSAL significantly promotes root length and plant growth of both monocot and dicot plant species due to improving nutrient bioavailability. The HSAL-mediated increase in iron bioavailability is comparable to the well-known metal chelator ethylenediaminetetraacetic acid. Therefore, HSAL promises to be a sustainable nutrient chelator to provide an attractive avenue for sustainable utilization of the waste lignin from the biorefinery industry.

The old Ginkgo biloba L. trees often develop cylindrical woody structures that grow downward from the underside of the large branches near the trunks. This structure is traditionally called “chi-chi” (a breast) in Japan. The structure of chi-chi has not been investigated in detail because of the rarity of such old trees of G. biloba . This study examined the chi-chi from wood anatomy and chemistry viewpoints. After debarking, there were many woody bulges and latent buds. There were hollows corresponding to these latent buds on the inner side of the bark. In the transverse section obtained from the tip part of the chi-chi sample, we found tracheids curving in a horizontal plane, and the parenchymatous latent bud tissue is the center of the swirl. Microscopic observations and X-ray micro-computed tomography suggest the growing course of the chi-chi due to several swirls contiguous to each other. From these observations, the downward growth of the chi-chi starting from the cambium cell division might be driven by the woody bulges with latent buds. The cell walls of the curved tracheids were not thickened, not rounded, and had no intercellular spaces, but their S 3 layers disappeared. Furthermore, the results of thioacidolysis and acetyl bromide analyses exhibited p -hydroxyphenyl subunits deposition to lignin and high lignin content on the tip part of the chi-chi. These results suggest that the curved tracheids in the chi-chi have compression-wood-like properties to some extent.

IntroductionPhellodendron amurense Rupr. contains rich alkaloids, which have been extensively applied in clinical treatments for their various biological activities. However, detailed microscopic distribution and roles of such alkaloids in P. amurense stem still need to be clarified. MethodsIn this study, the distribution of eight alkaloids in the transverse surface of freeze-fixed P. amurense stems in fall and summer has been visualized by cryo-time-of-flight secondary ion mass spectrometry and scanning electron microscopy (cryo-TOF-SIMS/SEM), which was found in living tissues with relative contents of different alkaloids varying with the position. In addition, the contents of these alkaloids quantified by high-performance liquid chromatography (HPLC) analysis suggested the seasonal variation from fall to the following summer. Results and discussionDistribution of eight alkaloids in the freeze-fixed stems of P. amurense from fall and summer seasons has been visualized and assigned into specific living tissues, with relative contents varying in different positions with seasons, which suggested their possible roles in the physiological processes of the plant itself or plant responding to changes in the surrounding conditions. ConclusionThis study provided a significant basis for further discussion of the genes or enzymes involved in these processes, which will contribute to investigating biosynthetic pathways and specific in planta roles of alkaloids.

We performed in-situ analysis on a ~ 1 mm-sized grain A0080 returned by the Hayabusa2 spacecraft from near-Earth asteroid (162173) Ryugu to investigate the relationship of soluble organic matter (SOM) to minerals. Desorption electrospray ionization-high resolution mass spectrometry (DESI-HRMS) imaging mapped more than 200 CHN, CHO, CHO–Na (sodium adducted), and CHNO soluble organic compounds. A heterogeneous spatial distribution was observed for different compound classes of SOM as well as among alkylated homologues on the sample surface. The A0080 sample showed mineralogy more like an Ivuna-type (CI) carbonaceous chondrite than other meteorites. It contained two different lithologies, which are either rich (lithology 1) or poor (lithology 2) in magnetite, pyrrhotite, and dolomite. CHN compounds were more concentrated in lithology 1 than in lithology 2; on the other hand, CHO, CHO–Na, and CHNO compounds were distributed in both lithologies. Such different spatial distribution of SOM is likely the result of interaction of the SOM with minerals, during precipitation of the SOM via fluid activity, or could be due to difference in transportation efficiencies of SOMs in aqueous fluid. Organic-related ions measured by time-of-flight secondary ion mass spectrometry (ToF–SIMS) did not coincide with the spatial distribution revealed by DESI-HRMS imaging. This result may be because the different ionization mechanism between DESI and SIMS, or indicate that the ToF–SIMS data would be mainly derived from methanol-insoluble organic matter in A0080. In the Orgueil meteorite, such relationship between altered minerals and SOM distributions was not observed by DESI-HRMS analysis and field-emission scanning electron microscopy, which would result from differences of SOM formation processes and sequent alteration process on the parent bodies or even on the Earth. Alkylated homologues of CHN compounds were identified in A0080 by DESI-HRMS imaging as observed in the Murchison meteorite, but not from the Orgueil meteorite. These compounds with a large C number were enriched in Murchison fragments with abundant carbonate grains. In contrast, such relationship was not observed in A0080, implying different formation or growth mechanisms for the alkylated CHN compounds by interaction with fluid and minerals on the Murchison parent body and asteroid Ryugu. Graphical Abstract