Sugar Moiety Research Articles

Free and Glycosidically Bound Volatile Compounds in Okinawan Pineapple (Ananas comosus)

Fruit plants produce various volatile compounds that emit distinct aroma characteristics and contribute to their flavor qualities. However, some of these substances, especially hydroxyl-group molecules, are in non-volatile glycosylated forms. This study aimed to determine free and glycosidically bound volatile compounds in three Okinawan pineapple cultivars (‘N67-10′, ‘Yugafu’, and ‘Yonekura’). The free volatile components of squashed pineapple juice were analyzed using solid-phase microextraction (SPME)–arrow-gas chromatography–flame ionization detection/mass spectrometry (GC-FID/MS). The glycosides were collected through solid-phase extraction, hydrolyzed by β-glucosidase, and the released volatile compounds were measured. The sugar moieties of the glycosides were confirmed using GC-MS, and their glycoside constituents were analyzed using liquid chromatography (LC)-MS. Okinawan pineapple varied in its content and composition of free volatile components, which were predominantly comprised of esters, followed by alcohols, terpenes, and ketones. Eight hydroxyl-group compounds, including chavicol, eugenol, geraniol, phenylethyl alcohol, benzyl alcohol, 2-ethyl-1-hexanol, 1-hexanol, and 3-methyl-2-butenol, were released from their glycosylated forms via enzymatic hydrolysis, wherein the amounts of most of them were greater in ‘Yonekura’ than in the other cultivars. Moreover, two glycosides, chavicol-O-β-D-glucopyranoside and eugenol-O-β-D-glucopyranoside, were identified in all the cultivars, wherein the aglycones of both glycosides could be potential odor sources of the medicinal-herbal aromas. These results provide important information regarding both volatile-aroma qualities and bounded-aroma resources in Okinawan pineapple for fresh consumption and agroindustrial processing.

Sesquiterpenoids with Phytotoxic and Antifungal Activities from a Pathogenic Fungus Aspergillus alabamensis.

Five new carotane sesquiterpenoids (asperalacids A-E (1-5)), one new tricyclic sesquiterpenoid (4-hydroxy-5(6)-dihydroterrecyclic acid A (6)), and two known analogues (7-8) were obtained from a seagrass-derived fungus Aspergillus alabamensis, which was speculated to be a phytopathogenic fungus, isolated from the necrotic leaves of Enhalus acoroides. The structures of 1-6 were established by a combination of spectroscopic methods, including comprehensive NMR analysis, mass spectrometry, conformational analysis, NMR computational methods, and ECD calculations. Compound 4, with higher inhibitory activity on wheat (Triticum aestivum L.) root and shoot elongation than the positive control terbutryn, a broad-spectrum systemic herbicide, is a new natural plant growth inhibitor. Compound 5, belonging to the rare glycosylated sesquiterpenoid class, represents the first example of glycosylated carotane sesquiterpenoid whose sugar moiety was identified as α-d-glucose. Compounds 1-4 and 6 displayed weak to potent antimicrobial activity against the plant pathogenic fungi Fusarium oxysporum, Fusarium graminearum, and Penicillium italicum and the Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus.

Rapid Assessment of Lipidomics Sample Purity and Quantity Using Fourier-Transform Infrared Spectroscopy.

Despite the increasing popularity of liquid chromatography–mass spectrometry (LC-MS)-based lipidomics, there is a lack of accepted and validated methods for lipid extract quality and quantity assessment prior to LC-MS. Fourier-Transform Infrared Spectroscopy (FTIR) has been reported for quantification of pure lipids. However, the impact of complex lipid sample complexity and purity on total lipid quantification accuracy has not been investigated. Here, we report comprehensive assessment of the sample matrix on the accuracy of lipid quantification using Attenuated Total Reflectance (ATR)-FTIR and establish a simple workflow for lipidomics sample quantification. We show that both pure and complex lipids show characteristic FTIR vibrations of CH- and C=O-stretching vibrations, with a quantitative range of 40–3000 ng and a limit of detection of 12 ng, but sample extraction method and local baseline subtraction during FTIR spectral processing significantly impact lipid quantification via CH stretching. To facilitate sample quality screening, we developed the Lipid Quality (LiQ) score from a spectral library of common contaminants, using a ratio of peak heights between CH stretching vibrations maxima and the collective vibrations from amide/amine, CH-stretching minima and sugar moieties. Taking all tested parameters together, we propose a rapid FTIR workflow for routine lipidomics sample quality and quantity assessment and tested this workflow by comparing to the total LC-MS intensity of targeted lipidomics of 107 human plasma lipid extracts. Exclusion of poor-quality samples based on LiQ score improved the correlation between FTIR and LC-MS quantification. The uncertainty of absolute quantification by FTIR was estimated using a 795 ng SPLASH LipidoMix standard to be <10%. With low sample requirement, we anticipate this simple and rapid method will enhance lipidomics workflow by enabling accurate total lipid quantification and normalization of lipid quantity for MS analysis.

Intermolecular Interactions of Nucleoside Antibiotic Tunicamycin with On-Target MraYCB-TUN and Off-Target DPAGT1-TUN in the Active Sites Delineated by Quantum Mechanics/Molecular Mechanics Calculations.

Tunicamycin (TUN) is a nucleoside antibiotic with a complex structure comprising uracil, tunicamine sugar, N-acetylglucosamine (GlcNAc), and fatty acyl tail moieties. TUN, known as a canonical inhibitor, blocks vital functions of certain transmembrane protein families, for example, the insect enzyme dolichyl phosphate α-N-acetylglucosaminylphosphotransferase (DPAGT1) of Spodoptera frugiperda and the bacterial enzyme phospho-N-acetylmuramoylpentapeptide translocase (MraYCB) of Clostridium bolteae. Accurate description of protein–drug interactions has an immense impact on structure-based drug design, while the main challenge is to create proper topology and parameter entries for TUN in modeling protein–TUN interactions given the structural complexity. Starting from DPAGT1–TUN and MraYCB–TUN crystal structures, we first sketched these structural complexes on the basis of the CHARMM36 force field and optimized each of them using quantum mechanics/molecular mechanics (QM/MM) calculations. By continuing calculations on the active site (QM region) of each optimized structure, we specified the characteristics of intermolecular interactions contributing to the binding of TUN to each active site by quantum theory of atoms in molecules (QTAIM) and natural bond orbital (NBO) analyses at the M06-2X/6-31G** level. The results outlined that TUN insertion into each active site requires multiple weak, moderate, and strong hydrogen bonds accompanying charge–dipole, dipole–dipole, and hydrophobic interactions among different TUN moieties and adjacent residues. The water-mediated interactions also play central roles in situating the uracil and tunicamine moieties of TUN within the DPAGT1 active site as well as in preserving the uracil-binding pocket in the MraYCB active site. The TUN binds more strongly to DPAGT1 than to MraYCB. The information garnered here is valuable particularly for better understanding mode of action at the molecular level, as it is conducive to developing next generations of nucleoside antibiotics.

Influence of Sugar Modifications on the Nucleoside Conformation and Oligonucleotide Stability: A Critical Review.

Ribofuranose sugar conformation plays an important role in the structure and dynamics of functional nucleic acids such as siRNAs, AONs, aptamers, miRNAs, etc. To improve their therapeutic potential, several chemical modifications have been introduced into the sugar moiety over the years. The stability of the oligonucleotide duplexes as well as the formation of stable and functional protein-oligonucleotide complexes are dictated by the conformation and dynamics of the sugar moiety. In this review, we systematically categorise various ribofuranose sugar modifications employed in DNAs and RNAs so far. We discuss different stereoelectronic effects imparted by different substituents on the sugar ring and how these effects control sugar puckering. Using this data, it would be possible to predict the precise use of chemical modifications and design novel sugar-modified nucleosides for therapeutic oligonucleotides that can improve their physicochemical properties.

Response to Oswald etal.: ST6Gal1 in plasma is dispensable for IgG sialylation.

The presence or absence of terminal sialic acid residues in the sugar moiety attached to the Fc-domain of IgG molecules modulates IgG activity and is associated with autoimmune or infection related inflammation. In a recent paper, Oswald and colleagues suggest that IgG sialylation may occur post IgG secretion from plasma cells, which would be a major issue for therapeutic antibodies injected into patients. In contrast, we argue that previous work rather demonstrates that IgG sialylation occurs within B cells and that the experimental system used by the authors is not suitable to address this critical question.

Subcellular localization of the enterobacterial common antigen GT-E-like glycosyltransferase, WecG.

Enterobacterales have developed a specialized outer membrane polysaccharide (enterobacterial common antigen [ECA]). ECA biosynthesis begins on the cytoplasmic side of the inner membrane (IM) where glycosyltransferases sequentially add sugar moieties to form a complete repeat unit which is then translocated across the IM by WzxE before being polymerized into short linear chains by WzyE/WzzE. Research into WecG, the enzyme responsible for generating ECA lipid-II, has not progressed beyond Barr et al. (1988) who described WecG as a membrane protein. Here we revise our understanding of WecG and re-characterize it as a peripherally associated membrane protein. Through the use of Western immunoblotting we show that WecG in Shigella flexneri is maintained to the IM via its three C-terminal helices and further identify key residues in helix II which are critical for this interaction which has allowed us to identify WecG as a GT-E glycosyltransferase. We investigate the possibility of protein complexes and ultimately show that ECA lipid-I maintains WecG to the membrane which is crucial for its function. This research is the first since Barr et al. (1988) to investigate the biochemistry of WecG and reveals possible novel drug targets to inhibit WecG and thus ECA function and cell viability.

Engineering of Leuconostoc citreum for Efficient Bioconversion of Soy Isoflavone Glycosides to Their Aglycone Forms.

Soy isoflavones are phytochemicals that possess various beneficial physiological properties such as anti-aging, anti-tumor, and antioxidant properties. Since soy isoflavones exist in glycoside forms, their bioavailability requires initial hydrolysis of the sugar moieties bound to them to be efficiently absorbed through the gut epithelium. Instead of conventional chemical hydrolysis using acids or organic solvents, alternative strategies for enhancing the bioavailability of soy isoflavones using biological methods are gaining attention. Here, we engineered Leuconostoc citreum isolated from Korean kimchi for efficient bioconversion of soy isoflavone glycosides into their aglycone forms to enhance their bioavailability. We first constructed an expression module based on the isoflavone hydrolase (IH)-encoding gene of Bifidobacterium lactis, which mediates conversion of isoflavone glycosides to aglycone forms. Using a high copy number plasmid and bicistronic expression design, the IH was successfully synthesized in L. citreum. Additionally, we determined enzymatic activity of the IH using an in vivo β-glucosidase assay and confirmed its highly efficient bioconversion efficiency for various types of isoflavone glycosides. Finally, we successfully demonstrated that the engineered L. citreum could convert isoflavone glycosides present in fermented soymilk into aglycones.

Changes in physicochemical and anticancer properties modulated by chemically modified sugar moieties within sequence-related G-quadruplex structures.

We systematically investigated the influence of locked nucleic acid (LNA), unlock nucleic acid (UNA), and 2’-O-methyl-RNA (2’-O-Me-RNA) residues on the thermal stability, structure folding topology, biological activity and enzymatic resistance of three sequence-related DNA G-quadruplexes. In order to better understand the mechanism of action of the studied modifications, a single-position substitution in the loops or G-tetrads was performed and their influence was analyzed for a total of twenty-seven modified G-quadruplex variants. The studies show that the influence of each modification on the physicochemical properties of G-quadruplexes is position-dependent, due to mutual interactions between G-tetrads, loops, and additional guanosine at 5’ or 3’ end. Nevertheless, the anticancer activity of the modified G-quadruplexes is determined by their structure, thus also by the local changes of chemical character of sugar moieties, what might influence the specific interactions with therapeutic targets. In general, UNA modifications are efficient modulators of the G-quadruplex thermodynamic stability, however they are poor tools to improve the anticancer properties. In contrast, LNA and 2’-O-Me-RNA modified G-quadruplexes demonstrated certain antiproliferative potential and might be used as molecular tools for designing novel G-quadruplex-based therapeutics.

Contribution of TagA-Like Glycosyltransferases to the Assembly of the Secondary Cell Wall Polysaccharide in Bacillus anthracis.

Bacillus anthracis elaborates a secondary cell wall polysaccharide (SCWP) made of 6 to 12 trisaccharide units. Pyruvyl and acetyl substitutions of the distal unit are prerequisites for the noncovalent retention of 22 secreted Bacillus S-layer (Bsl)-associated proteins bearing an S-layer homology (SLH) domain. Surface display of Bsl proteins contributes to cell separation as well as virulence. Earlier work suggested that TagO initiates the synthesis of SCWP while GneY and GneZ, two UDP-GlcNAc 2-epimerases, synthesize ManNAc that is later incorporated in the repeat unit (→4)-ManNAc-(β1→4)-GlcNAc-(β1→6)-GlcNAc-(α1→). In organisms that synthesize wall teichoic acid, TagA catalysts have been shown to form the glycosidic bond ManNAc-(β1→4)-GlcNAc. Here, we show that genes bas2675 and bas5272, predicted to encode glycosyltransferases of the WecB/TagA/CpsF family (PFAM03808; CAZy GT26), are required for B. anthracis SCWP synthesis and S-layer assembly. Similar to tagO or gneY gneZ mutants, B. anthracis strains depleted of tagA1 (bas5272) cannot maintain cell shape, support vegetative growth, or synthesize SCWP. Expression of tagA2 (bas2675), or Staphylococcus aureus tagA on a plasmid, rescues the nonviable tagA1 mutant. We propose that TagA1 and TagA2 fulfill overlapping and key glycosyltransferase functions for the synthesis of repeat units of the SCWP of B. anthracis. IMPORTANCE Glycosyltransferases (GTs) catalyze the transfer of sugar moieties from activated donor molecules to acceptor molecules to form glycosidic bonds using a retaining or inverting mechanism. Based on the structural relatedness of their catalytic and carbohydrate-binding modules, GTs have been grouped into 115 families in the Carbohydrate-Active EnZyme (CAZy) database. For complex products, the functional assignment of GTs remains highly challenging without the knowledge of the chemical structure of the assembled polymer. Here, we propose that two uncharacterized GTs of B. anthracis belonging to the WecB/TagA/CpsF family incorporate ManNAc in repeat units of the secondary cell wall polymer of bacilli species.

Preparation of RNAs with non-canonical 5' ends using novel di- and trinucleotide reagents for co-transcriptional capping.

Many eukaryotic and some bacterial RNAs are modified at the 5′ end by the addition of cap structures. In addition to the classic 7-methylguanosine 5′ cap in eukaryotic mRNA, several non-canonical caps have recently been identified, including NAD-linked, FAD-linked, and UDP-glucose-linked RNAs. However, studies of the biochemical properties of these caps are impaired by the limited access to in vitro transcribed RNA probes of high quality, as the typical capping efficiencies with NAD or FAD dinucleotides achieved in the presence of T7 polymerase rarely exceed 50%, and pyrimidine derivatives are not incorporated because of promoter sequence limitations. To address this issue, we developed a series of di- and trinucleotide capping reagents and in vitro transcription conditions to provide straightforward access to unconventionally capped RNAs with improved 5′-end homogeneity. We show that because of the transcription start site flexibility of T7 polymerase, R1ppApG-type structures (where R1 is either nicotinamide riboside or riboflavin) are efficiently incorporated into RNA during transcription from dsDNA templates containing both φ 6.5 and φ 2.5 promoters and enable high capping efficiencies (∼90%). Moreover, uridine-initiated RNAs are accessible by transcription from templates containing the φ 6.5 promoter performed in the presence of R2ppUpG-type initiating nucleotides (where R2 is a sugar or phosphate moiety). We successfully employed this strategy to obtain several nucleotide-sugar-capped and uncapped RNAs. The capping reagents developed herein provide easy access to chemical probes to elucidate the biological roles of non-canonical RNA 5′ capping.

Benzothioate Glycoside from a terrestrial Streptomyces sp. TBRC 11511 from Thailand.

A new benzothioate glycoside metabolite, phitsanoside A, together with its new and known derivatives were isolated from Streptomyces sp. TBRC 11511 collected from sediment of a dry evergreen forest located in Phitsanulok Province, Thailand. The structure elucidation of the new compound was interpreted on the basis of spectroscopic data analysis. The configuration of the sugar moiety was derived based on NOESY nuclear magnetic resonance correlations, a vicinal coupling constant analysis, and the measurement of an optical rotation from the hydrolyzed sugar unit, which was identified as β-D-glucopyranose. Phitsanoside A did not exhibit antibacterial activity against Bacillus cereus, Mycobacterium tuberculosis or Staphylococcus aureus, but phitsanoside B showed activity against all of them with MIC values of 3.13, 25 and 12.5 μg ml-1, respectively.

Antibody Recognition of Different Staphylococcus aureus Wall Teichoic Acid Glycoforms.

Wall teichoic acids (WTAs) are glycopolymers decorating the surface of Gram-positive bacteria and potential targets for antibody-mediated treatments against Staphylococcus aureus, including methicillin-resistant (MRSA) strains. Through a combination of glycan microarray, synthetic chemistry, crystallography, NMR, and computational studies, we unraveled the molecular and structural details of fully defined synthetic WTA fragments recognized by previously described monoclonal antibodies (mAbs 4461 and 4497). Our results unveiled the structural requirements for the discriminatory recognition of α- and β-GlcNAc-modified WTA glycoforms by the complementarity-determining regions (CDRs) of the heavy and light chains of the mAbs. Both mAbs interacted not only with the sugar moiety but also with the phosphate groups as well as residues in the ribitol phosphate (RboP) units of the WTA backbone, highlighting their significant role in ligand specificity. Using elongated WTA fragments, containing two sugar modifications, we also demonstrated that the internal carbohydrate moiety of α-GlcNAc-modified WTA is preferentially accommodated in the binding pocket of mAb 4461 with respect to the terminal moiety. Our results also explained the recently documented cross-reactivity of mAb 4497 for β-1,3/β-1,4-GlcNAc-modified WTA, revealing that the flexibility of the RboP backbone is crucial to allow positioning of both glycans in the antibody binding pocket.

Unusual ring B-seco isosteroidal alkaloid, yibeiglycoalkaloids A−E from Fritillaria pallidiflora schrenk

Five previously undescribed steroidal glycoalkaloids(SGAs)and a rare ring B-seco isosteroidal alkaloid, were isolated from Fritillaria pallidiflora Schrenk, along with six known alkaloids. The structures of these alkaloids were established by comprehensive analyses of the 1D, 2D-NMR and HR-ESI-MS data. Configurations of sugar moieties were resolved by chemical derivations. The isolated compounds showed nitric oxide (NO) inhibitory activities in lipopolysaccharide (LPS) induced RAW264.7 cells, and yibeinone exhibited the strongest inhibitory effects among them. This study revealed that the alkaloids from F. pallidiflora might have significant anti-inflammatory potentials.

Biocatalyzed Synthesis of Glycostructures with Anti-infective Activity.

ConspectusMolecules containing carbohydrate moieties play essential roles in fighting a variety of bacterial and viral infections. Consequently, the design of new carbohydrate-containing drugs or vaccines has attracted great attention in recent years as means to target several infectious diseases.Conventional methods to produce these compounds face numerous challenges because their current production technology is based on chemical synthesis, which often requires several steps and uses environmentally unfriendly reactants, contaminant solvents, and inefficient protocols. The search for sustainable processes such as the use of biocatalysts and eco-friendly solvents is of vital importance. Therefore, their use in a variety of reactions leading to the production of pharmaceuticals has increased exponentially in the last years, fueled by recent advances in protein engineering, enzyme directed evolution, combinatorial biosynthesis, immobilization techniques, and flow biocatalysis. In glycochemistry and glycobiology, enzymes belonging to the families of glycosidases, glycosyltransferases (Gtfs), lipases, and, in the case of nucleoside and nucleotide analogues, also nucleoside phosphorylases (NPs) are the preferred choices as catalysts.In this Account, on the basis of our expertise, we will discuss the recent biocatalytic and sustainable approaches that have been employed to synthesize carbohydrate-based drugs, ranging from antiviral nucleosides and nucleotides to antibiotics with antibacterial activity and glycoconjugates such as neoglycoproteins (glycovaccines, GCVs) and glycodendrimers that are considered as very promising tools against viral and bacterial infections.In the first section, we will report the use of NPs and N-deoxyribosyltransferases for the development of transglycosylation processes aimed at the synthesis of nucleoside analogues with antiviral activity. The use of deoxyribonucleoside kinases and hydrolases for the modification of the sugar moiety of nucleosides has been widely investigated.Next, we will describe the results obtained using enzymes for the chemoenzymatic synthesis of glycoconjugates such as GCVs and glycodendrimers with antibacterial and antiviral activity. In this context, the search for efficient enzymatic syntheses represents an excellent strategy to produce structure-defined antigenic or immunogenic oligosaccharide analogues with high purity. Lipases, glycosidases, and Gtfs have been used for their preparation.Interestingly, many authors have proposed the use Gtfs originating from the biosynthesis of natural glycosylated antibiotics such as glycopeptides, macrolides, and aminoglycosides. These have been used in the chemoenzymatic semisynthesis of novel antibiotic derivatives by modification of the sugar moiety linked to their complex scaffold. These contributions will be described in the last section of this review because of their relevance in the fight against the spreading phenomenon of antibiotic resistance. In this context, the pioneering in vivo synthesis of novel derivatives obtained by genetic manipulation of producer strains (combinatorial biosynthesis) will be shortly described as well.All of these strategies provide a useful and environmentally friendly synthetic toolbox. Likewise, the field represents an illustrative example of how biocatalysis can contribute to the sustainable development of complex glycan-based therapies and how problems derived from the integration of natural tools in synthetic pathways can be efficiently tackled to afford high yields and selectivity. The use of enzymatic synthesis is becoming a reality in the pharmaceutical industry and in drug discovery to rapidly afford collections of new antibacterial or antiviral molecules with improved specificity and better metabolic stability.

Design, synthesis, molecular docking and antimicrobial activities of novel triazole‐ferulic acid ester hybrid carbohydrates

In spite of search for newer and effective antibacterial drugs, nature has paved way to identify lead molecules from plants to treat chronic diseases. In an effort to develop biologically active antibacterial and antifungal agents, the ester of a natural compound ferulic acid (FA) bearing 1,2,3-triazole core and different sugar moieties have been designed and synthesized by Cu(I)-catalysed click dipolar cycloaddition. The chemical structures of the new derivatives were characterized using different spectroscopic techniques. The bactericidal and fungicidal activities were evaluated for synthesized clicked 1,2,3-triazoles compounds. Most of the compounds exhibited moderate to good activity against some strains of bacteria and fungi. Derivative 12 have shown superior antibacterial activity against G-positive Bacillus cereus. Appreciable antifungal activity was observed for all compounds in comparison with cyclohexamide the reference standard drug used. The molecular docking assessments indicated that most compounds exhibited good binding affinity with the target proteins of the microorganisms. Moreover, reports of physicochemical properties, pharmacokinetics, and drug-likeness for all derivatives showed that compound 12 is a potential candidate to develop novel antimicrobial drug.

Pro-Apoptotic and Pro-Autophagic Properties of Cardenolides from Aerial Parts of Pergularia tomentosa

Pergularia tomentosa L., a milkweed tropical plant belonging to the family Asclepiadaceae, is a rich source of unusual cardiac glycosides, characterised by transfused A/B rings and a sugar moiety linked by a double link, generating a dioxanoid structure. In the present report, five cardenolides isolated from the aerial parts of the plant (calactin, calotropin, 12β-hydroxycalactin, 12β,6′-dihydroxycalotropin, and 16α-hydroxycalotropin) were investigated for their biological effects on a human hepatocarcinoma cell line. Cell viability was monitored by an MTT assay. The occurrence of apoptosis was evaluated by detecting caspase-3 activation and chromatin fragmentation. The ability of these compounds to induce autophagy was analysed by monitoring two markers of the autophagic process, LC3 and p62. Our results indicated that all cardenolides had cytotoxic effects, with IC50 ranging from 0.127 to 6.285 μM. All compounds were able to induce apoptosis and autophagy, calactin being the most active one. Some of them also caused a reduction in cell migration and a partial block of the cell cycle into the S-phase. The present study suggests that selected cardenolides from aerial parts of P. tomentosa, particularly calactin, possess potentially desirable properties for further investigation as anticancer agents.

Design, Synthesis, Characterization, DFT Calculations, Molecular Docking Study, and Antimicrobial Activity of Hydrazones Bearing Pyrimidine and Sugar Moieties

Design, Synthesis, Characterization, DFT Calculations, Molecular Docking Study, and Antimicrobial Activity of Hydrazones Bearing Pyrimidine and Sugar Moieties

Integrated strategy for identifying isoflavones in Belamcandae Rhizoma based on the combination of mass defect filtering and neutral-loss-triggered multistage fragmentation

Belamcandae Rhizoma is a widely used traditional Chinese herbal medicine with isoflavones as the main active ingredient. In this paper, an integrated strategy was developed to discover and identify new isoflavones in Belamcandae Rhizoma by an ultra-high-performance liquid chromatography coupled with high resolution multistage mass spectrometry. Different characterization methods were used based on structural features of isoflavone aglycones and glycosides. On one hand, we adopted a data-dependent acquisition mode incorporated into intelligent AcquireX deep scan algorithms to analyze crude extract, and used a mass defect filtering technique to filter out two kinds of isoflavone aglycones from the extract. On the other hand, neutral-loss-triggered MSn was used to analyze isoflavone glycosides, and under this acquisition mode, MSn scan only took place when chemical components exhibited specific neutral losses. Identification of isoflavones was achieved either by comparison with reference compounds or analysis of characteristic product ions based on MS2 or MSn fragmentation patterns. UV absorbance spectra also contributed to the confirmation of isoflavones. As a result, a total of 65 isoflavone aglycones (42 new aglycones) and 142 isoflavone glycosides (122 new glycosides) were discovered, including a number of trace components. Meanwhile, modifications of new sugar moieties, such as xylose, rhamnose and 6-O-(4‑hydroxy-3,5-dimethoxybenzoyl)-β-D-glucose, were discovered in Belamcandae Rhizoma for the first time. These results indicated the feasibility of this established strategy for in-depth identification of new isoflavone aglycones and glycosides.

Lipase-Mediated Mechanoenzymatic Synthesis of Sugar Esters in Dissolved Unconventional and Neat Reaction Systems.

Mechanochemical and biocatalytic approaches in modern research are two major assets to develop greener processes. In the present study, these modular tools of sustainability are pointed toward the production of versatile and daily employed compounds such as surfactants. Toward this aim, glycolipids, a class of nonionic surfactants composed of ubiquitous and primary metabolites such as sugar and fatty acid moieties, represent a promising alternative to petroleum-derived surface-active agents. Therefore, the combination of biocatalysis with mechanochemistry aiming at glycolipid synthesis seemed a logical step that was taken in this study for the first time. The monoacylated model compound glucose-6-O-decanoate was synthesized with the help of a bead mill apparatus using two different unconventional dissolved reaction systems, namely, menthol-based hydrophobic deep eutectic solvents and 2-methyl-2-butanol, thus reaching up to 12% yield in the latter based on the conversion of vinyl decanoate, after only 90 min of reaction. In addition, a neat reaction system using an excess of vinylated fatty ester as an adjuvant allowed a 27 mM/h space-time yield. The overall significant increase in productivities, up to 6 times, compared to standard heating and shaking methods, shows the tremendous potential of mechanoenzymatic synthesis.