High-resolution Electrospray Ionization Mass Spectrometry Research Articles

LC-MS/MS Analysis of Reaction Products of Arginine/Methylarginines with Methylglyoxal/Glyoxal.

Methylglyoxal (MGO) and glyoxal (GO) are toxic α-dicarbonyl compounds that undergo reactions with amine containing molecules such as proteins and amino acids and result in the formation of advanced glycation end products (AGEs). This study aimed at investigating the reactivity of arginine (Arg) or dimethylarginine (SDMA or ADMA) with MGO or GO. The solutions of arginine and MGO or GO were prepared in PBS buffer (pH 7.4) and incubated at 37 °C. Direct electrospray ionization-high-resolution mass spectrometry (ESI-HRMS) analysis of the reaction mixture of Arg and MGO revealed the formation of Arg-MGO (1:1) and Arg-2MGO (1:2) products and their corresponding dehydrated products. Further liquid chromatography (LC)-MS analyses revealed the presence of isomeric products in each 1:1 and 1:2 product. The [M + H]+ of each isomeric product was subjected to MS/MS experiments for structural elucidation. The MS/MS spectra of some of the products showed a distinct structure indicative fragment ions, while others showed similar data. The types of products formed by the arginines with GO were also found to be similar to that of MGO. The importance of the guanidine group in the formation of the AGEs was reflected in similar incubation experiments with ADMA and SDMA. The structures of the products were proposed based on the comparison of the retention times and HRMS and MS/MS data interpretation, and some of them were confirmed by drawing analogy to the data reported in the literature.

Anti-Wrinkling Effect of 3,4,5-tri-O-caffeoylquinic Acid from the Roots of Nymphoides peltata through MAPK/AP-1, NF-κB, and Nrf2 Signaling in UVB-Irradiated HaCaT Cells.

Nymphoides peltata has been widely used pharmacologically in traditional Chinese medicine to treat heat strangury and polyuria. The aim of this study was to isolate the bioactive components from N. peltata and evaluate their potential use as antioxidant and anti-wrinkle agents. Phytochemical investigation of the methanolic extract of N. peltata roots led to the isolation of 15 compounds (1-15), which were structurally determined as α-spinasterol (1), 3-O-β-D-glucopyranosyl-oleanolic acid 28-O-β-D-glucuronopyranoside (2), 4-hydroxybenzoic acid (3), protocatechuic acid (4), vanillic acid (5), p-coumaric acid (6), caffeic acid (7), ferulic acid (8), neochlorogenic acid (neo-CQA) (9), chlorogenic acid (CQA) (10), cryptochlorogenic acid (crypto-CQA) (11), isochlorogenic acid B (3,4-DCQA) (12), isochlorogenic acid A (3,5-DCQA) (13), isochlorogenic acid C (4,5-DCQA) (14), and 3,4,5-tri-O-caffeoylquinic acid (TCQA) (15). Of these 15 compounds, compound 2 was a new oleanane saponin, the chemical structure of which was characterized by 1D and 2D nuclear magnetic resonance (NMR) spectroscopic data and high-resolution electrospray ionization mass spectrometry (HRESIMS), as well as chemical reaction. Biological evaluation of the isolated compounds revealed that 3,4,5-tri-O-caffeoylquinic acid (TCQA) significantly improved Nrf2 levels in an Nrf2-ARE reporter HaCaT cell screening assay. TCQA was found to potently inhibit the Nrf2/HO-1 pathway and to possess strong anti-wrinkle activity by modulating the MAPK/NF-κB/AP-1 signaling pathway and thus inhibiting MMP-1 synthesis in HaCaT cells exposed to UVB. Our results suggest that TCQA isolated from N. peltata might be useful for developing effective antioxidant and anti-wrinkle agents.

Subplenones A-J: Dimeric Xanthones with Antibacterial Activity from the Endophytic Fungus Subplenodomus sp. CPCC 401465.

Subplenones A-J (1-10), 10 new xanthone dimers, have been isolated and characterized from the endophytic fungus Subplenodomus sp. CPCC 401465, which resides within the Chinese medicinal plant Gentiana straminea. The isolation process was guided by antibacterial assays and molecular-networking-based analyses. The chemical structures of these compounds were elucidated through the interpretation of nuclear magnetic resonance (NMR) and high-resolution electrospray ionization mass spectrometry (HRESIMS) data. Furthermore, the relative configuration of the compounds was determined using NMR and single-crystal X-ray diffraction analyses, and the absolute configuration was established using electronic circular dichroism calculations. All of the isolated compounds exhibited significant inhibitory activity against Gram-positive bacteria. Notably, compounds 1, 5, and 7 displayed remarkable inhibitory activity against methicillin-resistant Staphylococcus aureus (MRSA) ATCC 700698, with a minimum inhibitory concentration (MIC) of 0.25 μg/mL, and against vancomycin-resistant Enterococcus faecium (VRE) ATCC 700221, with MIC values ranging from 0.5 to 1.0 μg/mL.

Ferrous determination in ground and brackish water by high-resolution electrospray ionization mass spectrometry

Generally, methods using nebulizers such as in Electrospray Ionization Mass Spectrometry (ESI-MS) or Induced Coupled Plasma Mass Spectrometry(ICP-MS) for the determination of Fe(II) in seawater are difficult because of the high concentration of salt. In earlier studies, we determined Fe(II) in river water with 1,10-phenanthroline (phen) and measured the complexed species by ESI-MS, but a method to reduce the interference of salts in seawater is lacking. After filtration of water samples with a 0.45 µm membrane on site, the sample was immediately added into a previously prepared solution containing sodium perchlorate (NaClO4) and phen in nitrobenzene (NB). After shaking for 20 min, Fe(II) was extracted into nitrobenzene (NB) with the perchlorate ions. The NB was introduced to an ESI-MS, using a syringe. The Fe(II)-phen complex and perchlorate ions in NB were more stable than in the aqueous solution.Approximately 99.6 % of Fe(II) was extracted into the organic phase. In our previously presented data, Fe(II) was complexed with 1,10-phenanthroline in aqueous solution, and measured by HR-ESI-MS, a standard addition method was used, and a calibration curve was obtained. In this method, the samples were added to 0.025, 0.05, 0.1 and 0.2 Fe(II)/µmol/L and a calibration curve (R2 = 0.993) of the peak intensity was determined by HR-ESI-MS from the total peak areas of [Fe(phen)₂-NB]2+m/z 269.5 and [Fe(phen)3]2+m/z 298.2. In the present study, the concentration of Fe(II) in brackish water containing 1 % salt was determined to be 0.184 ± 0.05 µM, and the total iron concentration was determined by ICP-MS and found to be 0.70 µM. Fe(II)/Fetotal was 26 %, the groundwater sample contained Fe(II) 0.146 ± 0.002 µM, Fetotal was 0.23 ± 0.02 µM, and Fe(II)/Fetotal was 63 %. Groundwater is less sensitive to oxygen and therefore has a higher proportion of Fe(II) to total iron. In brackish water, on the other hand, the proportion of Fe(II) can be higher or lower because it is affected by oxygen supply from the surface and reduction from the bottom sediment. Therefore, the Fe(II) concentration was relatively high, but in brackish water, Fe(II) was low, but Fetotal was high.

Targeted Isolation of N-Acetylcysteine-Containing Angucycline Derivatives from Streptomyces sp. MC16 and Their Antiproliferative Effects.

Liquid chromatography-mass spectrometry (LC-MS/MS)-based molecular networking analysis was applied to Streptomyces sp. MC16. The automatic classification of the MolNetEnhancer module revealed that its major constituent was an angucycline derivative. By targeted isolation of unique clusters in the molecular network, which showed different patterns from typical angucycline compounds, two new N-acetylcysteine-attached angucycline derivatives (1 and 2) were isolated. The structures were elucidated based on intensive NMR analysis and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). All isolated compounds (1-4) were tested for their inhibitory effects on the proliferation of A431, A549, and HeLa cell lines. Antibiotics 100-1 (3) and vineomycinone B2 (4) showed moderate inhibitory effects on these three cell lines with IC50 values ranging from 18.5 to 59.0 μM, while compounds 1 and 2 with an additional N-acetylcysteine residue showed weak inhibitory effects only on the HeLa cell line with IC50 values of 54.7 and 65.2 μM, respectively.

Anti-inflammatory activity via NO production inhibition of compounds from Vietnamese Lycopodium casuarinoides Spring

In our investigation of the methanol extract derived from the aerial parts of Vietnamese Lycopodium casuarinoides Spring, we successfully isolated two novel secondary metabolites: 3β,21α,24-trihydroxyserrat-14-en-29-yl-E-4′-hydroxy-3′-methoxycinnamate (1) and lycocasuarinol A (2), and eight known compounds, namely α-onocerin (3), salcolin A (4), salcolin B (5), (2 R,3 R)-dihydrokaempferol (6), apigenin (7), huperzinine (8), 4-methoxycinnamic acid (9), and (E)-p-coumaric acid (10). The structural elucidation of these compounds was accomplished through high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) and one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR) spectroscopy techniques. All isolates were tested for their potential to inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-induced RAW264.7 cells. Notably, the flavonoids (2 R,3 R)-dihydrokaempferol (6) and apigenin (7) exhibited significant inhibitory activity against NO production, with IC50 values of 3.4 µM and 3.0 µM, respectively. These values surpassed the inhibition observed with the positive control, celastrol. Furthermore, a comparison of the guaiacyl glyceryls salcolin A (4) and salcolin B (5), differing in their threo-β- and erythro-guaiacyl glyceryl moieties, revealed potential structure-activity relationships. These findings suggest that the extract from Lycopodium casuarinoides and its phytochemical constituents hold promise as potential anti-inflammatory agents

Bidesmosidic and monodesmosidic triterpenoid saponins from the grain of Amaranthus hypochondriacus

Five known bidesmosidic triterpenoid saponins (1–5) and their hydrolysates, namely two new monodesmosidic triterpenoid saponins (1′ and 2′) and three known saponins (3′–5′), were isolated from Amaranthus hypochondriacus seeds. The structures of the new compounds were determined using interpreting 1D and 2D nuclear magnetic resonance (NMR) data and high-resolution electrospray ionisation mass spectrometry (HRESI-MS) analyses as 3-O-β-D-glucuronopyranosyl-30-normedicagenic acid (1′) and 3-O-[α-L-rhamnopyranosyl (1 → 3)-β-D-glucuronopyranosyl] bayogenin (2′). For the first time, 5 and 5′ were isolated from Medicago plants as the first examples of glycosides in medicagenic acid from Amaranthus plants. While bidesmosidic saponins showed no cytotoxicity, monodesmosidic saponins, except for 1′, showed moderate cytotoxicity against human cervix adenocarcinoma (HeLa) cells. In addition, monodesmosidic saponins showed anti-fungal activity against the rice blast fungus (Pyricularia oryzae).

Macrosphelides from Antarctic fungus Pseudogymnoascus sp. (strain SF-7351) and their neuroprotective effects on BV2 and HT22 cells

Strategies for reducing inflammation in neurodegenerative diseases have attracted increasing attention. Herein, we discovered and evaluated the neuroprotective potential of fungal metabolites isolated from the Antarctic fungus Pseudogymnoascus sp. (strain SF-7351). The chemical investigation of the EtOAc extract of the fungal strain isolate revealed a novel naturally occurring epi-macrosphelide J (1), a novel secondary metabolite macrosphelide N (2), and three known compounds, namely macrosphelide A (3), macrosphelide B (4), and macrosphelide J (5). Their structures were established unambiguously using spectroscopic methods, such as one-dimensional and two-dimensional nuclear magnetic resonance (1D and 2D-NMR) spectroscopy, high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), and gauge-including atomic orbital (GIAO) NMR chemical shift calculations, with the support of the advanced statistical method DP4+. Among the isolated metabolites, the absolute configuration of epi-macrosphelide J (1) was further confirmed using single-crystal X-ray diffraction analysis. The neuroprotective effects of the isolated metabolites were evaluated in lipopolysaccharide (LPS)-induced BV2 and glutamate-stimulated HT22 cells. Only macrosphelide B (4) displayed substantial protective effects in both BV2 and HT22 cells. Molecular mechanisms underlying this activity were investigated using western blotting and molecular docking studies. Macrosphelide B (4) inhibited the inflammatory response by reducing the nuclear translocation of NF-κB (p65) in LPS-induced BV2 cells and induced the Nrf2/HO-1 signaling pathway in both BV2 and HT22 cells. The neuroprotective effect of macrosphelide B (4) is related to the interaction between Keap1 and p65. These results suggest that macrosphelide B (4), present in the fungus Pseudogymnoascus sp. (strain SF-7351), may serve as a candidate for the treatment of neurodegenerative diseases.

Ergosterol and Its Metabolites Induce Ligninolytic Activity in the Lignin-Degrading Fungus Phanerochaete sordida YK-624.

White-rot fungi are the most important group of lignin biodegraders. Phanerochaete sordida YK-624 has higher ligninolytic activity than that of model white-rot fungi. However, the underlying mechanism responsible for lignin degradation by white-rot fungi remains unknown, and the induced compounds isolated from white-rot fungi for lignin degradation have never been studied. In the present study, we tried to screen ligninolytic-inducing compounds produced by P. sordida YK-624. After large-scale incubation of P. sordida YK-624, the culture and mycelium were separated by filtration. After the separation and purification, purified compounds were analyzed by high-resolution electrospray ionization mass spectrometry and nuclear magnetic resonance. The sterilized unbleached hardwood kraft pulp was used for the initial evaluation of ligninolytic activity. Ergosterol was isolated and identified and it induced the lignin-degrading activity of this fungus. Moreover, we investigated ergosterol metabolites from P. sordida YK-624, and the ergosterol metabolites ergosta-4,7,22-triene-3,6-dione and ergosta-4,6,8(14),22-tetraen-3-one were identified and then chemically synthesized. These compounds significantly improved the lignin-degrading activity of the fungus. This is the first report on the ligninolytic-inducing compounds produced by white-rot fungi.

A New Harziane Diterpene, Harziaketal A, and a New Sterol, Trichosterol A, from the Marine-Alga-Epiphytic Trichoderma sp. Z43.

One new diterpene, harziaketal A (1), and one new highly degraded sterol, trichosterol A (2), along with three known compounds, including one diterpene, harzianone (3), and two steroids, (22E,24R)-5α,6α-epoxy-ergosta-8(14),22-dien-3β,7α-diol (4) and isoergokonin B (5), were isolated from the culture of the marine-alga-epiphytic fungus Trichoderma sp. Z43 by silica gel column chromatography (CC), Sephadex LH-20 CC, and preparative thin-layer chromatography (TLC). Their structures and relative configurations were assigned by nuclear magnetic resonance (NMR) and high resolution electrospray ionisation mass spectrometry (HR-ESI-MS) data, and the absolute configuration of 1 was established by X-ray diffraction. Compound 1 features a hemiketal unit situated at the four-membered ring of harziane-type diterpenes for the first time, while 2 represents the rare occurrence of sterols with rings A and B being degraded. Compounds 1 and 2 displayed weak inhibition against the tested phytoplankton (Amphidinium carterae, Heterocapsa circularisquama, Heterosigma akashiwo, and Prorocentrum donghaiense) with half maximal inhibitory concentration (IC50 ) ranging from 14 to 53 μg/mL.

Furanonaphthoquinones, Diterpenes, and Flavonoids from Sweet Marjoram and Investigation of Antimicrobial, Bacterial Efflux, and Biofilm Formation Inhibitory Activities.

The chloroform extract of Origanum majorana exhibited high antibacterial and antifungal activities against 12 bacterial and 4 fungal strains; therefore, it was subjected to bioassay-guided isolation to afford six compounds (1-6). The structures were determined via one- and two-dimensional nuclear magnetic spectroscopy and high-resolution electrospray ionization mass spectrometry experiments. The compounds were identified as furanonaphthoquinones [majoranaquinone (1), 2,3-dimethylnaphtho[2,3-b]furan-4,9-dione (2)], diterpenes [19-hydroxyabieta-8,11,13-trien-7-one (3), 13,14-seco-13-oxo-19-hydroxyabieta-8-en-14-al (4)], and flavonoids [sterubin (5) and majoranin (6)]. Compounds 1 and 2 were first obtained from a natural source and compounds 3 and 4 were previously undescribed. Majoranaquinone (1) exhibited a high antibacterial effect against 4 Staphylococcus, 1 Moraxella, and 1 Enterococcus strains (MIC values between 7.8 μM and 1 mM). In the efflux pump inhibition assay, majoranaquinone (1) showed substantial activity in Escherichia coli ATCC 25922 strain. Furthermore, 1 was found to be an effective biofilm formation inhibitor on E. coli ATCC 25922 and E. coli K-12 AG100 bacteria. Our findings proved that bioactivities of majoranaquinone (1) significantly exceed those of the essential oil constituents; therefore, it should also be considered when assessing the antimicrobial effects of O. majorana.

Polyporenic Acids from the Mushroom Buglossoporus quercinus Possess Chemosensitizing and Efflux Pump Inhibitory Activities on Colo 320 Adenocarcinoma Cells.

Polyporenic acids N-R (1-5), five novel 24-methylene lanostane triterpenes along with seven known polyporenic acids (6-12), were identified from the fruiting bodies of Buglossoporus quercinus. The isolation of compounds 1-12 was performed by a combination of multistep flash chromatography and reversed-phase high-performance liquid chromatography (HPLC). The structure determination was carried out by extensive spectroscopic analysis, including 1D and 2D nuclear magnetic resonance (NMR) and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) experiments. The isolated fungal metabolites were investigated for their antiproliferative activity in vitro by 3-(4,5-dimethylthiazol2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay on the resistant Colo 320 human colon adenocarcinoma cell line expressing P-glycoprotein (ABCB1). The lanostane triterpenes exerted moderate antiproliferative activity with IC50 values in the range of 20.7-106.2 μM. A P-glycoprotein efflux pump modulatory test on resistant Colo 320 cells highlighted that fungal metabolites 3, 5, 8, and 10-12 have the ability to inhibit the efflux pump activity of cancer cells. Moreover, the drug interactions of triterpenes with doxorubicin were studied by the checkerboard method. Compounds 3-4, and 7-12 interacted in a synergistic manner, while an outstanding potency was detected for compound 9, which was defined as strong synergism (CI = 0.276). The current study reveals that B. quercinus is a remarkable source of fungal steroids with considerable chemosensitizing activity on cancer cells.

Anti-leishmanial and cytotoxic compounds isolated from marine sponge Hemimycale sp.

The methanolic extract of the marine sponge Hemimycale sp. yielded two new compounds; 1-(2’-methyl heptadecyl) phenol (1) and a new pyrazole derivative; 4-(hydroxymethyl)-1H-pyrazol-3-ol (2), together with previously isolated (2′R)-2′-hydroxy-N-((2S,3S,4R)-1,3,4-trihydroxy-16-methylpentadecan-2-yl)docosanamide (3), cholesterol (4), 5, 8-epi-dioxycholest-6-en-3-ol (5) and 3-acetylsesterstatin 3 (6), which were firstly reported from family Hymedesmiidae. Their structure elucidation was based on extensive nuclear magnetic resonance spectroscopy and high resolution-electrospray ionization-mass spectrometry. The isolated compounds were evaluated for their anti-leishmanial and cytotoxic activities. Compound 5 showed remarkable anti-leishmanial activity with IC50 value of 15.8 ± 0.92 µg/mL comparable with the standard miltefosine (IC50 = 3.2 ± 0.07 µg/mL), while compound 3 exhibited noteworthy cytotoxicity against A594 cell line with IC50 value of 29.6 ± 1.68 µg/mL compared to etoposide (IC50 = 10.9 ± 1.30 µg/mL).

Six New Phenolic Glycosides from the Seeds of Moringa oleifera Lam. and Their α-Glucosidase Inhibitory Activity.

Plant-derived phytochemicals have recently drawn interest in the prevention and treatment of diabetes mellitus (DM). The seeds of Moringa oleifera Lam. are widely used in food and herbal medicine for their health-promoting properties against various diseases, including DM, but many of their effective constituents are still unknown. In this study, 6 new phenolic glycosides, moringaside B-G (1-6), together with 10 known phenolic glycosides (7-16) were isolated from M. oleifera seeds. The structures were elucidated by 1D and 2D NMR spectroscopy and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) data analysis. The absolute configurations of compounds 2 and 3 were determined by electronic circular dichroism (ECD) calculations. Compounds 2 and 3 especially are combined with a 1,3-dioxocyclopentane moiety at the rhamnose group, which are rarely reported in phenolic glycoside backbones. A biosynthetic pathway of 2 and 3 was assumed. Moreover, all the isolated compounds were evaluated for their inhibitory activities against α-glucosidase. Compounds 4 and 16 exhibited marked activities with IC50 values of 382.8 ± 1.42 and 301.4 ± 6.22 μM, and the acarbose was the positive control with an IC50 value of 324.1 ± 4.99 μM. Compound 16 revealed better activity than acarbose.

Determination of Umifenovir and Its Metabolites by High-Performance Liquid Chromatography with Combined Mass Spectrometric Detection

An approach to the highly sensitive and selective determination of the bromine-containing antiviral drug umifenovir (Arbidol) and its metabolites in natural and waste water, activated sludge, and bottom sediments by chromatography-mass spectrometry based on a combination of two techniques– inductively coupled plasma mass spectrometry (ICP MS) and electrospray ionization high-resolution mass spectrometry (ESI HRMS) is developed. Reversed-phase chromatographic separation and detection based on ESI HRMS provide the reliable detection and identification of analytes in complex matrices, while the use of ICP-MS with 79Br signal detection makes it possible to exclude matrix effects and use a single analytical standard for quantitative analysis. The use of solid-phase extraction and pressurized liquid extraction as sample preparation methods made it possible to achieve limits of detection at a level of 0.2 ng/L and 2 µg/kg for liquid and solid samples, respectively. The developed approach was successfully tested in the analysis of real samples. It is shown that the concentrations of analytes in urban wastewaters are in the range 4.4–260 ng/L. The highest concentration (up to 3.7 mg/kg) is characteristic of activated sludge, which acts as an effective sorbent for umifenovir and its transformation products.

Dryoptkirbioside, A New Fructofuranoside glycerol, and Other Constituents from Dryopteris kirbi Hook et Grav Rhizomes.

A new fructofuranoside glycerol, dryoptkirbioside (1), along with fourteen known compounds (2-15), was isolated from the MeOH extract of Dryopteris kirbi rhizomes by silica gel column chromatography, Sephadex LH-20 column chromatography, and semipreparative HPLC. The structure of the new compound was determined by analyses of its spectroscopic data including nuclear magnetic resonance (NMR), and high-resolution electrospray ionisation mass spectrometry (HRESI-MS) and chemical conversions. The n-hexane-soluble portion and the EAFA fraction showed strong activities against lung (A549), breast (MCF-7), and cervical (HeLa) human cancer cell lines (IC50 values ranging from 4.0 to 8.8 µg/mL). Aspidinol P (5) and aspidinol B (6) exhibited moderate to low cytotoxicity on the three cell lines (IC50 values ranging from 20.4 to 58.7 µM). The MeOH extract and n-hexane-soluble portion had excellent activities against Staphylococcus aureus and Bacillus subtilis (MICs 11.7 and 23.4 µg/mL), whereas the EtOAc- and n-BuOH-soluble portions were significantly active on S. aureus (MICs 46.9 and 93.8 µg/mL). The main fractions EAFB, EAFC and nBFB displayed excellent activity against S. aureus (MICs 11.7 and 23.4 µg/mL). Aspidinol B (6) had significant activity, while aspidinol P (5) was moderately active against S. aureus and B. subtilis (MICs 42.0 and 89.5 µM).

Sarcodonol A–D from fruiting bodies of Sarcodon imbricatus inhibits HCoV-OC43 induced apoptosis in MRC-5 cells

Four new 26-carboxylated ergostane-type sterols (Sarcodonol A–D) were isolated from 70% ethanol extracts of dried fruiting bodies of Sarcodon imbricatus. Their chemical structures were elucidated using 1D- and 2D-nuclear magnetic resonance and high-resolution electrospray ionization mass spectrometry, and confirmed by comparison with previously reported data. As far as we know, this is the first instance of isolating a 26-carboxylated ergostane-type sterol from nature. The determined antiviral efficacy of sarcodonol A–D (1–4) against HCoV-OC43 in MRC-5 cells confirmed that sarcodonol D (4) had significant antiviral activity. Notably, sarcodonol D (4) potently blocked virus infection at low-micromolar concentration and showed high SI (IC50 = 2.26 μM; CC50 > 100 μM; SI > 44.2). In addition, this research shows that the antiviral effect of sarcodonol D (4) via reduced apoptosis increased by viral infection is through mitochondrial stress regulation. This suggests that sarcodonol D (4) is a potential candidate for use as an antiviral treatment.

Identification of compounds using HPLC-QTOF-MS online antioxidant activity mapping from aerial parts of Ligularia stenocephala

Inflammation, diabetes, and even malignancies are pharmacological effects connected by antioxidant capacity and free radicals. Many antioxidants scavenge free radicals originating from dietary sources such as fruits, vegetables, and teas. To identify the bioactive components of Ligularia stenocephala, an effective method combining HPLC-QTOF-MS and bioactivity evaluation was investigated for the first time. Antioxidant agents were isolated from L. stenocephala, a folk medicine used for edema and scrofula in Korea, Japan, and China. The phytochemical investigation of the aerial parts of L. stenocephala resulted in the separation and determination of six compounds (1–6). In particular, the chemical structures were identified as hyperoside (1), 3,5-dicaffeoylquinic acid (2), 3,5-dicaffeoylquinic acid methyl ester (3), trifolin (4), rutin (5), and 3,4-dicaffeoylquinic acid (6). Their structures were identified using 1D and 2D NMR spectroscopy and high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) data analysis. The results showed that phenolic components were responsible for the antioxidant inhibitory activity of L. stenocephala. Additionally, to understand the mechanisms of the antioxidant inhibitory activity of L. stenocephala, a docking simulation study was performed to support the in vitro results. Taken together, this new method is rapid, inexpensive, and can be applied to identify the active components of medicinal herbs without separation.

Two New Compounds from Allii Macrostemonis Bulbus and Their In Vitro Antioxidant Activities.

Two new compounds named 4,4'-bis(β-D-glucopyranosyloxy)biphenyl (1) and spirostane-25(27)-en-2α,3β-diol-3-O-β-D-xylopyranosyl(1→3)-β-D-glucopyranosyl(1→4)-β-D-galactopyranoside (2) were isolated from n-butanol extraction part of 80% ethanol extract of Allii Macrostemonis Bulbus. Alongside these, ten known compounds (3-12) were also identified, including a flavonoid glycoside (3), seven steroids (4-10), a nucleoside (11), and a phenylpropanoid glycoside (12) were found. Notably, compounds 3-6 were isolated from this plant for the first time. The structures of all compounds were confirmed using high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), 1D, and 2D NMR spectroscopy. Some of these compounds showed strong antioxidant activity, and compound 1 demonstrated the most potent reduction of ferric ions (Fe3+) with an IC50 value of 0.59 ± 0.18 mg/mL. Compounds 2 and 3 exhibited the highest scavenging activity against superoxide anion radicals (O2-·) with an IC50 value of 0.02 ± 0.01 mg/mL. Additionally, compound 3 displayed substantial scavenging activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) with IC50 values of 0.21 ± 0.17 mg/mL and 0.02 ± 0.01 mg/mL, respectively. The discovery of these two new compounds is a reference for identifying Allii Macrostemonis Bulbus quality markers. Moreover, their exceptional antioxidant activity offers a promising avenue for uncovering novel natural antioxidants.

Identification of glycine betaine as a host-derived molecule required for the vegetative proliferation of the protozoan parasite Perkinsus olseni.

Perkinsus olseni is an industrially significant protozoan parasite of Manila clam, Ruditapes philippinarum. So far, various media, based on Dulbecco's Modified Eagle Medium and Ham's F-12 nutrient mixture with supplementation of fetal bovine serum (FBS), have been developed to proliferate the parasitizing trophozoite stage of P. olseni. The present study showed that P. olseni did not proliferate in FBS-deficient Perkinsus broth medium (PBMΔF), but proliferated well in PBMΔF supplemented with tissue extract of host Manila clams, indicating that FBS and Manila clam tissue contained molecule(s) required for P. olseni proliferation. Preliminary characterization suggested that the host-derived molecule(s) was a heat-stable molecule(s) with a molecular weight of less than 3 kDa, and finally a single molecule required for the proliferation was purified by high-performance liquid chromatography processes. High-resolution electrospray ionization mass spectrometry and nuclear magnetic resonance analyses identified this molecule as glycine betaine (=trimethylglycine), and the requirement of this molecule for P. olsseni proliferation was confirmed by an assay using chemically synthesized, standard glycine betaine. Although glycine betaine was required for the proliferation of all examined Perkinsus species, supplementation of glycine betaine precursors, such as choline and betaine aldehyde, enhanced the proliferation of 4 Perkinsus species (P. marinus, P. chesapeaki, P. mediterraneus and P. honshuensis), but not of 2 others (P. olseni and P. beihaiensis). Thus, it was concluded that the ability to biosynthesise glycine betaine from its precursors varied among Perkinsus species, and that P. olseni and P. beihaiensis lack the ability required to biosynthesize glycine betaine for proliferation.