Preparative High-performance Liquid Chromatography Research Articles

Establishing aqueous two-phase flotation coupled with preparative high performance liquid chromatography and its application for the purification of astragalin from Flaveria bidentis

The strategy of aqueous two-phase flotation (ATPF) followed by preparative high performance liquid chromatography (prep-HPLC) was established and used for the separation of astragalin from Flaveria bidentis. In the ATPF, the effects of sublation solvent, solution pH, (NH4)2SO4 concentration in aqueous solution, cosolvent, N2 flow rate, flotation time and volumes of the PEG phase on the recovery of astragalin were investigated in detail, and the optimal conditions of ATPF were selected: 50 wt% PEG1000 ethanol solvent as the flotation solvent, pH 4, 350 g/L of (NH4)2SO4 concentration in 5 % ethanol aqueous phase, 40 mL/min of N2 flow rate, 30 min of flotation time, 10.0 mL of flotation solvent volume and twice. After ATPF enrichment, the flotation product was further purified by prep-HPLC. As determined by HPLC, the purity of astragalin was 98.8 %.

An efficient method for the preparative isolation and purification of alkaloids from Gelsemium by using high speed counter-current chromatography and preparative HPLC

We established an efficient method using high-speed countercurrent chromatography (HSCCC) combined with preparative high-performance liquid chromatography (prep-HPLC) for isolating and purifying Gelsemium elegans (G. elegans) alkaloids. First, the two-phase solvent system composed of 1% triethylamine aqueous solution/n-hexane/ethyl acetate/ethanol (volume ratio 4:2:3:2) was employed to separate the crude extract (350 mg) using HSCCC. Subsequently, the mixture that resulted from HSCCC was further separated by Prep-HPLC, resulting in seven pure compounds including: 14-hydroxygelsenicine (1, 12.1 mg), sempervirine (2, 20.8 mg), 19-(R)-hydroxydihydrogelelsevirine (3, 10.1 mg), koumine (4, 50.5 mg), gelsemine (5, 32.2 mg), gelselvirine (6, 50.5 mg), and 11-hydroxyhumanmantenine (7, 12.5 mg). The purity of these seven compounds were 97.4, 98.9, 98.5, 99, 99.5, 96.8, and 85.5%, as determined by HPLC. The chemical structures of the seven compounds were analyzed and confirmed by electrospray ionization mass spectrometry (ESI-MS), 1H-nuclear magnetic resonance (1H NMR), and 13 C-nuclear magnetic resonance (13 C NMR) spectra. The results indicate that the HSCCC-prep-HPLC method can effectively separate the major alkaloids from the purified G. elegans, holding promising prospects for potential applications in the separation and identification of other traditional Chinese medicines.

Isolation and Identification of Bitter Compounds in Ginseng (Panax ginseng C. A. Mey.) Based on Preparative High Performance Liquid Chromatography, UPLC-Q-TOF/MS and Electronic Tongue

As a traditional Chinese medicinal herb, ginseng (Panax ginseng C. A. Mey.) is commonly used to treat common diseases, for example, esophageal cancer and myasthenia gravis. Furthermore, ginseng is also processed into a functional food additive that is utilized to improve the freshness of chicken soup and make health wine. Unfortunately, ginseng (Panax ginseng C. A. Mey.) has already shown a noticeable bitterness during its application process. In this research, the bitter substances in ginseng (Panax ginseng C. A. Mey.) after two common preparation processes (water extraction and ethanol extraction) were separated, purified and identified by preparative high performance liquid chromatography (prep-HPLC), high performance liquid chromatography with diode array detector (HPLC-DAD), ultra-performance liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) and an electronic tongue. The results indicated that compared with the other four bitter compounds, the ginsenoside Rb1 had the highest bitterness value, followed by 20(S)-ginsenoside Rg2, ginsenoside Rg1, ginsenoside Rf and ginsenoside Rb3. Upon the evaluation of results to reduce the bitterness of ginseng extract, we found that the composite embedding system of chitosan adsorption in the ginseng carrageenan gel microsphere (K/MC/MCG) could effectively reduce the bitterness.

Self-forming barrier layer for carbon dioxide corrosion protection of mild steel in NaCl solution containing hydroxychavicol isolated from Piper betle L. leaf

Hydroxychavicol compound was isolated from Piper betle L. leaf and identified via preparative high performance liquid chromatography (Prep-HPLC) and nuclear magnetic resonance (NMR) spectroscopy. Hydroxychavicol compound was demonstrated to be a potential inhibitor for mild steel corrosion in carbon dioxide-containing environments. This is due to the formation of a barrier layer on the steel surface. Scanning electron microscopy and atomic force microscopy results indicated that the inclusion of hydroxychavicol in the aggressive environment made steel surfaces more homogeneous and smoother with a rough mean square roughness of 7.78 nm, whereas the steel surface was severely corroded in the blank solution. Moreover, X-ray photoelectron spectroscopy results showed an enrichment of hydroxychavicol species combined with iron oxides and hydroxides on the mild steel surface. This can lead to the creation of a protective layer on the mild steel surface, thereby enhancing corrosion protection and localized inhibition. The work suggested a novel approach to isolate corrosion inhibitors from natural products, particularly agricultural by-products for mild steel in carbon dioxide and chloride ion environment with high inhibition performance.

Chemical profiling, anticonvulsant and anxiolytic effects of the smoke constituents isolated from Leonotis leonurus (L.) R.Br.

The use of medicinal plants for central nervous system (CNS)-related ailments, such as epilepsy and anxiety, is prevalent in South Africa. Plants from the Lamiaceae family are commonly used for their therapeutic benefits. Leonotis leonurus (L.) R.Br. has been reported in ethnobotanical literature to have anticonvulsant and anxiolytic effects through the inhalation of pyrolysis products obtained by combustion of the aerial parts. To explore the chemical profiles and CNS activity of the smoke extract and isolated constituents of L. leonurus in zebrafish larvae, through anticonvulsive and anxiolytic activity assays. The smoke extract of L. leonurus was obtained through the combustion of the aerial parts of the plant using a custom-built smoke recovery apparatus. The chemical profile of the smoke constituents was determined using the Ultra-Performance Liquid Chromatography coupled with Mass Spectrometry (UPLC-MS). Targeted compounds were subjected to preparative High-Performance Liquid Chromatography for separation before structure elucidation using Nuclear Magnetic Resonance (NMR). The maximum tolerated concentrations, as well as the anxiolytic activity of the smoke extract were determined in five days post fertilisation zebrafish larvae. Reverse-thigmotaxis and locomotor activity of larvae in the light/dark transition assay were used to determine anxiolytic activity. Zebrafish larvae at six days post fertilisation were subjected to several concentrations of the smoke constituents of L. leonurus. The baseline locomotor activity of the larvae was tracked for 30 min, prior to addition of pentylenetetrazole (PTZ) to induce seizure-like behaviour in the larvae, after which the locomotor activity of the larvae was once again tracked for an additional 30 min. and Discussion: The UPLC-MS profiles of the smoke extract revealed the presence of two main compounds, leoleorin A and leoleorin B, which were targeted and isolated. Upon subjection to NMR spectroscopy for structure elucidation, the compounds were confirmed to be labdane diterpenoids. Both leoleorin A and leoleorin B, and the smoke extract displayed suppression of the PTZ induced seizure-like behaviour in six days post fertilisation (dpf) in zebrafish larvae. Under light and dark conditions, the smoke extract and compounds displayed potential anxiolytic activity at different concentrations. Our results suggest that the smoke constituents of L. leonurus may exert anticonvulsant and anxiolytic effects which align with the traditional indications and the mode of administration.

Stereochemical Heterogeneity Analysis of Polylactides by Multidimensional Liquid Chromatography.

A new and robust high-performance liquid chromatography (HPLC) method that separates poly(lactic acid) (PLA) according to its stereochemical composition is presented. Using this method, poly(l-lactide) incorporating trace amounts of meso-lactide resulting from the racemization is separated from the pristine polymer. To prove this aspect in more detail, a representative poly(l-lactic acid) standard, assumed to be highly homogeneous, was separated using this method. The result showed that this was not the case as a fraction incorporating meso-lactide due to racemization occurring during the synthesis is separated. Employing two-dimensional liquid chromatography (2D-LC), the molar mass differences of the separated species were investigated, and fractions with similar molecular sizes were detected, confirming that the LC separation is solely based on stereochemical heterogeneity. The sample was further fractionated by preparative HPLC, followed by an in-depth analysis of the fractions using homonuclear decoupling in proton nuclear magnetic resonance (1H NMR). Convincing results that unveiled significant differences in the stereochemistry of the isolated PLA fractions were obtained. Subsequent analysis by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) also confirmed oligomer series with different end group structures, indicating that the applied HPLC method is very sensitive to minor variations in stereochemistry and end groups. This integrated approach offers detailed insight into the structural characteristics of PLA polymers, contributing to a better understanding of their composition and potential applications.

Hydrophobic Components in Light-Yellow Pulp Sweet Potato (Ipomoea batatas (L.) Lam.) Tubers Suppress LPS-Induced Inflammatory Responses in RAW264.7 Cells via Activation of the Nrf2 Pathway.

Sweet potato is a crop that is widely consumed all over the world and is thought to contribute to health maintenance due to its abundant nutrients and phytochemicals. Previous studies on the functionality of sweet potatoes have focused on varieties that have colored pulp, such as purple and orange, which contain high levels of specific phytochemicals. Therefore, in the present study, we evaluated the anti-inflammatory effects of light-yellow-fleshed sweet potatoes, which have received little attention. After freeze-drying sweet potatoes harvested in 2020, extracts were prepared from the leaves, stems, roots, and tubers in 100% ethanol. Mouse macrophage-like cell line RAW264.7 cells were cultured with 10 µg/mL of the extracts and induced lipopolysaccharide (LPS)-stimulated inflammation. Of the extracts, the tuber extracts showed the highest suppression of LPS-induced interleukin-6 (IL-6) gene expression and production in RAW264.7, which was attributed to the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) oxidative stress response pathway. In addition, preparative high-performance liquid chromatography (HPLC) experiments suggested that hydrophobic components specific to the tuber were the main body of activity. In previous studies, it has been shown that the tubers and leaves of sweet potatoes with colored pulp exhibit anti-inflammatory effects due to their rich phytochemicals, and our results show that the tubers with light-yellow pulp also exhibit the effects. Furthermore, we were able to show a part of the mechanism, which may contribute to the fundamental understanding of the treatment and prevention of inflammation by food-derived components.

Bioprospection of Tenellins Produced by the Entomopathogenic Fungus Beauveria neobassiana.

Fungi are known as prolific producers of bioactive secondary metabolites with applications across various fields, including infectious diseases, as well as in biological control. However, some of the well-known species are still underexplored. Our current study evaluated the production of secondary metabolites by the entomopathogenic fungus Beauveria neobassiana from Thailand. The fermentation of this fungus in a liquid medium, followed by preparative high-performance liquid chromatography (HPLC) purification, resulted in the isolation of a new tenellin congener, namely pretenellin C (1), together with five known derivatives (2-6). Their chemical structures were elucidated by 1D and 2D nuclear magnetic resonance (NMR) spectroscopy in combination with high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). We evaluated the antimicrobial and cytotoxic activities from all isolated compounds, as well as their inhibitory properties on biofilm formation by Staphylococcus aureus. Generally, tenellins displayed varying antibiofilm and cytotoxic effects, allowing us to propose preliminary structure-activity relationships (SARs). Among the tested compounds, prototenellin D (2) exhibited the most prominent antibiofilm activity, while its 2-pyridone congener, tenellin (4), demonstrated potent cytotoxic activity against all tested cell lines. Given the fact that the biological activities of the tenellins have so far been neglected in the past, our study could provide a good starting point to establish more concise structure-activity relationships in the near future.

Preparation and characterization of the aflatoxin B1 purity certified reference material (GBW (E) 100599)

Aflatoxin B1 (AFB1), the most harmful mycotoxin, is widely distributed in many important foodstuffs, especially in peanuts, maize, and oil products. AFB1 certified reference material (CRM), essential for an accurate and reliable detection, is not easily obtained due to the difficulty of preparing high purity candidate. In this study, the high-purity AFB1 candidate was prepared by the preparative high performance liquid chromatography (Pre-HPLC) after high-yields of AFB1 on the maize medium by Aspergillus flavus. The structure of AFB1 was successively characterized by UV, IR, LC-TOF-MS/MS, 1H NMR, and 13C NMR. According to the principle of ISO 17034: 2016 and ISO guide 35: 2017, the purity of the candidate was accurately determined by the mass balance method (MB) and the quantitative nuclear magnetic resonance spectroscopy method (qNMR), while the homogeneity, long-term stability, short-term stability, and uncertainty were systematically studied by the MB method. Under the optimum conditions, the certified purity value for the AFB1 CRM was 99.53 % with an extended uncertainty of 0.56 % (k = 2, 95 % confidence interval), which was also sufficiently homogeneous and stable for 6 months at 4 °C and −20 °C. After approval by State Administration for Market Regulation, a new AFB1 purity CRM (GBW (E) 100599) was finally prepared. This is the first report for the preparation of a reliable AFB1 purity CRM, benefit for enhancing the accuracy, traceability, and comparability of measurements of AFB1 in various foodstuffs, and finally ensure the food quality and safety.

Melinjo Seeds (Gnetum gnemon L.) Antioxidant Activity and Cytotoxic Effects on MCF-7 Breast Cancer Cells: A Study Based on Tracing of Resveratrol Compound.

Breast cancer is one of the deadliest cancers worldwide. One of the polyphenols, namely, resveratrol, has been proven to have anticancer activity. Melinjo seeds which contain resveratrol need to be tested for their potential as an anti-breast cancer agent. This study aims to determine the antioxidant activity and cytotoxic effect of melinjo seeds based on solvent variations and resveratrol tracing. Extraction of melinjo seeds was performed using the soxhletation method. Antioxidant test was performed using the 2,2-diphenyl-1-picrylhydrazil method. The in vitro cytotoxic test was carried out using the microtetrazolium method. Cytotoxic test was carried out on MCF-7 breast cancer cells using a concentration range of melinjo seeds between 31,25 and 1000 μg/mL. Antioxidant and anticancer potentials are expressed in inhibitory concentration (IC)50 values. Resveratrol was traced using preparative high-performance liquid chromatography (Prep-HPLC). Melinjo seed ethanol extract provided the largest total phenolics (126,154 ± 0,865 mg GAE/g sample) and total flavonoids (44,576 ± 0,611 mg QE/g sample) among all solvent fractions. The antioxidant activity of melinjo seeds from ethanol extract, n-hexane fraction, ethyl acetate fraction, and ethanol fraction was 263,307 ppm, 317,595 ppm, 160,878 ppm, and 181,159 ppm, respectively. The ethyl acetate fraction of melinjo seeds showed the strongest cytotoxic effect (94.6 μg/mL) among all extracts and solvent fractions. Prep-HPLC showed that the ethanol extract of melinjo seeds contained resveratrol, while the ethanol and ethyl acetate fractions of melinjo seeds were thought to contain resveratrol derivatives. The antioxidant activity of melinjo seeds showed a cytotoxic effect on MCF-7 cells, which varied based on solvent polarity and total phenolic and total flavonoid. The ethyl acetate fraction which is thought to contain resveratrol derivatives provides the most potent antioxidant activity and cytotoxic effect. These results indicate that melinjo seeds containing resveratrol and its derivatives have the potential for anticancer of the breast. Further studies are still needed in determining the structure of resveratrol compounds and their derivatives to ensure their biological activity and mechanism of action.

New isoindolinone derivatives isolated from the fruiting bodies of the basidiomycete Hericium coralloides

Preparative high-performance liquid chromatography (HPLC) purification of the ethyl acetate extract derived from dried basidiomes of the European mushroom Hericium coralloides led to the identification of two previously undescribed isoindolinone derivatives named corallocins D and E (1-2). The structures of the compounds were elucidated based on HR-ESI-MS (high-resolution electron spray ionization mass spectrometry), interpretation of 1D and 2D NMR spectra, electronic circular dichroism (ECD) experiments, and comparisons with published and theoretical data. The metabolites were tested for their cytotoxic and antimicrobial effects in vitro where weak to moderate biological effects were observed against HeLa cells (KB 3.1), Mucor hiemalis and Bacillus subtilis.

The Identification and Role of the Key Mycotoxin of Pestalotiopsis kenyana Causing Leaf Spot Disease of Zanthoxylum schinifolium.

Leaf spot is a common disease of Zanthoxylum schinifolium (Z. schinifolium), which can seriously harm the plant's ability to grow, flower, and fruit. Therefore, it is important to identify the mechanism of leaf spot caused by Pestalotiopsis kenyana (P. kenyana) for thorough comprehension and disease control. In this study, to verify whether the mycotoxins produced by P. kenyana cause leaf spot disease, the best medium for P. kenyana, namely PDB, was used. The mycotoxins were determined by ammonium sulfate precipitation as non-protein substances. The crude mycotoxin of P. kenyana was prepared, and the optimal eluent was eluted with petroleum either/ethyle acetate (3:1, v/v) and purified by silica gel column chromatography and preparative high-performance liquid chromatography to obtain the pure mycotoxins PK-1, PK-2, and PK-3. The PK-3 had the highest toxicity to Z. schinifolium, which may be the primary mycotoxin, according to the biological activity test using the spray method. The physiological and biochemical indexes of Z. schinifolium plants treated with PK-3 mycotoxin were determined. Within 35 days after mycotoxin treatment, the results showed that the protein content and malondialdehyde content of leaves increased over time. The soluble sugar and chlorophyll content decreased over time. The superoxide dismutase activity and catalase activity of the leaves increased first and then decreased, and the above changes were the same as those of Z. schinifolium inoculated with the spore suspension of the pathogen. Therefore, it is believed that the mycotoxin pestalopyrone could be a virulence factor that helps P. kenyana induce the infection of Z. schinifolium. In this study, the pathogenic mechanism of Z. schinifolium leaf spot was discussed, offering a theoretical foundation for improved disease prevention and control.

Identification of novel umami peptides from yeast extract and the mechanism against T1R1/T1R3.

Yeast extract was separated by using ultrafiltration, gel filtration chromatography, and preparative high-performance liquid chromatography for analyzing the umami mechanism. 13 kinds of umami peptides were screened out from 73 kinds of peptides which were identified in yeast extract using nanoscale ultra-performance liquid chromatography-tandem mass spectrometry and virtual screening. The umami peptides were found to have a threshold range of 0.07-0.61mM. DWTDDVEAR exhibited a strong umami taste with a pronounced enhancement effect for monosodium glutamate. Molecular docking studies revealed that specific amino acid residues in the T1R1 subunit, including Arg316, Ser401, and Asp315, played a critical role in the umami perception with these peptides. Overall, the study highlights the potential of natural flavor enhancers and provides insights into the mechanism of umami taste perception.

Separation and antioxidant activities of new acetylated EGCG compounds

Acetylation could improve the bioavailability of (-)-Epigallocatechin-3-Gallate (EGCG), but the relationship of substitution degree and antioxidant capacity of acetylated EGCG was unclear. The acetylated EGCG products were separated by preparation high performance liquid chromatography (HPLC). Two mono substituted acetylated EGCG, three substituted acetylated EGCG (T-AcE), eight substituted acetylated EGCG (E-AcE) and (-)-Epigallocatechin gallate (EGCG) were isolated. The 7-acetyl-EGCG (S7-ACEGCG) and 7-acetyl-EGCG (T-AcE) were identified for the first time. The antioxidant capacity, superoxide anion radical scavenging capacities, and hydroxyl radical scavenging capacities of EGCG decreased significantly after acetylation modification. The more EGCG acetylation modification sites, the lower the total antioxidant capacity, superoxide anion radical scavenging capacities, and hydroxyl radical scavenging capacities. The antioxidant capacity, superoxide anion radical scavenging capacities, and hydroxyl radical scavenging capacities of 5-acetyl-EGCG (S5-ACE) were higher than 7-acetyl-EGCG (S7-AcE). Combining all the results in this and previous studies, acetylation modification is not conducive to the performance of EGCG antioxidant capacity.

Discovery of Anti-Coronavirus Cinnamoyl Triterpenoids Isolated from Hippophae rhamnoides during a Screening of Halophytes from the North Sea and Channel Coasts in Northern France

The limited availability of antiviral therapy for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spurred the search for novel antiviral drugs. Here, we investigated the potential antiviral properties of plants adapted to high-salt environments collected in the north of France. Twenty-five crude methanolic extracts obtained from twenty-two plant species were evaluated for their cytotoxicity and antiviral effectiveness against coronaviruses HCoV-229E and SARS-CoV-2. Then, a bioguided fractionation approach was employed. The most active crude methanolic extracts were partitioned into three different sub-extracts. Notably, the dichloromethane sub-extract of the whole plant Hippophae rhamnoides L. demonstrated the highest antiviral activity against both viruses. Its chemical composition was evaluated by ultra-high performance liquid chromatography (UHPLC) coupled with mass spectrometry (MS) and then it was fractionated by centrifugal partition chromatography (CPC). Six cinnamoyl triterpenoid compounds were isolated from the three most active fractions by preparative high-performance liquid chromatography (HPLC) and identified by high resolution MS (HR-MS) and mono- and bi-dimensional nuclear magnetic resonance (NMR). Specifically, these compounds were identified as 2-O-trans-p-coumaroyl-maslinic acid, 3β-hydroxy-2α-trans-p-coumaryloxy-urs-12-en-28-oic acid, 3β-hydroxy-2α-cis-p-coumaryloxy-urs-12-en-28-oic acid, 3-O-trans-caffeoyl oleanolic acid, a mixture of 3-O-trans-caffeoyl oleanolic acid/3-O-cis-caffeoyl oleanolic acid (70/30), and 3-O-trans-p-coumaroyl oleanolic acid. Infection tests demonstrated a dose-dependent inhibition of these triterpenes against HCoV-229E and SARS-CoV-2. Notably, cinnamoyl oleanolic acids displayed activity against both SARS-CoV-2 and HCoV-229E. Our findings suggest that Hippophae rhamnoides could represent a source of potential antiviral agents against coronaviruses.

Pharmacokinetics of cyanidin-3-O-galactoside and cyanidin-3-O-arabinoside after intravenous administration in rats

Cyanidin-3-O-galactoside and cyanidin-3-O-arabinoside (purity >98%) were isolated from black chokeberry by preparative high-performance liquid chromatography, and an animal experiment was conducted to investigate the pharmacokinetics of two anthocyanin monomers after intravenous administration. The results showed that cyanidin-3-O-galactoside has preferable druggability than cyanidin-3-O-arabinoside in pharmacokinetic area.

Bioactive Compounds from an Endophytic Pezicula sp. Showing Antagonistic Effects against the Ash Dieback Pathogen.

A fungal endophyte originating from the Canary Islands was identified as a potent antagonist against the fungal phytopathogen Hymenoscyphus fraxineus, which causes the devastating ash dieback disease. This endophyte was tentatively identified as Pezicula cf. ericae, using molecular barcoding. Isolation of secondary metabolites by preparative high-performance liquid chromatography (HPLC) yielded the known compounds CJ-17,572 (1), mycorrhizin A (3) and cryptosporioptides A-C (4-6), besides a new N-acetylated dihydroxyphenylalanin derivative 2, named peziculastatin. Planar structures were elucidated by NMR and HRMS data, while the relative stereochemistry of 2 was assigned by H,H and C,H coupling constants. The assignment of the unknown stereochemistry of CJ-17,572 (1) was hampered by the broadening of NMR signals. Nevertheless, after semisynthetic conversion of 1 into its methyl derivatives 7 and 8, presumably preventing tautomeric effects, the relative configuration could be assigned, whereas comparison of ECD data to those of related compounds determined the absolute configuration. Metabolites 1 and 3 showed significant antifungal effects in vitro against H. fraxineus. Furthermore, 4-6 exhibited significant dispersive effects on preformed biofilms of S. aureus at concentrations up to 2 µg/mL, while the biofilm formation of C. albicans was also inhibited. Thus, cryptosporioptides might constitute a potential source for the development of novel antibiofilm agents.

Metabolomics insights into the polyketide-lactones produced by Diaporthe caliensis sp. nov., an endophyte of the medicinal plant Otoba gracilipes.

Plants of the genus Otoba have been the basis for the treatment of tropical diseases in indigenous communities of countries like Colombia. Despite the lack of knowledge about their bioactive principles, endophytic fungi derived from medicinal plants are a prolific source of innovative chemistry. We systematically investigated the secondary metabolite production of a previously undescribed species of Diaporthe, herein introduced as Diaporthe caliensis sp. nov., using different metabolomics approaches together with classical chemical screening. To get an outline of the chemical space produced by this fungus, an exploratory molecular networking (MN) analysis was undertaken. A major molecular family was found to contain the known 10-membered lactone phomol (1), together with other putative congeners as compound 3. After isolation by preparative high-performance liquid chromatography, we confirmed phomol (1) as the main reason for the antimicrobial activity of the crude extract. The unknown absolute configuration of 1 was determined by the synthesis of α-methoxy-α-trifluoromethylphenylacetyl (MTPA)-esters and chemical degradation experiments. Moreover, caliensolides A (2) and B (3) were isolated, and their structures were elucidated as novel butenolides structurally unrelated to 1. Overall, the initial MN analysis incorrectly clustered compounds 1 and 3 within a single molecular family, despite evident differences in chemical structures and biosynthetic origin. Contrariwise, the unsupervised substructure discovery algorithm MS2LDA provided a deeper understanding of the fragmentation patterns and correctly clustered the polyketide-lactones produced by D. caliensis sp. nov. Our findings encourage the exploration of Colombian fungal diversity, which as demonstrated here could result in the discovery of new natural products.IMPORTANCEThe integration of metabolomics-based approaches into the discovery pipeline has enabled improved mining and prioritization of prolific secondary metabolite producers such as endophytic fungi. However, relying on automated untargeted analysis tools might lead to misestimation of the chemical complexity harbored in these organisms. Our study emphasizes the importance of isolation and structure elucidation of the respective metabolites in addition to deep metabolome analysis for the correct interpretation of untargeted metabolomics approaches such as molecular networking. Additionally, it encourages the further exploration of endophytic fungi from traditional medicinal plants for the discovery of natural products.

[A new allo-aromadendrane sesquiterpene from Dendrobium nobile].

To study the chemical constituents in the non-alkaloid part of stems of Dendrobium nobile. The macroporous adsorption resin, MCI, silica gel, RP-C_(18), and Sephadex LH-20 gel, preparative thin layer chromatography, and preparative high-performance liquid chromatography(HPLC) were used to isolate and purify the compounds. The structures of the compound were determined according to the spectra data, physicochemical properties, and relevant references. A total of 8 compounds were isolated from D. nobile, which were soltorvum F(1), p-hydroxyphenylpropionic acid(2), vanillic acid(3), p-hydroxybenzoic acid(4), N-trans-cinnamic acid acyl-p-hydroxybenzene ethylamine(5),(+)-(1R,2S,3R,4S,5R,6S,9R)-2,11,12-trihydroxypicrotoxane-3(15)-lactone(6), dendronobilin H(7), soltorvum E(8). Compound 1 was a novel compound, named as soltorvum F. Compound 8 was isolated from Dendrobium species for the first time.

[Chemical constituents from whole herb of Hedyotis scandens].

This study aimed to investigate the chemical constituents in the water extract of the whole herb of Hedyotis scandens by silica gel, ODS, and MCI column chromatographies together with preparative high-performance liquid chromatography(HPLC). The structures of isolated constituents were identified by NMR, HR-ESI-MS, etc. Thirteen compounds were isolated and identified as methyl 4-benzoyloxy-3-methoxybenzeneacetate(1), 4-benzoyloxy-3-methoxybenzeneacetic acid(2), 3-(4-hydroxy-3-methoxyphenyl)-propanoic acid(3), salicylic acid(4), 3-hydroxy-4-methoxypyridine(5), syringic acid(6), hydroxycinnamic acid(7),(R)-6-methyl-4,6-bis(4-methylpent-3-enyl)cyclohexa-1,3-dienecarbaldehyde(8), 1,2-bis(4-hydroxy-3-methoxyphenyl)-1,3-propanediol(9), 1H-indole-3-carboxaldehyde(10), isoscopoletin(11), syringaresinol(12), and pinoresinol(13). Among them, compounds 1 and 2 were new phenolic acid compounds, compounds 3-5, 8-11, and 13 were isolated from this genus for the first time, and compounds 6, 7, and 12 were obtained from H. scandens for the first time. The activity test showed that compounds 1 and 10 had a certain inhibitory effect on Mycobacterium smegmatis, with MIC_(50) values of 58.5 and 33.3 μg·mL~(-1), respectively.