HPLC Retention Times Research Articles

Segment-Based Peptide Design Reveals the Importance of N-Terminal High Cationicity for Antimicrobial Activity Against Gram-Negative Pathogens.

Host defense antimicrobial peptides (AMPs) are recognized candidates to develop a new generation of peptide antibiotics. While high hydrophobicity can be deployed in peptides for eliminating Gram-positive bacteria, high cationicity is usually observed in AMPs against Gram-negative pathogen. This study investigates how the sequence distribution of basic amino acids affects peptide activity. For this purpose, we utilized human cathelicidin LL-37 as a template and designed four highly selective ultrashort peptides with similar length, net charge, and hydrophobic content. LL-10 + , RK-9 + , KR-8 + , and RIK-10 + showed similar activity against methicillin-resistant Staphylococcus aureus in vitro and comparable antibiofilm efficacy in a murine wound model. However, these peptides showed clear activity differences against Gram-negative pathogens with RIK-10 + (i.e., LL-37mini2) being the strongest and LL-10 + the weakest. To understand this activity difference, we characterized peptide toxicity; the effects of salts, pH, and serum on peptide activity; and the mechanism of action and determined the membrane-bound helical structure for RIK-10 + by two-dimensional NMR spectroscopy. By writing an R program, we generated charge density plots for these peptides and uncovered the importance of the N-terminal high-density basic charges for antimicrobial potency. To validate this finding, we reversed the sequences of two peptides. Interestingly, sequence reversal weakened the activity of RIK-10 + but increased the activity of LL-10 + especially against Escherichia coli, Pseudomonas aeruginosa, and Acinetobacter baumannii. Those more active peptides with high cationicity at the N-terminus are also more hydrophobic based on HPLC retention times. A database search found numerous natural sequences that arrange basic amino acids primarily at the N-terminus. Combined, this study not only obtained novel peptide leads but also discovered one useful strategy for designing novel antimicrobials to control drug-resistant Gram-negative pathogens.

Quality assessment and characterization of unregulated antiviral drugs for feline infectious peritonitis: implications for treatment, safety, and efficacy.

Feline infectious peritonitis is fatal, and due to lack of approved treatments, unregulated antiviral drugs are used to treat this disease. This study set out to determine the purity of various batches of these drugs from several companies, characterize them, and note any impurities or other unusual characteristics. We also developed a method to qualitatively assess the primary components before administration. We tested 30 vials from 17 brands of GS-441524 and 5 vials from 1 brand of GC376. We compared the GS-441524 to a control standard from Ambeed and the GC376 to a standard from Cayman Chemical. We recorded physical appearance, pH, absorbance, HPLC retention times, and thin-layer chromatography retention factors for all of the samples. Some samples were used for nuclear magnetic resonance and mass spectrometric analysis. Some of the GS-441524 vials were 10% to 25% more concentrated than advertised, but most of the GS-441524 samples tested were similar in purity and composition, both between batches and between brands. We also tested 5 vials of GC376 and found that 1 of the 5 vials contained GS-441524 rather than GC376 and the other 4 vials contained molnupiravir. While all of the GS-441524 vials contained GS-441524, none of the GC376 vials tested contained GC376. GC376 is used in cats that are unresponsive to GS-441524, and use of the wrong antiviral can cause serious side effects. We provide suggested methods for distinguishing one drug from the other in new batches.

Identification and structural characterization of a novel (+)-catechin-caffeic acid adduct present in wines

A new compound with a molecular ion mass of m/z 467 in the negative ion mode was found to occur in a white wine aged 30 months in bottle. In this latter, fragment ions compatible with the loss of a carboxylic acid (-44 a.m.u.), a caffeic acid unit (-178 a.m.u.), and a Retro-Diels Alder (-152 a.m.u.) were observed. The present work reports the synthesis of a (+)-catechin-caffeic acid adduct resulting from the condensation reaction between caffeic acid and (+)-catechin. The structural characterization by NMR showed that this adduct is formed by the linkage between carbon 8 at ring A from (+)-catechin and carbon 9 from caffeic acid. In addition, the similarity in the HPLC retention time and UV–Visible spectra of the synthesized compound with the one detected in white wine and the bottling experiments, confirms the presence of this novel (+)-catechin-derived compound in those matrices.

Total Synthesis of Stachybotrys microspora Triprenol Phenol-7 (SMTP-7).

The first total synthesis of Stachybotrys microspora triprenol phenol (SMTP)-7 is described. Establishment of the two pyran ring stereogenic centers and key reactions featuring a double reductive amination and a double lactam ring formation in flow are described. The (2R,3S)-trans-benzopyran intermediate 7A, isolated by chiral preparative SFC chromatography, was carried forward to afford SMTP-7. Analytical data for synthetic SMTP-7, including 1H and 13C NMR data, HPLC retention time, and UV spectrum, were in excellent agreement with those for natural SMTP-7.

Identification of Genes Responsible for the Synthesis of Glycitein Isoflavones in Soybean Seeds.

Soybean (Glycine max (L.) Merrill) isoflavones are among the most important secondary metabolites, with functional benefits for human health. Soybeans accumulate three aglycone forms of isoflavones: genistein, daidzein, and glycitein. Soybean landrace Kumachi-1 does not accumulate malonylglycitin at all. Gene structure analysis indicated that Glyma.11G108300 (F6H4) of Kumachi-1 has a 3.8-kbp insertion, resulting in a truncated flavonoid 6-hydroxylase (F6H) sequence compared to the wild-type sequence in Fukuyutaka. Mapping experiments using a mutant line (MUT1246) with a phenotype similar to that of Kumachi-1, with a single-nucleotide polymorphism (SNP) in F6H4, revealed co-segregation of this mutation and the absence of glycitein isoflavones. We also identified a mutant line (K01) that exhibited a change in the HPLC retention time of glycitein isoflavones, accumulating glycoside and malonylglycoside forms of 6-hydroxydaidzein. K01 contains an SNP that produces a premature stop codon in Glyma.01G004200 (IOMT3), a novel soybean isoflavone O-methyltransferase (IOMT) gene. We further analyzed transgenic hairy roots of soybeans expressing Glyma.11G108300 (F6H4) and Glyma.01G004200 (IOMT3). Those overexpressing F6H4 accumulated malonylglycoside forms of 6-hydroxydaidzein (M_6HD), and co-expression of F6H4 and IOMT3 increased the level of malonylglycitin but not of M_6HD. These results indicate that F6H4 and IOMT3 are responsible for glycitein biosynthesis in soybean seed hypocotyl.

Colossal C130 Fullertubes: Soluble [5,5] C130-D5h(1) Pristine Molecules with 70 Nanotube Carbons and Two 30-Atom Hemifullerene End-caps.

We report the seminal experimental isolation and DFT characterization of pristine [5,5] C130-D5h(1) fullertubes. This achievement represents the largest soluble carbon molecule obtained in its pristine form. The [5,5] C130 species is the highest aspect ratio fullertube purified to date and now surpasses the recent gigantic [5,5] C120-D5d(1). In contrast to C90, C100, and C120 fullertubes, the longer C130-D5h has more nanotubular carbons (70) than end-cap fullerenyl atoms (60). Starting from 39,393 possible C130 isolated pentagon rule (IPR) structures and after analyzing polarizability, retention time, and UV-vis spectra, these three layers of data remarkably predict a single candidate isomer and fullertube, [5,5] C130-D5h(1). This structural assignment is augmented by atomic resolution STEM data showing distinctive and tubular "pill-like" structures with diameters and aspect ratios consistent with [5,5] C130-D5h(1) fullertubes. The high selectivity of the aminopropanol reaction with spheroidal fullerenes permits facile separation and removal of fullertubes from soot extracts. Experimental analyses (HPLC retention time, UV-vis, and STEM) were synergistically used (with polarizability and DFT property calculations) to down select and confirm the C130 fullertube structure. Achieving the isolation of a new [5,5] C130-D5h fullertube opens the door to application development and fundamental studies of electron confinement, fluorescence, and metallic character for a fullertube series of molecules with systematic tubular elongation. This [5,5] fullertube family also invites comparative studies with single-walled carbon nanotubes (SWCNTs), nanohorns (SWCNHs), and fullerenes.

Isolation, synthesis and identification of degraded impurities in Letermovir

Letermovir is a cytomegalovirus inhibitor for cytomegalovirus infection a hematopoietic-cell transplantation. In the degradation test of Letermovir, five new impurities were detected at levels of ND ∼ 2.21 % (by oxide, thermal or photolytic). These impurities were synthesized directly, characterized and identified by HRMS NMR spectra and X-ray crystallography. Then co-injected with commercial products to confirm their retention times in HPLC. The possible formation pathways and synthetic methods of these impurities were discussed in details. Furthermore, the toxicological properties of impurities were evaluated by ACD/Percepta 14.52.0 (Build 3525) software.

Isolation and stereospecific synthesis of indole alkaloids with lipid-lowering effects from the marine-derived fungus Colletotrichum gloeosporioides BB4

Seven undescribed compounds, colletotrichindoles A–E, colletotrichaniline A, and colletotrichdiol A, as well as three known compounds, (−)-isoalternatine A, (+)-alternatine A and 3-hydroxybutan-2-yl 2-phenylacetate were isolated from the marine-derived fungus Colletotrichu gloeosporioides BB4. The racemic mixtures colletotrichindole A，colletotrichindole C, and colletotrichdiol A were further separated by chiral chromatography to give three pairs of enantiomers (10S,11R,13S)/(10R,11S,13R)-colletotrichindole A, (10R,11R,13S)/(10S,11S,13R)-colletotrichindole C, and (9S,10S)/(9R,10R)-colletotrichdiol A, respectively. The chemical structures of seven undescribed compounds and the known compounds, (−)-isoalternatine A, and (+)-alternatine A were determined using a combination of NMR, MS, X-ray diffraction, ECD calculations, and/or chemical synthesis. All possible enantiomers of colletotrichindoles A–E were synthesized and used to determine the absolute configurations of the natural products by comparing their spectroscopic data and HPLC retention times on a chiral column. In addition, the X-ray crystal structures of the known compounds (−)-isoalternatine A and (+)-alternatine A were also obtained to confirm their absolute configurations. (10S,11R,13S)-Colletotrichindole A, colletotrichindole B, and (+)-alternatine A significantly reduced triglyceride levels in 3T3-L1 cells with EC50 values of 5.8, 9.0, and 1.3 μM, respectively.

Identification of Novel Parishin Compounds from the Twig of Maclura tricuspidata and Comparative Analysis of Parishin Derivatives in Different Parts

Parishin compounds are rare polyphenolic glucosides mainly found in the rhizome of the traditional Chinese medicinal plant, Gastrodia elata. These constituents are reported to have several biological and pharmacological activities. In the present study, two novel parishin derivatives not previously reported as plant-based phytochemicals were identified from a twig of Maclura tricuspidata (MT) and two new compounds were elucidated as 1-(4-(β-d-glucopyranosyloxy)benzyl)-3-hydroxy-3-methylpentane-1,5-dioate (named macluraparishin E) and 1,3-bis(4-(β-d-glucopyranosyloxy)benzyl)-3-hydroxy-3-methylpentane- 1,5-dioate (macluraparishin C), based on the experimental data obtained by UV-Visible (UV-Vis) spectroscopy, high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOF-MS) and nuclear magnetic resonance (NMR) spectroscopy. Additionally, gastrodin, parishin A and parishin B were positively identified by spectroscopic evidence and the comparison of HPLC retention time with the corresponding authentic standards. Gastrodin, parishin A and parishin B, macluraparishin E and macluraparishin C were found to be the most abundant constituents in the MT twig. The compositions and contents of these constituents were found to vary depending on the different parts of the MT plant. In particular, the contents of parishin A, parishin B, macluraparishin C and macluraparishin E were higher in the twig, bark and root than in the leaves, xylem and fruit.

Absolute Configuration of the Polyketide Natural Product (-)-Enterocin.

The absolute configuration of the polyketide natural product (-)-enterocin was established by two independent approaches. In the first approach, synthetic enterocin with a defined configuration was compared to the natural product. While identical in all scalar properties, the compound displayed an opposite specific rotation and a different chiral HPLC retention time when compared with (-)-enterocin. In a second approach, the vibrational circular dichroism (VCD) of the natural product was measured and shown to be opposite to the calculated VCD of its enantiomer.

Experimental design approach, screening and optimization of system variables, analytical method development of flurbiprofen in nanoparticles and stability-indicating methods for high-pressure liquid chromatography

BackgroundThe development of chromatographic method and the validation of a sensitive, simple, efficient, and reversed-phase high-performance liquid chromatography (RP-HPLC) approach were adopted for the drug flurbiprofen (FBP) in nanoparticles formulation by using a design of experiment (DoE). The critical method variables (CMVs) were screened using a statistical two-level fractional factorial design (FFD) followed by optimization of the selected CMVs that influence the analytical responses (ARs) of the RP-HPLC process by using two-level full factorial design.ResultsStatistical models are used to investigate the effects of system factors including column temperature, flow rate, and methanol in orthophosphoric acid (OPA) on the dependent responses, retention time, peak area, tailing factor, and theoretical plates in HPLC. The ideal column temperature (25 °C), flow rate (1 ml/min), and mobile phase (methanol 85 percent v/v in 0.05 percent OPA in water) were selected independently from the response surface at three levels (1, + 1, and 0) for further validation at constant solvent pH 2.75. Optimized method in the RP-HPLC resulted a retention time of 4.75 min, a peak area of 3975.12, a tailing factor of 0.73, and a total of 9697.7 theoretical plates followed by validation in accordance with the current ICH recommendations Q2 (R1). Linearity, precision, accuracy, assay, limit of detection (LOD), limit of quantification (LOQ), and robustness were all included in validation. The calibration curve was linear (r2 = 0.9997, slope = 70.72) for the concentration of 10 to 50 µg/ml, with a limit of detection of 0.14 µg/ml. Furthermore, stability-indicating methods demonstrate that drug degradation is highest in the presence of basic circumstances (about 96.49%), followed by oxidation (about 76.41%), and acidic conditions (about 48.12%), whereas drug is stable in some extent under neutral, photo (sunlight), and dry heat conditions.ConclusionsEffect of independent variables on dependent responses was screened and optimized by using statistical software design. A method for drug development could be successfully implemented for the estimation of drug in nanoparticles formulation as well as for the routine analysis in bulk and pharmaceutical formulations. The high recovery and low relative standard deviation support the suitability of proposed method that could be employed.Graphical

Gigantic C120 Fullertubes: Prediction and Experimental Evidence for Isomerically Purified Metallic [5,5] C120-D5d(1) and Nonmetallic [10,0] C120-D5h(10766).

We report the first experimental characterization of isomerically pure and pristine C120 fullertubes, [5,5] C120-D5d(1) and [10,0] C120-D5h(10766). These new molecules represent the highest aspect ratio fullertubes isolated to date; for example, the prior largest empty cage fullertube was [5,5] C100-D5d(1). This increase of 20 carbon atoms represents a gigantic leap in comparison to three decades of C60-C90 fullerene research. Moreover, the [10,0] C120-D5d(10766) fullertube has an end-cap derived from C80-Ih and is a new fullertube whose C40 end-cap has not yet been isolated experimentally. Theoretical and experimental analyses of anisotropic polarizability and UV-vis assign C120 isomer I as a [5,5] C120-D5d(1) fullertube. C120 isomer II matches a [10,0] C120-D5h(10766) fullertube. These structural assignments are further supported by Raman data showing metallic character for [5,5] C120-D5d(1) and nonmetallic character for C120-D5h(10766). STM imaging reveals a tubular structure with an aspect ratio consistent with a [5,5] C120-D5d(1) fullertube. With microgram quantities not amenable to crystallography, we demonstrate that DFT anisotropic polarizability, augmented by long-accepted experimental analyses (HPLC retention time, UV-vis, Raman, and STM) can be synergistically used (with DFT) to down select, predict, and assign C120 fullertube candidate structures. From 10 774 mathematically possible IPR C120 structures, this anisotropic polarizability paradigm is quite favorable to distinguish tubular structures from carbon soot. Identification of isomers I and II was surprisingly facile, i.e., two purified isomers for two possible structures of widely distinguishing features. These metallic and nonmetallic C120 fullertube isomers open the door to both fundamental research and application development.

Simultaneous quantification of 18 bioactive constituents in Ziziphus jujuba fruits by HPLC coupled with a chemometric method

The fruit of Ziziphus jujuba Mill., known as Hongzao (or Hong-Zao) in Chinese and cultivated in China for more than 4000 years, has shown to have hepatoprotective property. In previous study, we have isolated and identified 27 known compounds from Z. jujuba fruits, which demonstrated anti-tumor activity. In this study, a high-performance liquid chromatography-diode-array detection-mass spectrometry (HPLC-DAD-MS) method was successfully applied to the simultaneous characterization and quantitation of 18 constituents in 28 Z. jujuba samples, comprised of 12 cultivars from different regions in China, by comparing their HPLC retention times, MS spectra, UV spectra, and NMR data with those of reference compounds. The quantitative method was validated with excellent linearity (R2 > 0.999 1), preferable intra- and inter-day precisions (RSD < 2.78%), and good recoveries (94.96%–102.65%). The content variation of 18 compounds was analyzed by a chemometric method (hierarchical cluster analysis). In addition, these constituents showed protection against carbon tetrachloride (CCl4) intoxicated HepG2 cell lines by decreasing lactic dehydrogenase (LDH) levels. Results in this study illustrated that the content of all 18 compounds examined has significant difference and variation among cultivars and extracts. The proposed method can serve as a prerequisite for quality control of bioactive compounds in Z. jujuba products.

Lead optimisation efforts on a molecular prototype of the immunomodulatory parasitic protein ES-62

Abstract The immunomodulatory property of some parasitic helminths is well documented. The glycoprotein ES-62 from the nematode, acanthocheilonema viteae has been found to possess immunomodulatory properties. Two small molecule analogues (SMA’s) of ES-62 (S3 and S5) were found to mimic its immunomodulatory properties in vivo and were active in animal models of allergic, inflammatory and autoimmune diseases. In this work, new efforts were made to further optimise the activities of compound S3 by making small but directed structural changes. A variety of analogues based on the S3 prototype were simulated by making variations at one position and then screened in silico. The best compounds were selected based on predicted physiochemical properties and medicinal chemistry indices and synthesised. Structural elucidation was done via HNMR, LCMS, FTIR and HRESIMS. The predicted properties were evaluated by HPLC method. A total of 11 novel molecules were synthesised and characterised. Significant correlation was obtained between the predicted physicochemical properties and their HPLC retention times (RT) for eight of our novel compounds. This suggests that these compounds may behave in a physiological environment as closely as computationally predicted. This entails, lesser host toxicity while maintaining good or better activities compared to the earlier prototype. They hence provide a good opportunity for development of drugs for immune conditions such as asthma, inflammation and autoimmune diseases.

LDAmy, an α-amylase from Colorado potato beetle (Leptinotarsa decemlineata) with transglycosylation activity

Potential of α-amylase from the gut of Leptinotarsa decemlineata (LDAmy) to catalyse transfer reactions was investigated. LDAmy as a component of insect gut extract showed significant transfer activity on reducing-end- and both-end-protected maltoheptamer substrates. Transfer reaction was examined using purified enzyme, 2-chloro-4-nitrophenyl β-d-glucopyranoside as acceptor and starch and maltooligosaccharides as donors. In addition to suitability of various donors, effect of pH and acetonitrile (MeCN) concentration were also studied. The reactions were followed using separation of reaction products by a reversed phase HPLC method. LDAmy catalysed the hydrolysis and transglycosylation of the both-end-protected substrate 4,6-O-benzylidene-4-nitrophenyl β-maltoheptaoside in parallel reactions. Shorter and longer both-end-protected products with degree of polymerization 4–10 were formed. Identification of products was carried out based on HPLC retention time, UV and mass spectra. Ratio of transglycosylation to hydrolysis reached 0.5 in presence of 20% MeCN as organic solvent. Aromatic protecting group at the non-reducing end was favourable for transfer reaction. Lack of the mobile loop and presence of more nonpolar aromatic moieties near to the active site may be the reason for the enhanced transfer activity of LDAmy based on the comparison of the sequence and structure of mammalian and insect-derived α-amylases. Highlights Transferase activity of Colorado potato beetle derived α-amylase LDAmy is presented. Effect of pH and organic co-solvent on transfer reaction of LDAmy were studied. Shorter and longer products were formed from a both-end protected maltoheptamer. The unusual transfer ability was explained by sequence differences of α-amylases.

Isolation, synthesis, identification of new process-related impurities in evocalcet

Evocalcet is a calcimimetic for the treatment of secondary hyperparathyroidism. In the preparation of evocalcet, eleven process-related impurities were detected in the reaction solution of the last step at levels of 0.05%− 2.50% by a new HPLC method. Ten impurities were separated from the enriched mother liquor and synthesized directly. These impurities were characterized and identified by HRMS and NMR spectra and then coinjected with commercial products to confirm their retention times in HPLC. The possible formation pathways and synthetic methods of ten process-related impurities ware discussed in detail.

Study of the relationship between applied transmembrane pressure and antimicrobial activity of lysozyme

During the processing of biomolecules by ultrafiltration, the lysozyme enzyme undergoes conformational changes, which can affect its antibacterial activity. Operational conditions are considered to be one of the main parameters responsible for such changes, especially when using the same membrane and molecule. The present study demonstrates that, the same cut-off membrane (commercial data) can result in different properties of the protein after filtration, due to their different pore network. The filtration of lysozyme, regardless of the membrane, produces a decrease in the membrane hydraulic permeability (between 10 and 30%) and an increase in its selectivity in terms of observed rejection rate (30%). For the filtrated lysozyme, it appears that the HPLC retention time increases depending on the membrane used. The antibacterial activity of the filtrated samples is lower than the native protein and decreases with the increase of the applied pressure reaching 55–60% loss for 12 bar which has not been reported in the literature before. The observed results by SEC-HPLC and bacteriological tests, suggest that the conformation of the filtrated molecules are indeed modified. These results highlight the relationship between protein conformation or activity and the imposed shear stress.

Fixed Dose Single Tablet Formulation with Differential Release of Amlodipine Besylate and Simvastatin and Its Pharmacokinetic Profile: QbD and Risk Assessment Approach.

PurposeA differential release fixed dose matrix tablet of amlodipine besylate (AML-B) and simvastatin (SIM) was formulated to enhance patient compliance.Material and MethodIn the first phase, release controlling parameters of AML-B and SIM granules were identified and in the second phase a fixed dose AML-B and SIM tablet formulation was prepared and optimized for a differential release of the drugs using a quality by design (QbD) and risk assessment approach. A validated HPLC method was employed for simultaneous determination of AML-B and SIM for FDC formulation. A pharmacokinetics of the above drugs was studied in healthy dogs in the third phase.ResultsIn QbD-based optimized formulation, Eudragit® RSPO-dicalcium phosphate (DCP) blend controlled the release of AML-B over 8 h, though this diffusion-controlled release assumed first order kinetics. DCP and Eudragit® RS 100 also retarded release of SIM causing SIM release over 8 h after AML-B release from the optimized FDC tablet formulation. The HPLC retention times of AML-B and SIM were 2.10 and 15.52 min, respectively. Linearity for AML-B was 5.0–50 ng/mL and 0.01–2.0 µg/mL for SIM with percent recoveries of 92.85–101.53% and 94.51–117.75% for AML-B and SIM. AUC0-∞ of AML-B was increased 3 fold, while AUC0-∞ of SIM was decreased 2 fold. The tmax values for AML-B and SIM were 12 and 6 h, respectively. AML-B was absorbed without any lag time (tlag) while tlag was 6.33 ± 0.81 h for SIM, thus met the study objective.ConclusionThe pharmacokinetic study showed an immediate absorption of AML-B while that of SIM was withheld for 6 h, close to the desired delay time of 8 h.

Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS/MS data of reference desulfoglucosinolates

A library of ion trap MS2 spectra and HPLC retention times reported here allowed distinction in plants of at least 70 known glucosinolates (GSLs) and some additional proposed GSLs. We determined GSL profiles of selected members of the tribe Cardamineae (Brassicaceae) as well as Reseda (Resedaceae) used as outgroup in evolutionary studies. We included several accessions of each species and a range of organs, and paid attention to minor peaks and GSLs not detected. In this way, we obtained GSL profiles of Barbarea australis, Barbarea grayi, Planodes virginica selected for its apparent intermediacy between Barbarea and the remaining tribe and family, and Rorippa sylvestris and Nasturtium officinale, for which the presence of acyl derivatives of GSLs was previously untested. We also screened Armoracia rusticana, with a remarkably diverse GSL profile, the emerging model species Cardamine hirsuta, for which we discovered a GSL polymorphism, and Reseda luteola and Reseda odorata. The potential for aliphatic GSL biosynthesis in Barbarea vulgaris was of interest, and we subjected P-type and G-type B. vulgaris to several induction regimes in an attempt to induce aliphatic GSL. However, aliphatic GSLs were not detected in any of the B. vulgaris types. We characterized the investigated chemotypes phylogenetically, based on nuclear rDNA internal transcribed spacer (ITS) sequences, in order to understand their relation to the species B. vulgaris in general, and found them to be representative of the species as it occurs in Europe, as far as documented in available ITS-sequence repositories. In short, we provide GSL profiles of a wide variety of tribe Cardamineae plants and conclude aliphatic GSLs to be absent or below our limit of detection in two major evolutionary lines of B. vulgaris. Concerning analytical chemistry, we conclude that availability of authentic reference compounds or reference materials is critical for reliable GSL analysis and characterize two publicly available reference materials: seeds of P. virginica and N. officinale.

Neobulgarones Revisited: Anti and Syn Bianthrones from an Australian Mud Dauber Wasp Nest-Associated Fungus, Penicillium sp. CMB-MD22.

We report on the chemical analysis of a mud dauber wasp nest-associated fungus, Penicillium sp. CMB-MD22, leading to the discovery and structure elucidation of three known (1-3) and two new (4 and 5) anthrones, and a family of new and known bianthrones, neobulgarones 6-23. Detection and structure elucidation of 1-23 was supported by detailed spectroscopic analysis, as well as chemical (thermal) transformations, and global natural products social (GNPS) molecular networking. An empirical approach using HPLC retention times was effective at differentiating anti from syn bianthrone isomers, while a facile thermal equilibration was shown to favor anti over syn isomers. The neobulgarones 6-23 are natural products, and a crude extract rich in 6-23 exhibits selective antifungal activity against a co-isolated mud dauber wasp nest-associated fungus, suggestive of a possible ecological role as an antifungal chemical defense.