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Rufomycin and ilamycin are synonymous for the same class of cyclopeptides, currently encompassing 33 structurally characterized isolates and 9 semisynthetic derivatives. Elucidation of new structures prioritized the consolidation of the names and established the structures of four diastereoisomeric rufomycins with a 2-piperidinone, named rufomycins 4–7, including full (1)H/(13)C NMR assignments. The characteristic HSQC cross-peak for the CH-5, the hemiaminal carbon in amino acid #5, allows assignment of the stereocenters C-4 and C-5 within this ring. Semisynthetic derivatives (rufomycinSS 1, 2, and 3) were prepared from a rufomycins 4 and 6 mixture to validate the structural assignments. Based on the X-ray crystal structures of rufomycins 2 and 4, considering the NMR differences of rufomycins 7 vs 4–6 compared to rufomycinSS 1 vs 2 and 3, and taking into account that two major conformers, A and B, occur in both rufomycinSS 2 and 3, structural modeling was pursued. Collectively, this paper discusses the NMR spectroscopic differences of the stereoisomers and their possible 3D conformers and correlates these with the anti-Mycobacterium tuberculosis activity. In addition, a look at the history prioritizes names and numbering schemes for this group of antibiotics and leads to consolidated nomenclature for all currently known members, natural and semisynthetic derivatives, and serves to accommodate future discoveries.

Favipiravir is an established antiviral that is currently being assessed as an investigational drug for the treatment of COVID-19. Favipiravir is strikingly similar to two molecules that the World Health Organization (WHO) lists as essential medicines, which also consist of a six-membered aromatic N-heterocycle bearing a carboxamide function: the anti-tuberculosis agent, pyrazinamide, and nicotinamide, also known as vitamin B3 . We demonstrate the utility of 1 H nuclear magnetic resonance (NMR) profiling, an emerging pharmacopoeial tool, for the highly specific identification, selective differentiation of congeners, and subsequent detection of drug falsification or adulteration of these medicines. The straightforward comparison of basic 1-D 1 H NMR spectra, obtained with benchtop or advanced NMR instruments alike, offers a rapid identity assay and works independently of physical reference materials. This approach accelerates and advances pharmaceutical quality control measures under situations of increased drug demand and altered economy, such as during a pandemic.

Following activation, macrophages undergo extensive metabolic rewiring1,2. Production of itaconate through the inducible enzyme IRG1 is a key hallmark of this process3. Itaconate inhibits succinate dehydrogenase4,5, has electrophilic properties6 and is associated with a change in cytokine production4. Here, we compare the metabolic, electrophilic and immunologic profiles of macrophages treated with unmodified itaconate and a panel of commonly used itaconate derivatives to examine its role. Using wild-type and Irg1-/- macrophages, we show that neither dimethyl itaconate, 4-octyl itaconate nor 4-monoethyl itaconate are converted to intracellular itaconate, while exogenous itaconic acid readily enters macrophages. We find that only dimethyl itaconate and 4-octyl itaconate induce a strong electrophilic stress response, in contrast to itaconate and 4-monoethyl itaconate. This correlates with their immunosuppressive phenotype: dimethyl itaconate and 4-octyl itaconate inhibited IκBζ and pro-interleukin (IL)-1β induction, as well as IL-6, IL-10 and interferon-β secretion, in an NRF2-independent manner. In contrast, itaconate treatment suppressed IL-1β secretion but not pro-IL-1β levels and, surprisingly, strongly enhanced lipopolysaccharide-induced interferon-β secretion. Consistently, Irg1-/- macrophages produced lower levels of interferon and reduced transcriptional activation of this pathway. Our work establishes itaconate as an immunoregulatory, rather than strictly immunosuppressive, metabolite and highlights the importance of using unmodified itaconate in future studies.

Thiocolchicoside is a natural product analogue often used for its spasmolytic action. To know more about its stability under various stress conditions, the drug was stirred in acid, base and peroxide solutions. In acid hydrolysis, two products were obtained and in both, the glucose got cleaved. In one of them the acetyl group also got cleaved. A set of two diastereomers were formed during the peroxide mediated hydrolysis. The base mediated hydrolysis resulted in formation of three novel degradants. They have six membered rings in their structures instead of a seven membered cycloheptatrienone. Structures of known and novel degradation products have been elucidated by extensive analysis of HRMS, 1D and 2D NMR spectroscopic techniques.

APOE is the strongest genetic risk factor for late-onset Alzheimer’s disease. ApoE exacerbates tau-associated neurodegeneration by driving microglial activation. However, how apoE regulates microglial activation and whether targeting apoE is therapeutically beneficial in tauopathy is unclear. Here we show that overexpressing an apoE metabolic receptor LDLR (low-density lipoprotein receptor) in P301S tauopathy mice markedly reduces brain apoE, and ameliorates tau pathology and neurodegeneration. LDLR overexpression (OX) in microglia cell-autonomously downregulates microglial Apoe expression, and is associated with suppressed microglial activation as in apoE-deficient microglia. Both apoE-deficiency and LDLR-OX strongly drive microglial immunometabolism towards enhanced catabolism over anabolism, whereas LDLR-overexpressing microglia also uniquely upregulate specific ion channels and neurotransmitter receptors upon activation. ApoE-deficient and LDLR-overexpressing mice harbor enlarged pools of oligodendrocyte progenitor cells (OPCs), and show greater preservation of myelin integrity under neurodegenerative conditions. They also show less reactive astrocyte activation in the setting of tauopathy.

Plasmacytoid dendritic cells (pDCs) specialize in the production of type I IFN (IFN-I). pDCs can be depleted in vivo by injecting diphtheria toxin (DT) in a mouse in which pDCs express a diphtheria toxin receptor (DTR) transgene driven by the human CLEC4C promoter. This promoter is enriched for binding sites for TCF4, a transcription factor that promotes pDC differentiation and expression of pDC markers, including CLEC4C. Here, we found that injection of DT in CLEC4C-DTR(+) mice markedly augmented Th2-dependent skin inflammation in a model of contact hypersensitivity (CHS) induced by the hapten fluorescein isothiocyanate. Unexpectedly, this biased Th2 response was independent of reduced IFN-I accompanying pDC depletion. In fact, DT treatment altered the representation of conventional dendritic cells (cDCs) in the skin-draining lymph nodes during the sensitization phase of CHS; there were fewer Th1-priming CD326(+) CD103(+) cDC1 and more Th2-priming CD11b(+) cDC2. Single-cell RNA-sequencing of CLEC4C-DTR(+) cDCs revealed that CD326(+) DCs, like pDCs, expressed DTR and were depleted together with pDCs by DT treatment. Since CD326(+) DCs did not express Tcf4, DTR expression might be driven by yet-undefined transcription factors activating the CLEC4C promoter. These results demonstrate that altered DC representation in the skin-draining lymph nodes during sensitization to allergens can cause Th2-driven CHS.

Seven piperic acid amides along with their lower homologs (12) were synthesized using HATU-DIPEA coupling reagent. All the synthesized derivatives were evaluated for their antibacterial activities against Staphylococcus aureus, Pseudomonas aeruginosa, and vancomycin-resistant P. aeruginosa. They were found to be more active on P. aeruginosa than on S. aureus. However, they did not exhibit potent activity on Vancomycin resistant P. aeruginosa. Among the tested compounds, methylenedioxycinnamic acid amide of anthranilic acid (MDCA-AA, 2a) was found to be most active against S. aureus with MIC of 3.125 μg/ml. The PAS and INH amides of piperic acid were screened against Mycobacterium tuberculosis H37Ra strain. They were found to be most active among all the tested compounds but were found to be less active than the standard drug, isoniazid.

Summary The apolipoprotein E (APOE) gene is the strongest genetic risk factor for Alzheimer’s disease and directly influences tauopathy and tau-mediated neurodegeneration. ApoE4 has strong deleterious effects on both parameters. In the brain, apoE is produced and secreted primarily by astrocytes and by activated microglia. The cell-specific role of each form of apoE in the setting of neurodegeneration has not been determined. We generated P301S Tau/Aldh1l1-CreERT2/apoE3flox/flox or Tau/Aldh1l1-CreERT2/apoE4flox/flox mice. At 5.5 months of age, after the onset of tau pathology, we administered tamoxifen or vehicle and compared mice at 9.5 months of age. Removing astrocytic APOE4 markedly reduced tau-mediated neurodegeneration and decreased phosphorylated tau (pTau) pathology. Single-nucleus RNA sequencing analysis revealed striking gene expression changes in all cell types, with astrocytic APOE4 removal decreasing disease-associated gene signatures in neurons, oligodendrocytes, astrocytes, and microglia. Removal of astrocytic APOE4 decreased tau-induced synaptic loss and microglial phagocytosis of synaptic elements, suggesting a key role for astrocytic apoE in synaptic degeneration.

Issues related to pharmaceutical quality are arising at an alarming rate. Pharmaceutical quality concerns both the Active Pharmaceutical Ingredients (APIs) and the Finished Drug Product/ Formulation. Recently, there has been a significant increase in the number of reports of harmful impurities in marketed drug formulations. Impurities range from solvents, reactants, adulterants, and catalysts to synthetic byproducts. Quality concerns in commercial preparations may also arise due to shelf life stability. Furthermore, a number of falsified and substandard drug cases have been reported. Most of the techniques which are currently in place can, at best, detect the impurities, but cannot identify them unless they are already known and can be compared to a standard. On the other hand, (1)H NMR spectroscopy detects all the hydrogen containing species, typically provides information to elucidate structures partially or even completely, and through its absolute quantitative capabilities even can detect the presence hydrogen-free species indirectly. The structural properties that produce (1)H NMR signals as characteristic representations of a given molecule are the chemical shifts (δ in ppm) and coupling constants (J in Hz). Along with the line widths (ω(1/2) in Hz), these parameters are bound to both the molecule and the NMR experimental conditions by the quantum mechanical (QM) principles. This means that the (1)H NMR spectra of APIs can be precisely calculated and compared to the experimental data. This review explains how (1)H NMR spectroscopy coupled with Full Spin Analysis can contribute towards the quality control of pharmaceuticals by improving structural dereplication and achieving simultaneous quantification of both APIs and their contaminants.

Chikungunya virus (CHIKV), an emerging alphavirus, has infected millions of people. However, the factors modulating disease outcome remain poorly understood. Here, we show in germ-free mice or in oral antibiotic-treated conventionally housed mice with depleted intestinal microbiomes that greater CHIKV infection and spread occurs within 1 day of virus inoculation. Alteration of the microbiome alters TLR7-MyD88 signaling in plasmacytoid dendritic cells (pDCs) and blunts systemic production of type I interferon (IFN). Consequently, circulating monocytes express fewer IFN-stimulated genes and become permissive for CHIKV infection. Reconstitution with a single bacterial species, Clostridium scindens, or its derived metabolite, the secondary bile acid deoxycholic acid, can restore pDC- and MyD88-dependent type I IFN responses to restrict systemic CHIKV infection and transmission back to vector mosquitoes. Thus, symbiotic intestinal bacteria modulate antiviral immunity and levels of circulating alphaviruses within hours of infection through a bile acid-pDC-IFN signaling axis, which affects viremia, dissemination, and potentially transmission.

Domperidone, a gastroprokinetic agent, is a common drug to treat emesis. It was subjected to acid, base-mediated hydrolysis, peroxide-mediated oxidation, photolysis and thermal degradation according to ICH guidelines to observe stability of the selected drug under the stress conditions. Although the drug is resistant to base hydrolysis, photolysis and thermal stressors, two degradants (DP-ISO1 and DP-ISO2) were formed in acid mediated hydrolysis. Oxidation with hydrogen peroxide also resulted in one product (DP-OX). All three degradants were isolated from the crude reaction mixture by preparative high-performance liquid chromatography and supercritical fluid chromatography. Structures of isolated compounds were unambiguously characterized as 5-chloro-1-(1-(3-(6-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-1-yl)propyl)piperidin-4-yl)-1,3-dihydro-2H-benzo[d]imidazol-2-one (DP-ISO1), 5-chloro-1-(3-(4-(5-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-1-yl)piperidin-1-yl)propyl)-1,3-dihydro-2H-benzo[d]imidazol-2-one (DP-ISO2), 4-(5-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazol-1-yl)-1-(3-(2-oxo-2,3-dihydro-1H-benzo[d]imidazol-1-yl)propyl)piperidine 1-oxide (DP-OX) by analysis of mass spectrometry, 1D and 2D nuclear magnetic resonance spectra. To the best of our knowledge, DP-ISO1 and DP-ISO2 are new and DP-OX was previously reported as domperidone impurity.

Keywords: Stereochemistry; 2,2,5-trisubsituted tetrahydrofuran; Malabaricanes; Ocotillone type dammaranes; 1H NMR splitting

Levodopa is effective for the motor symptoms of Parkinson's disease (PD), but is associated with motor fluctuations and dyskinesia. Many patients require add-on therapy to improve motor fluctuations without exacerbating dyskinesia. The objective of this Phase III, multicenter, double-blind, placebo-controlled, parallel-group study was to evaluate the efficacy and safety of safinamide, an α-aminoamide with dopaminergic and nondopaminergic mechanisms, as add-on to l-dopa in the treatment of patients with PD and motor fluctuations. Patients were randomized to oral safinamide 100 mg/day (n = 224), 50 mg/day (n = 223), or placebo (n = 222) for 24 weeks. The primary endpoint was total on time with no or nontroublesome dyskinesia (assessed using the Hauser patient diaries). Secondary endpoints included off time, Unified Parkinson's Disease Rating Scale (UPDRS) Part III (motor) scores, and Clinical Global Impression-Change (CGI-C). At week 24, mean ± SD increases in total on time with no or nontroublesome dyskinesia were 1.36 ± 2.625 hours for safinamide 100 mg/day, 1.37 ± 2.745 hours for safinamide 50 mg/day, and 0.97 ± 2.375 hours for placebo. Least squares means differences in both safinamide groups were significantly higher versus placebo. Improvements in off time, UPDRS Part III, and CGI-C were significantly greater in both safinamide groups versus placebo. There were no significant between-group differences for incidences of treatment-emergent adverse events (TEAEs) or TEAEs leading to discontinuation. The addition of safinamide 50 mg/day or 100 mg/day to l-dopa in patients with PD and motor fluctuations significantly increased total on time with no or nontroublesome dyskinesia, decreased off time, and improved parkinsonism, indicating that safinamide improves motor symptoms and parkinsonism without worsening dyskinesia.

In a 6-month double-blind, placebo-controlled study of Parkinson's disease patients with motor fluctuations, safinamide 50 and 100 mg/d significantly increased ON-time without increasing dyskinesia. Further long-term safinamide use in these patients was evaluated over an additional 18 months. Patients continued on their randomized placebo, 50, or 100 mg/d safinamide. The primary endpoint was change in Dyskinesia Rating Scale total score during ON-time over 24 months. Other efficacy endpoints included change in ON-time without troublesome dyskinesia, changes in individual diary categories, depressive symptoms, and quality of life measures. Change in Dyskinesia Rating Scale was not significantly different in safinamide versus placebo groups, despite decreased mean total Dyskinesia Rating Scale with safinamide compared with an almost unchanged score in placebo. Ad hoc subgroup analysis of moderate to severe dyskinetic patients at baseline (36% of patients) showed a decrease with safinamide 100 mg/d compared with placebo (P = 0.0317). Improvements in motor function, activities of daily living, depressive symptoms, clinical status, and quality of life at 6 months remained significant at 24 months. Adverse events and discontinuation rates were similar with safinamide and placebo. This 2-year, controlled study of add-on safinamide in mid-to-late Parkinson's disease with motor fluctuations, although not demonstrating an overall difference in dyskinesias between patients and controls, showed improvement in dyskinesia in patients at least moderately dyskinetic at baseline. The study additionally demonstrated significant clinical benefits in ON-time (without troublesome dyskinesia), OFF-time, activities of daily living, motor symptoms, quality of life, and symptoms of depression.

In the first phytochemical investigation of Trichuriella monsoniae, three known flavonoidal C-glycosides, isoswertisin 1, 2″-O-β-d-galactosyl isoswertisin 2 and 2″-O-β-d-xylosyl isoswertisin 3 were isolated from the methanolic extract of the whole plant. Their structures were elucidated by extensive NMR spectroscopic studies including 2D NMR and HRMS, and the structure of 2 was supported by single crystal X-ray data studies. Further, NMR assignments for 3 are being reported for the first time.

Summary An instrument to measure the seed coat adhesion, cotyledon adhesion and shear strength of pigeonpea grain was constructed using a peizo-electric transducer. Urea was applied to scarified and non-scarified grain and part was then heat treated. Seed coat adhesion was slightly reduced by scarification of grain and further reduced by urea plus heat treatment. Cotyledon adhesion was slightly reduced by urea plus heat but shear strength was unaffected. Pretreatments may lead to an improved quality product (good quality splits and less breakage). The instrument may be useful in assessing improved cultivars.

Ten malabaricane type triterpenes were isolated from the oleoresin of Ailanthus malabarica, out of which six (1-6) were new. For three of the known compounds (7-9), NMR assignments are being reported for the first time. Compound 10, a known one, is a new report from this source. The structures were established by extensive 1D and 2D NMR spectroscopy. The oleoresin and some of the isolates did not possess antimicrobial activity and did not lyse RBCs.

Summary Pigeonpea grain (c.v. C11) was used for optimization of dehulling pretreatments. Treatments applied for dehulling were soaking in urea solution and hot milling. Second order polynomial equations were developed to describe a dehulling index and dehulling time. Response surface methodology was also used. Treatment with urea solution was applied to both nonscarified and scarified grain. Different concentrations, solution-to-grain ratios and duration of treatments were tested in five levels as independent factors in the first set of experiments. In the second set, hot milling, heating time and heating temperature were factors in five levels. Hot milling was followed by optimized treatment with urea solution. The levels of these factors were optimized to give a maximum dehulling index and minimum dehulling time, this would yield the maximum product recovery and reduce grain scouring. The results revealed that hot milling after urea treatment of scarified grain increased the dehulling index by about 11% and, consequently, reduced dehulling time by approximately 65% when compared with the treatment followed in commercial mills. This subsequently increased the product recovery and intact pearled grain by about 19 and 23%, respectively.

Summary The effect of different pre-treatments, i.e. grain scarification, chemical treatment and hot milling, on the strength and dehulling properties of pigeonpea grain was studied by measurement of the forces required for removal of the seed coat from cotyledons and of cotyledons from each other. The dehulling index, dehulling time, product recovery and amount of intact pearled grain were determined after dehulling the pre-treated grain. The results revealed that a significant (P

The radioactivity administered as a single oral dose of N-(3-chloro-4-({beta}-D-glycosyl({sup 14}C)methyl)phenyl)urea was rapidly absorbed from the gastrointestinal tract and eliminated in feces and urine by both the rat and the Japanese quail. Within 6 h after treatment 57% and 74% of the dose was excreted by rat and quail, respectively. Of the 78% of the dose excreted by rat within 12 h, 49% was the parent glucoside, 3.6% the aglycon, and 14.6% the aglycon acid (the carboxyphenyl derivative of the aglycon). However, within the same period quail excreted 90% of the dose, of which 25.5% was the parent glucoside, 26.6% the aglycon, and 12% the aglycon acid. Unidentified metabolites, two in rat and seven in quail, formed 10.8% and 25.5% of the dose, respectively. The aglycon in rat and both the aglycon and the aglycon acid in quail formed conjugates.

Immobilisation sur le copolymere methacrylate d'epoxy-2,3 propyle-acrylate de butyle reticule avec le triethylene glycol dimethacrylate modifie ou non par l'hexamethylenediamine ou la L-lysine

Penicillin acylase of E. coli NCIM 2400 has been purified to homogeneity using a combination of hydrophobic interaction chromatography and DEAE-cellulose treatment. A variety of substituted matrices were synthesized using D- or DL-phenylglycine, norleucine, ampicillin, or amoxycillin as ligands, all of which retained penicillin acylase at high concentrations of ammonium sulfate or sodium sulfate. The enzyme could be eluted nonbiospecifically by buffer of lower ionic strength with over 95% recovery of the activity. Ammonium chloride, ammonium nitrate, sodium chloride, sodium nitrate, and potassium chloride were ineffective in either adsorption or elution of the enzyme on these columns. Further purification of this partially pure enzyme with DEAE-cellulose at pH 7.0-7.2 yielded an enzyme preparation of very high purity according to electrophoretic and ultracentrifugal analyses, its specific activity being as high as 37 U/mg protein. The purified enzyme has a molecular weight of 67,000 a sedimentation coefficient of 4.0S, and resolves into two forms upon isoelectric focusing. Overall recoveries ranged between 75 and 85%. Ease of operation, high recoveries, high purity of the enzyme and prolonged reuse of the conjugates make the process economically feasible and possibly of great commercial importance.

A higher yielding mutant of Aspergillus candidus Link var. aureus has been isolated using the chemical mutagen N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) at 5 μg/ml concentration of the mutagen. This mutant produces 88 to 111 U/ml of glucoamylase compared with 40 to 50 U/ml produced by the parent strain. The enzyme from this mutant has been purified to homogeneity using hydrophobic interaction chromatography and a DEAE-cellulose column with over 65% recovery of the enzyme activity and 21-fold purification. The mutant protein is very similar to the enzyme from the parent strain in that it cochromatographs with the parent enzyme on phenylglycyl-Sepharose, norleucyl-Sepharose, Sepharose-CL-6B, Concanavalin A-Sepharose, DEAE-cellulose and DEAE-Sephadex-A-50. The two proteins exhibit similar electrophoretic behavior and have similar molecular weights and amino acid composition. However, the specific activity of the purified mutant protein is 1125 U/mg compared with 560 U/mg for the parent enzyme. Further genetic analysis of the mutant is needed to explain these observations.

Amyloglucosidase (glucoamylase; EC 3.2.1.3) has been purified from the culture filtrates of Aspergillus candidus Link var. aureus using hydrophobic interaction chromatography and DEAE-cellulose treatment. The enzyme thus obtained has a specific activity of 329 units/mg protein with the overall recoveries between 70 and 75%. The process appears to be of industrial promise.