Large Scale Conversion of Trilobolide into the Payload of Mipsagargin: 8-O-(12-Aminododecanoyl)-8-O-Debutanoylthapsigargin.

In spite of the impressing cytotoxicity of thapsigargin (Tg), this compound cannot be used as a chemotherapeutic drug because of general toxicity, causing unacceptable side effects. Instead, a prodrug targeted towards tumors, mipsagargin, was brought into clinical trials. What substantially reduces the clinical potential is the limited access to Tg and its derivatives and cost-inefficient syntheses with unacceptably low yields. Laser trilobum, which contains a structurally related sesquiterpene lactone, trilobolide (Tb), is successfully cultivated. Here, we report scalable isolation of Tb from L. trilobum and a transformation of Tb to 8-O-(12-aminododecanoyl)-8-O-debutanoylthapsigargin in seven steps. The use of cultivated L. trilobum offers an unlimited source of the active principle in mipsagargin.

Maturation of the gut microbiome during the first year of life contributes to the protective farm effect on childhood asthma.

Growing up on a farm is associated with an asthma-protective effect, but the mechanisms underlying this effect are largely unknown. In the Protection against Allergy: Study in Rural Environments (PASTURE) birth cohort, we modeled maturation using 16S rRNA sequence data of the human gut microbiome in infants from 2 to 12 months of age. The estimated microbiome age (EMA) in 12-month-old infants was associated with previous farm exposure (ß = 0.27 (0.12-0.43), P = 0.001, n = 618) and reduced risk of asthma at school age (odds ratio (OR) = 0.72 (0.56-0.93), P = 0.011). EMA mediated the protective farm effect by 19%. In a nested case-control sample (n = 138), we found inverse associations of asthma with the measured level of fecal butyrate (OR = 0.28 (0.09-0.91), P = 0.034), bacterial taxa that predict butyrate production (OR = 0.38 (0.17-0.84), P = 0.017) and the relative abundance of the gene encoding butyryl-coenzyme A (CoA):acetate-CoA-transferase, a major enzyme in butyrate metabolism (OR = 0.43 (0.19-0.97), P = 0.042). The gut microbiome may contribute to asthma protection through metabolites, supporting the concept of a gut-lung axis in humans.

The characteristic smell emitted from two scale insects, Ceroplastes japonicus and Ceroplastes rubens.

The volatile components emitted from two scale insects, Ceroplastes japonicus and Ceroplastes rubens, were identified using GC-MS analysis. The major volatile components of the solvent extract from C. japonicus were α-humulene (35.8%) and δ-cadinene (17.0%), while those of C. rubens were ß-selinene (10.3%) and ß-elemene (5.1%). In GC/olfactometry, linalool, butyric acid, 3-methylbutyric acid, 2-methylbutyric acid, and vanillin were identified as the odor-active components of the extract from C. japonicus, in addition to trace amounts of trans-4,5-epoxy-(2E)-decenal, 4-methyl-(3E)-hexenoic acid, and phenylacetic acid. With regard to C. rubens, trans-4,5-epoxy-(2E)-decenal, 3-methylbutyric acid, and phenylacetic acid were identified as the odor-active components. Besides, decan-1,4-olide (γ-decalactone) with milky cherry-like note and 3-hydroxy-4,5-dimethylfuran-2(5H)-one (sotolone) with brown sugar-like note were also detected as the characteristic cherry-like sweet-and-sour note of these two scale insects. ABBREVIATIONS: GC: Gas chromatography; GC/O: gas chromatography/olfactometry.

Enantiomeric ratios of 2-methylbutanoic acid and its methyl ester: Elucidation of novel biogenetic pathways towards (R)-methyl 2-methylbutanoate in a beverage fermented with shiitake.

Up to 35% of (R)-methyl 2-methylbutanoate (M2MB) was observed in a beverage fermented with shiitake. As M2MB naturally occurs typically in high excesses of the (S)-enantiomer, the origin of the (R)-ester was elucidated by stable isotope labeled precursor-feeding studies. (R)-2-Methylbutanoic acid was identified as the main precursor in the substrate wort. Trace amounts of (R)-M2MB were produced by transformation of unsaturated secondary metabolites (tiglic aldehyde and tiglic acid) derived from l-isoleucine. Surprisingly, shiitake esterified (R)-2-methylbutanoic acid faster to (R)-M2MB than the corresponding (S)-enantiomer. Concurrently, spontaneous non-enantioselective degradation of M2MB occurred in shiitake. This explains diverse enantiomeric ratios of M2MB and different enantiomeric ratios of 2-methylbutanoic acid and M2MB in the beverage. As the odor threshold values of (R)-and (S)-M2MB differ significantly, these findings are of high relevance for the overall flavor of the fermented beverage and elucidate the discrepancy of enantiomeric ratios of 2-alkyl-branched acids and esters reported in nature.

Decursin and Z-Ligustilide in Angelica tenuissima Root Extract Fermented by Aspergillus oryzae Display Anti-Pigment Activity in Melanoma Cells.

The anti-melanogenic effects of the extract of Angelica tenuissima (AT) root and the extract of AT root fermented by Aspergillus oryzae (FAT) were investigated. These effects were determined by measuring the inhibitory activity of AT and FAT on melanin production in B16F10 melanocytes and with in vitro tyrosinase activity assays. The AT extract inhibited melanin production at concentrations above 250 µg/ml, and this inhibitory effect was significantly enhanced by the fermentation process with A. oryzae. HPLC analysis resulted in the isolation of two active compounds from both the AT and FAT extracts. Their chemical structures were identified as decursin and Z-ligustilide through comparison with previously reported NMR data. The decursin and Z-ligustilide contents were increased in the FAT extract and could be responsible for its enhanced inhibitory effects on melanin production and tyrosinase activity compared with that of the AT extract.

Two new lignans from the resin of Bursera microphylla A. gray and their cytotoxic activity.

Two new lignans, namely 7-O-podophyllotoxinyl butyrate (1) and dihydroclusin 9-acetate (2), were isolated from the dichloromethane fraction of a methanol extract of Bursera microphylla (Burseraceae), along with eight known lignans (3-10). Their structures were determined by means of comprehensive spectroscopic analysis. Lignans 2-6 were tested for their anti-proliferative activity on the cancer cell lines LS180, A549 and HeLa, and on a non-cancer cell line, ARPE-19. Only compounds 4 and 5 showed an interesting activity on HeLa cells.

Natural Korean Medicine Dang-Gui: Biosynthesis, Effective Extraction and Formulations of Major Active Pyranocoumarins, Their Molecular Action Mechanism in Cancer, and Other Biological Activities.

Angelica gigas Nakai (AGN) is a crucial oriental medicinal herb that grows especially in Korea and the Far-East countries. It contains chemically active compounds like pyranocoumarins, polyacetylenes and essential oils, which might be useful for treatment of several chronic diseases. It has been used for centuries as a traditional medicine in Southeast Asia, but in Western countries is used as a functional food and a major ingredient of several herbal products. The genus Angelica is also known as 'female ginseng' due to its critical therapeutic role in female afflictions, such as gynecological problems. However, it is well-documented that the AGN pyranocoumarins may play vital beneficial roles against cancer, neurodisorders, inflammation, osteoporosis, amnesia, allergies, depression, fungi, diabetes, ischemia, dermatitis, reactive oxygen species (ROS) and androgen. Though numerous studies revealed the role of AGN pyranocoumarins as therapeutic agents, none of the reviews have published their molecular mechanism of action. To the best of our knowledge, this would be the first review that aims to appraise the biosynthesis of AGN's major active pyranocoumarins, discuss effective extraction and formulation methods, and detail the molecular action mechanism of decursin (D), decursinol angelate (DA) and decursinol (DOH) in chronic diseases, which would further help extension of research in this area.

High metabolic versatility of different toxigenic and non-toxigenic Clostridioides difficile isolates.

Clostridioides difficile (formerly Clostridium difficile) is a major nosocomial pathogen with an increasing number of community-acquired infections causing symptoms from mild diarrhea to life-threatening colitis. The pathogenicity of C. difficile is considered to be mainly associated with the production of genome-encoded toxins A and B. In addition, some strains also encode and express the binary toxin CDT. However; a large number of non-toxigenic C. difficile strains have been isolated from the human gut and the environment. In this study, we characterized the growth behavior, motility and fermentation product formation of 17 different C. difficile isolates comprising five different major genomic clades and five different toxin inventories in relation to the C. difficile model strains 630Δerm and R20291. Within 33 determined fermentation products, we identified two yet undescribed products (5-methylhexanoate and 4-(methylthio)-butanoate) of C. difficile. Our data revealed major differences in the fermentation products obtained after growth in a medium containing casamino acids and glucose as carbon and energy source. While the metabolism of branched chain amino acids remained comparable in all isolates, the aromatic amino acid uptake and metabolism and the central carbon metabolism-associated fermentation pathways varied strongly between the isolates. The patterns obtained followed neither the classification of the clades nor the ribotyping patterns nor the toxin distribution. As the toxin formation is strongly connected to the metabolism, our data allow an improved differentiation of C. difficile strains. The observed metabolic flexibility provides the optimal basis for the adaption in the course of infection and to changing conditions in different environments including the human gut.

In situ esterification and extractive fermentation for butyl butyrate production with Clostridium tyrobutyricum.

Butyl butyrate (BB) is a valuable chemical that can be used as flavor, fragrance, extractant, and so on in various industries. Meanwhile, BB can also be used as a fuel source with excellent compatibility as gasoline, aviation kerosene, and diesel components. The conventional industrial production of BB is highly energy-consuming and generates various environmental pollutants. Recently, there have been tremendous interests in producing BB from renewable resources through biological routes. In this study, based on the fermentation using the hyper-butyrate producing strain Clostridium tyrobutyricum ATCC 25755, efficient BB production through in situ esterification was achieved by supplementation of lipase and butanol into the fermentation. Three commercially available lipases were assessed and the one from Candida sp. (recombinant, expressed in Aspergillus niger) was identified with highest catalytic activity for BB production. Various conditions that might affect BB production in the fermentation have been further evaluated, including the extractant type, enzyme loading, agitation, pH, and butanol supplementation strategy. Under the optimized conditions (5.0 g L-1 of enzyme loading, pH at 5.5, butanol kept at 10.0 g L-1 ), 34.7 g L-1 BB was obtained with complete consumption of 50 g L-1 glucose as the starting substrate. To our best knowledge, the BB production achieved in this study is the highest among the ever reported from the batch fermentation process. Our results demonstrated an excellent biological platform for renewable BB production from low-value carbon sources. Biotechnol. Bioeng. 2017;114: 1428-1437. © 2017 Wiley Periodicals, Inc.

Efficient biosynthesis of ethyl (R)-4-chloro-3-hydroxybutyrate using a stereoselective carbonyl reductase from Burkholderia gladioli.

BACKGROUND: Ethyl (R)-4-chloro-3-hydroxybutyrate ((R)-CHBE) is a versatile chiral precursor for many pharmaceuticals. Although several biosynthesis strategies have been documented to convert ethyl 4-chloro-3-oxobutanoate (COBE) to (R)-CHBE, the catalytic efficiency and stereoselectivity are still too low to be scaled up for industrial applications. Due to the increasing demand of (R)-CHBE, it is essential to explore more robust biocatalyst capable of preparing (R)-CHBE efficiently. RESULTS: A stereoselective carbonyl reductase toolbox was constructed and employed into the asymmetric reduction of COBE to (R)-CHBE. A robust enzyme designed as BgADH3 from Burkholderia gladioli CCTCC M 2012379 exhibited excellent activity and enantioselectivity, and was further characterized and investigated in the asymmetric synthesis of (R)-CHBE. An economical and satisfactory enzyme-coupled cofactor recycling system was created using recombinant Escherichia coli cells co-expressing BgADH3 and glucose dehydrogenase genes to regenerate NADPH in situ. In an aqueous/octanol biphasic system, as much as 1200 mmol COBE was completely converted by using substrate fed-batch strategy to afford (R)-CHBE with 99.9 % ee at a space-time yield per gram of biomass of 4.47 mmol∙L-1∙h-1∙g DCW-1. CONCLUSIONS: These data demonstrate the promising of BgADH3 in practical synthesis of (R)-CHBE as a valuable chiral synthon. This study allows for the further application of BgADH3 in the biosynthesis of chiral alcohols, and establishes a preparative scale process for producing (R)-CHBE with excellent enantiopurity.

Nanocomposites based on Soluplus and Angelica gigas Nakai extract fabricated by an electrohydrodynamic method for oral administration.

Nanocomposites (NCs) based on Soluplus (SP) were fabricated by an electrohydrodynamic (EHD) method for the oral delivery of Angelica gigas Nakai (AGN). Nano-sized particles were obtained after dispersing the resultant, produced by the EHD technique, in the aqueous environment. AGN/SP2 (AGN:SP=1:2, w/w) NC dispersion in aqueous media exhibited a 130nm mean diameter, narrow size distribution, and robust stability in the tested concentration range of the ethanol extract of AGN (AGN EtOH ext) and at pH 1.2 and 6.8. Amorphization of the components of AGN and their interactions with SP in the AGN/SP2 NC formulation were demonstrated by X-ray diffractometry (XRD) analysis. The released amounts of decursin (D) and decursinol angelate (DA), major components of AGN, from NCs were improved compared with those from the AGN EtOH ext group at both pH 1.2 and 6.8. As D and DA can be metabolized into decursinol (DOH) in the liver after oral administration, the DOH concentrations in plasma were quantitatively determined to evaluate the oral absorption of AGN. In a pharmacokinetic study in rats, higher oral absorption and the maximum concentration in plasma (Cmax) were presented in the AGN/SP2 NC group compared with the AGN EtOH ext and AGN NC groups. These findings indicate the successful application of developed SP-based NCs for the oral delivery of AGN.

Cardiorespiratory fitness as a predictor of intestinal microbial diversity and distinct metagenomic functions.

BACKGROUND: Reduced microbial diversity in human intestines has been implicated in various conditions such as diabetes, colorectal cancer, and inflammatory bowel disease. The role of physical fitness in the context of human intestinal microbiota is currently not known. We used high-throughput sequencing to analyze fecal microbiota of 39 healthy participants with similar age, BMI, and diets but with varying cardiorespiratory fitness levels. Fecal short-chain fatty acids were analyzed using gas chromatography. RESULTS: We showed that peak oxygen uptake (VO2peak), the gold standard measure of cardiorespiratory fitness, can account for more than 20 % of the variation in taxonomic richness, after accounting for all other factors, including diet. While VO2peak did not explain variation in beta diversity, it did play a significant role in explaining variation in the microbiomes' predicted metagenomic functions, aligning positively with genes related to bacterial chemotaxis, motility, and fatty acid biosynthesis. These predicted functions were supported by measured increases in production of fecal butyrate, a short-chain fatty acid associated with improved gut health, amongst physically fit participants. We also identified increased abundances of key butyrate-producing taxa (Clostridiales, Roseburia, Lachnospiraceae, and Erysipelotrichaceae) amongst these individuals, which likely contributed to the observed increases in butyrate levels. CONCLUSIONS: Results from this study show that cardiorespiratory fitness is correlated with increased microbial diversity in healthy humans and that the associated changes are anchored around a set of functional cores rather than specific taxa. The microbial profiles of fit individuals favor the production of butyrate. As increased microbiota diversity and butyrate production is associated with overall host health, our findings warrant the use of exercise prescription as an adjuvant therapy in combating dysbiosis-associated diseases.

Decursin from Angelica gigas suppresses RANKL-induced osteoclast formation and bone loss.

Osteoclasts are the only cells capable of breaking down bone matrix, and excessive activation of osteoclasts is responsible for bone-destructive diseases. In this study, we investigated the effects of decursin from extract of Angelica gigas root on receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclast formation using mouse bone marrow-derived macrophages (BMMs). Decursin inhibited RANKL-induced osteoclast formation without cytotoxicity. In particular, decursin maintains the characteristics of macrophages by blocking osteoclast differentiation by RANKL. Furthermore, the RANKL-stimulated bone resorption was diminished by decursin. Mechanistically, decursin blocked the RANKL-triggered ERK mitogen-activated protein kinases (MAPK) phosphorylation, which results in suppression of c-Fos and the nuclear factor of activated T cells (NFATc1) expression. In accordance with the in vitro study, decursin reduced lipopolysaccharide (LPS)- or ovariectomy (OVX)-induced bone loss in vivo. Therefore, decursin exerted an inhibitory effect on osteoclast formation and bone loss in vitro and in vivo. Decursin could be useful for the treatment of bone diseases associated with excessive bone resorption.

[Search of Neurotrophin-mimic Natural Products for Prevention and Treatment of Neurodegenerative Disease].

As part of our continuing studies on neurotrophin-mimic active compounds in natural products, we investigated the chemical constituents of the pericarps of Illicium jiadifengpi and the roots of Indonesian ginger Zingiber purpureum, resulting in the isolation of new seco-prezizaane-type sesquiterpenoid 1 and phenylbutenoid dimer 3-4 and two new curcuminoids 5-6. The MeOH extract of I. jiadifengpi was fractionated, leading to the isolation of compound 1. Compound 1 significantly enhanced neurite outgrowth in primary cell cultures of fetal rat cortical neurons. It is noteworthy that compound 1 has potential significantly to promote differentiation of multipotent neural stem cell line (MEB5 cells) into neurons. Additionally, we investigated the MeOH extract of the root of Bangle (Z. purpureum) that exhibited neuritogenesis activity in PC12 cells at 25 µg/mL, resulting in the isolation of neurotrophic phenylbutenoid dimers 3-4 and new compounds 5-6. Compounds 3 and 4 were found not only significantly to induce neurite sprouting of PC12 cells but also to increase the neurite length and number of neurites in primary cultured rat cortical neurons, and also showed protective activity against cell death caused by deprivation of serum. Furthermore, chronic treatment with these compounds enhanced hippocampal neurogenesis in dementia model olfactory bulbectomized (OBX) mice. Compounds 5 and 6 had significant NGF-potentiating effects on PC12 cells whereas compound 5 enhanced prevention of amyloid ß (Aß) 42 aggregation.

The Neuroprotective Effects of Decursin Isolated from Angelica gigas Nakai Against Amyloid ß-Protein-Induced Apoptosis in PC 12 Cells via a Mitochondria-Related Caspase Pathway.

Decursin, purified from Angelica gigas Nakai, has been proven to exert neuroprotective property. Previous study revealed decursin protected the PC12 cells from Aß25-35-induced oxidative cytotoxicity. The present study aimed to investigate whether decursin could protect PC12 cells from apoptosis caused by Aß. Our results indicated that pretreatment of PC12 cells with decursin significantly inhibited Aß25-35-induced cytotoxicity and apoptosis. The mechanism of action is likely to reverse Aß25-35-induced mitochondrial dysfunction, including the reduction of mitochondrial membrane potential, the inhibition of reactive oxygen species production, and the decrease of mitochondrial release of cytochrome c in PC12 cells. In addition, decursin significantly suppressed the activity of caspase-3 and moderated the ratio of Bcl-2/Bax induced by Aß25-35. These findings indicate that decursin exerts a neuroprotective effect against Aß25-35-induced neurotoxicity in PC12 cells, at least in part, via suppressing the mitochondrial pathway of cellular apoptosis.

Entre o campo e o laboratório: a dinâmica de produção de conhecimento no Ambulatório de Menopausa do Caism/Unicamp / Between the country and the laboratory: the dynamics of the production of knowledge at the Menopause Outpatients Clinic at Caism, Unicamp

Este estudo investiga as práticas de produção de conhecimento sobre a menopausa no Caism/Unicamp, centro de referência para políticas públicas em saúde da mulher. Foram realizadas observações de consultas ginecológicas, entrevistas com mulheres e médicos e observação de reuniões de apoio psicológico, buscando identificar os discursos que circulam no lugar e o processo de alistamento de diferentes atores para que os conhecimentos ali produzidos alcancem credibilidade e “viajem” além dos limites do hospital-escola, tornando-se “universais”. A análise baseia-se nos “estudos localistas”, alinhados aos estudos sociais de ciência e tecnologia.

This study investigates the practices involved in the production of knowledge about menopause at Caism, Unicamp, a reference center for public policies for women’s health. Gynecological appointments and psychological support meetings were observed, and women and doctors were interviewed in order to identify what discourse circulates there and how different actors are brought in to ensure that the knowledge produced attains credibility and “travels” beyond the boundaries of the teaching hospital to become “universal”. The analysis is based on localized studies aligned with social studies of science and technology.

A new dihydrofuranocoumarin from Opopanax hispidus (Friv.) Griseb.

Phytochemical investigation of chloroform extract of the aerial parts of Opopanax hispidus afforded a new dihydrofuranocoumarin, 3'-isobutyryl-3'-hydroxymarmesin (1a), together with six known coumarins, oreoselon, peucedanin, officinalin, smirniorin, 4'-acetyl-3'-isobutyryl-3'-hydroxymarmesin and 3'-hydroxyprantschimgin. The structures of the compounds were determined by their comprehensive spectroscopic analysis (IR, EI-MS, (1)H NMR, (13)C NMR, HMBC, COSY, HMBC and HMQC), elemental analysis and comparison with literature data.

Lipase inhibition and antiobesity effect of Atractylodes lancea.

The ethanol extract of Atractylodes lancea rhizome displayed significant lipase inhibition with an IC50 value of 9.06 µg/mL in a human pancreatic lipase assay from high-throughput screening. Bioassay-guided isolation led to the identification of one new polyacetylene, syn-(5E,11E)-3-acetoxy-4-O-(3-methylbutanoyl)-1,5,11-tridecatriene-7,9-diyne-3,4-diol (7), along with six known compounds (1-6). The structure of compound 7 was determined based on the analysis of NMR and MS data. Among these seven lipase inhibitors, the major compound atractylodin (1) showed the highest lipase inhibitory activity (IC50 = 39.12 µM). The antiobesity effect of the ethanol extract of Atractylodes lancea rhizome was evaluated in a high-fat diet-induced obesity mice model at daily dosages of 250 mg/kg and 500 mg/kg body weight for 4 weeks, and treatment with this extract demonstrated a moderate efficacy at the 500 mg/kg dose level.

In vitro metabolism of pyranocoumarin isomers decursin and decursinol angelate by liver microsomes from man and rodents.

The aim of this study is to investigate and compare the metabolic rate and profiles of pyranocoumarin isomers decursin and decursinol angelate using liver microsomes from humans and rodents, and to characterize the major metabolites of decursin and decursinol angelate in human liver microsomal incubations using LC-MS/MS. First, we conducted liver microsomal incubations of decursin and decursinol angelate in the presence or absence of NADPH. We found that in the absence of NADPH, decursin was efficiently hydrolyzed to decursinol by hepatic esterase(s), but decursinol angelate was not. In contrast, formation of decursinol from decursinol angelate was mediated mainly by cytochrome P450(s). Second, we measured the metabolic rate of decursin and decursinol angelate in liver S9 fractions from mice and humans. We found that human liver S9 fractions metabolized both decursin and decursinol angelate more slowly than those of the mouse. Third, we characterized the major metabolites of decursin and decursinol angelate from human liver microsomes incubations using HPLC-UV and LC-MS/MS methods and assessed the in vivo metabolites in mouse plasma from a one-dose PK study. Decursin and decursinol angelate have different metabolite profiles. Nine metabolites of decursin and nine metabolites of decursinol angelate were identified in human liver microsome incubations besides decursinol using a hybrid triple quadruple linear ion trap LC-MS/MS system, and many of them were later verified to be also present in plasma samples from rodent PK studies.