Use of 13C-NMR Chemical Shifts; Application of Principal Component Analysis for Categorizing Structurally Similar Methoxyflavones and Correlation Analysis between Chemical Shifts and Cytotoxicity.

The 13C-NMR spectral data for the 15-carbon flavonoid skeleton in eleven methoxyflavones isolated from Kaempferia parviflora (Zingiberaceae) were processed by principal component analysis (PCA). Based on the PCA score plots, the methoxyflavones were categorized into three groups according to their structural features. The cytotoxicities of the methoxyflavones toward 3T3-L1 murine preadipocyte cells were evaluated by 3-(4,5-dimethylthiazole-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTT) assay and found to differ according to structure. The relationship between the 13C-NMR chemical shifts of the methoxyflavones and their cytotoxicities was investigated using Pearson's correlation analysis. The 13C-NMR signal at C-10, a quaternary carbon, was correlated with cytotoxicity. Based on these results, a structural design which lowers the 13C-NMR chemical shift at C-10 would be important for the development of cytotoxic compounds. Although quantitative structure-activity and structure-property relationships are well established paradigms for predicting trends among a series of compounds, quantitative property-activity relationships have been relatively unstudied. This approach offers a new strategy for directing structure-activity relationship research.

Insolation-driven 100,000-year glacial cycles and hysteresis of ice-sheet volume.

The growth and reduction of Northern Hemisphere ice sheets over the past million years is dominated by an approximately 100,000-year periodicity and a sawtooth pattern (gradual growth and fast termination). Milankovitch theory proposes that summer insolation at high northern latitudes drives the glacial cycles, and statistical tests have demonstrated that the glacial cycles are indeed linked to eccentricity, obliquity and precession cycles. Yet insolation alone cannot explain the strong 100,000-year cycle, suggesting that internal climatic feedbacks may also be at work. Earlier conceptual models, for example, showed that glacial terminations are associated with the build-up of Northern Hemisphere 'excess ice', but the physical mechanisms underpinning the 100,000-year cycle remain unclear. Here we show, using comprehensive climate and ice-sheet models, that insolation and internal feedbacks between the climate, the ice sheets and the lithosphere-asthenosphere system explain the 100,000-year periodicity. The responses of equilibrium states of ice sheets to summer insolation show hysteresis, with the shape and position of the hysteresis loop playing a key part in determining the periodicities of glacial cycles. The hysteresis loop of the North American ice sheet is such that after inception of the ice sheet, its mass balance remains mostly positive through several precession cycles, whose amplitudes decrease towards an eccentricity minimum. The larger the ice sheet grows and extends towards lower latitudes, the smaller is the insolation required to make the mass balance negative. Therefore, once a large ice sheet is established, a moderate increase in insolation is sufficient to trigger a negative mass balance, leading to an almost complete retreat of the ice sheet within several thousand years. This fast retreat is governed mainly by rapid ablation due to the lowered surface elevation resulting from delayed isostatic rebound, which is the lithosphere-asthenosphere response. Carbon dioxide is involved, but is not determinative, in the evolution of the 100,000-year glacial cycles.

Synthesis and anti-influenza virus evaluation of triterpene-sialic acid conjugates.

We are interested in new non-natural glycosides with sialic acid conjugates and their biological activities. We report the synthesis of eleven non-natural occurring glycosides, which are triterpene (glycyrrhetinic acid and its derivatives)-sialic acid conjugates, and their inhibitory activities against influenza virus sialidases and influenza virus multiplication in MDCK host cells. Deoxoglycyrrhetol-sialic acid conjugates (6d and 6e) and oleanolic acid-sialic acid conjugates (7d and 7e) showed strong inhibitory activities against three subtypes of influenza virus sialidases. These four compounds (6d, 6e, 7d and 7e) showed clear inhibition to influenza virus multiplication but not to MDCK host cell survival.

Establishment of a Direct-Injection Electron Ionization-Mass Spectrometry Metabolomics Method and Its Application to Lichen Profiling.

Direct-injection electron ionization-mass spectrometry (DI-EI-MS) is a multivariate analysis method useful for characterizing biological materials. We demonstrated the use of DI-EI-MS for metabolic profiling using several closely related lichen species: Cladonia krempelhuberi, C. gracilis, C. pseudogymnopoda, and C. ramulosa. The methodology involves conversion of total ion chromatograms to integrated chromatograms and assessment of reproducibility. The qualitative DI-EI-MS method was used to profile the major and/or minor constituents in extracts of lichen samples. It was possible to distinguish each lichen sample by altering the electron energy in DI-EI-MS and examining the resulting data using one-way analysis of variance. Previously undetectable peaks, which are easy to fragment could be revealed by varying the electron energy. Our results suggest that metabolic profiling using DI-EI-MS would be useful for discriminating between subgroups within the same species. This is the first study to report the use of DI-EI-MS in a metabolomics application.

Inhibition of amyloid ß aggregation and protective effect on SH-SY5Y cells by triterpenoid saponins from the cactus Polaskia chichipe.

Alzheimer's disease (AD) destroys brain function, especially in the hippocampus, and is a social problem worldwide. A major pathogenesis of AD is related to the accumulation of amyloid beta (Aß) peptides, resulting in neuronal cell death in the brain. Here, we isolated four saponins (1-4) and elucidated their structures from 1D and 2D NMR and HRFABMS spectral data. The structures of 1 and 2 were determined as new saponins which have cochalic acid as the aglycon, and 3 was determined as a new saponin with oleanolic acid as the aglycon. Compound 4 was confirmed as the known saponin chikusetsusaponin V (=ginsenoside R0). Isolated saponins (1-4) and six previously reported saponins (5-10) were tested for their inhibitory effects of Aß aggregation and their protective effects on SH-SY5Y cells against Aß-associated toxicity. As the results, compounds 3 and 4 showed inhibitory effect of Aß aggregation and compounds 5-8 exerted the protective effects on SH-SY5Y cells against Aß-associated toxicity.

Inhibitory activities of biflavonoids against amyloid-ß peptide 42 cytotoxicity in PC-12 cells.

A major hallmark of Alzheimer's disease is the cerebral accumulation and resulting cytotoxicity of amyloid-ß peptides, particularly Aß42. In this study, we used an MTT assay to investigate the inhibitory activity of biflavonoids 1-22 against Aß42 cytotoxicity in PC-12 cell cultures. Cytoprotective effects were observed for the following amentoflavone type biflavonoids: podocarpusflavone B 8, isoginkgetin 10, sciadopitysin 13, and kayaflavone 15. These biflavonoids exhibited strong activity in tested compounds, with EC50 values of 5.18, 10.77, 9.84, and 5.29 µM, respectively. Cell viability tests of PC-12 cells revealed that biflavonoids 13 and 15 had stronger inhibitory activities than apigenin 23 and (-)-epigallocatechin gallate 24.

Biosynthesis of Panaefluoroline B from the Cultured Mycobiont of Amygdalaria panaeola.

Panaefluoroline B (2) is a fluorescent yellowish-green pigment produced by the cultured mycobiont of a lichen, Amygdalaria panaeola. Panaefluoroline B (2) has an isoquinoline skeleton, a C5 unit, and an amino acid, glycine, in its structure. The biosynthetic pathway of 2 was revealed by feeding experiments using [1-(13)C]-sodium acetate and [1,2-(13)C2][(15)N]-glycine. The analysis of labeling patterns of 2 and its mass spectrum suggested the isoquinoline part is biosynthesized via the acetate-malonate pathway with glycine as the nitrogen source and that the C5 unit originates from the mevalonate pathway.

Commissioning and quality assurance of Dynamic WaveArc irradiation.

A novel three-dimensional unicursal irradiation technique "Dynamic WaveArc" (DWA), which employs simultaneous and continuous gantry and O-ring rotation during dose delivery, has been implemented in Vero4DRT. The purposes of this study were to develop a commissioning and quality assurance procedure for DWA irradiation, and to assess the accuracy of the mechanical motion and dosimetric control of Vero4DRT. To determine the mechanical accuracy and the dose accuracy with DWA irradiation, 21 verification test patterns with various gantry and ring rotational directions and speeds were generated. These patterns were irradiated while recording the irradiation log data. The differences in gantry position, ring position, and accumulated MU (EG, ER, and EMU, respectively) between the planned and actual values in the log at each time point were evaluated. Furthermore, the doses delivered were measured using an ionization chamber and spherical phantom. The constancy of radiation output during DWA irradiation was examined by comparison with static beam irradiation. The mean absolute error (MAE) of EG and ER were within 0.1° and the maximum error was within 0.2°. The MAE of EMU was within 0.7 MU, and maximum error was 2.7 MU. Errors of accumulated MU were observed only around control points, changing gantry, and ring velocity. The gantry rotational range, in which EMU was greater than or equal to 2.0 MU, was not greater than 3.2%. It was confirmed that the extent of the large differences in accumulated MU was negligibly small during the entire irradiation range. The variation of relative output value for DWA irradiation was within 0.2%, and this was equivalent to conventional arc irradiation with a rotating gantry. In conclusion, a verification procedure for DWA irradiation was designed and implemented. The results demonstrated that Vero4DRT has adequate mechanical accuracy and beam output constancy during gantry and ring rotation.

Climate change and Southern Ocean ecosystems I: how changes in physical habitats directly affect marine biota.

Antarctic and Southern Ocean (ASO) marine ecosystems have been changing for at least the last 30 years, including in response to increasing ocean temperatures and changes in the extent and seasonality of sea ice; the magnitude and direction of these changes differ between regions around Antarctica that could see populations of the same species changing differently in different regions. This article reviews current and expected changes in ASO physical habitats in response to climate change. It then reviews how these changes may impact the autecology of marine biota of this polar region: microbes, zooplankton, salps, Antarctic krill, fish, cephalopods, marine mammals, seabirds, and benthos. The general prognosis for ASO marine habitats is for an overall warming and freshening, strengthening of westerly winds, with a potential pole-ward movement of those winds and the frontal systems, and an increase in ocean eddy activity. Many habitat parameters will have regionally specific changes, particularly relating to sea ice characteristics and seasonal dynamics. Lower trophic levels are expected to move south as the ocean conditions in which they are currently found move pole-ward. For Antarctic krill and finfish, the latitudinal breadth of their range will depend on their tolerance of warming oceans and changes to productivity. Ocean acidification is a concern not only for calcifying organisms but also for crustaceans such as Antarctic krill; it is also likely to be the most important change in benthic habitats over the coming century. For marine mammals and birds, the expected changes primarily relate to their flexibility in moving to alternative locations for food and the energetic cost of longer or more complex foraging trips for those that are bound to breeding colonies. Few species are sufficiently well studied to make comprehensive species-specific vulnerability assessments possible. Priorities for future work are discussed.

Antiangiogenic activity of hypoxylonol C.

Hypoxylonol C (1), isolated from the inedible mushroom Hypoxylon truncatum, exhibited inhibitory activities against the migration and tube formation of HUVECs. A cDNA microarray analysis was performed to investigate the target of hypoxylonol C (1) in HUVECs, and it was found that the genes related to cell cycle and adhesion were down-regulated. The down-regulation of mRNA levels of cell cycle and adhesion genes was confirmed by real-time RT-PCR. Cell cycle arrest and suppression of adhesion molecule expression might be plausible mechanisms of actions for the antiangiogenic activity of hypoxylonol C (1).

Pyridone alkaloids from a marine-derived fungus, Stagonosporopsis cucurbitacearum, and their activities against azole-resistant Candida albicans.

Four new 4-hydroxy-2-pyridone alkaloids, didymellamides A-D (1-4), were isolated from the marine-derived fungus Stagonosporopsis cucurbitacearum. The structures of 1-4 were elucidated from spectroscopic data (NMR, MS, and IR), and the absolute configuration of 1 was determined by X-ray diffraction analysis. Didymellamide A (1) exhibited antifungal activity against azole-resistant Candida albicans.

Effect of audio instruction on tracking errors using a four-dimensional image-guided radiotherapy system.

The Vero4DRT (MHI-TM2000) is capable of performing X-ray image-based tracking (X-ray Tracking) that directly tracks the target or fiducial markers under continuous kV X-ray imaging. Previously, we have shown that irregular respiratory patterns increased X-ray Tracking errors. Thus, we assumed that audio instruction, which generally improves the periodicity of respiration, should reduce tracking errors. The purpose of this study was to assess the effect of audio instruction on X-ray Tracking errors. Anterior-posterior abdominal skin-surface displacements obtained from ten lung cancer patients under free breathing and simple audio instruction were used as an alternative to tumor motion in the superior-inferior direction. First, a sequential predictive model based on the Levinson-Durbin algorithm was created to estimate the future three-dimensional (3D) target position under continuous kV X-ray imaging while moving a steel ball target of 9.5 mm in diameter. After creating the predictive model, the future 3D target position was sequentially calculated from the current and past 3D target positions based on the predictive model every 70 ms under continuous kV X-ray imaging. Simultaneously, the system controller of the Vero4DRT calculated the corresponding pan and tilt rotational angles of the gimbaled X-ray head, which then adjusted its orientation to the target. The calculated and current rotational angles of the gimbaled X-ray head were recorded every 5 ms. The target position measured by the laser displacement gauge was synchronously recorded every 10 msec. Total tracking system errors (ET) were compared between free breathing and audio instruction. Audio instruction significantly improved breathing regularity (p < 0.01). The mean ± standard deviation of the 95th percentile of ET (E95T ) was 1.7 ± 0.5 mm (range: 1.1-2.6mm) under free breathing (E95T,FB) and 1.9 ± 0.5 mm (range: 1.2-2.7 mm) under audio instruction (E95T,AI). E95T,AI was larger than E95T,FB for five patients; no significant difference was found between E95T,FB and E95T,AI (p = 0.21). Correlation analysis revealed that the rapid respiratory velocity significantly increased E95T. Although audio instruction improved breathing regularity, it also increased the respiratory velocity, which did not necessarily reduce tracking errors.

The FORCIS database: A global census of planktonic Foraminifera from ocean waters.

Planktonic Foraminifera are unique paleo-environmental indicators through their excellent fossil record in ocean sediments. Their distribution and diversity are affected by different environmental factors including anthropogenically forced ocean and climate change. Until now, historical changes in their distribution have not been fully assessed at the global scale. Here we present the FORCIS (Foraminifera Response to Climatic Stress) database on foraminiferal species diversity and distribution in the global ocean from 1910 until 2018 including published and unpublished data. The FORCIS database includes data collected using plankton tows, continuous plankton recorder, sediment traps and plankton pump, and contains ~22,000, ~157,000, ~9,000, ~400 subsamples, respectively (one single plankton aliquot collected within a depth range, time interval, size fraction range, at a single location) from each category. Our database provides a perspective of the distribution patterns of planktonic Foraminifera in the global ocean on large spatial (regional to basin scale, and at the vertical scale), and temporal (seasonal to interdecadal) scales over the past century.

Positional accuracy of novel x-ray-image-based dynamic tumor-tracking irradiation using a gimbaled MV x-ray head of a Vero4DRT (MHI-TM2000).

PURPOSE: To verify the positional accuracy of a novel x-ray-image-based dynamic tumor-tracking (DTT) irradiation technique using the gimbaled MV x-ray head of a Vero4DRT (MHI-TM2000). METHODS: Verification of the x-ray-image-based DTT was performed using three components: a three-dimensional moving phantom with a steel ball target, a laser displacement gauge, and an orthogonal kV x-ray imaging subsystem with a gimbaled MV x-ray head and the system controller of the Vero4DRT. The moving phantom was driven based on seven periodic patterns [peak-to-peak amplitude (A): 20-40 mm, breathing period (T): 2-5 s] and 15 patients' aperiodic respiratory patterns (A: 6.5-22.9 mm, T: 1.9-5.8 s). The target position was detected in real time with the orthogonal kV x-ray imaging subsystem using the stereo vision technique. Subsequently, the Vero4DRT predicted the next position of the target, and then the gimbaled MV x-ray head tracked the corresponding orientation of the target. The displacements of the target were measured synchronously using the laser displacement gauge. The difference between the target positions predicted by the Vero4DRT and those measured by the laser displacement gauge was computed as the prediction error (E(P)), and the difference between the target positions tracked by the gimbaled MV x-ray head and predicted target positions was computed as the mechanical error (E(M)). Total tracking system error (E(T)) was defined as the difference between the tracked and measured target positions. RESULTS: The root mean squares (RMSs) of E(P), E(M), and E(T) were up to 0.8, 0.3, and 0.7 mm, respectively, for the periodic patterns. Regarding the aperiodic patterns, the median RMSs of E(P), E(M), and E(T) were 1.2 (range, 0.9-1.8) mm, 0.1 (range, 0.1-0.5) mm, and 1.2 (range, 0.9-1.8) mm, respectively. From the results of principal component analysis, tracking efficiency, defined as the ratio of twice the RMS of E(T) to A, was improved for patients with high respiratory function (R = 0.91; p < 0.01). CONCLUSIONS: The present study demonstrated that the Vero4DRT is capable of high-accuracy x-ray-image-based DTT. E(T) was caused primarily by E(P), and E(M) was negligible. Furthermore, principal component analysis showed that tracking efficiency could be improved with this system, especially for patients with high respiratory function.

New triterpenoid saponins from cacti and anti-type I allergy activity of saponins from cactus.

The research in our laboratory focuses on the isolation of saponins from cactus. In this study, we report five new triterpenoid saponins, dumortierinoside A methyl ester (1), pachanoside I1 (2), pachanoside D1 (3), gummososide A (4), and gummososide A methyl ester (5). Compounds 1-3 isolated from Isolatocereus dumortieri Backbg., and compounds 4 and 5 were isolated from Stenocereus alamosensis A. C. Gibson & K. E. Horak. Compound 2 possessed a new pachanane-type triterpene skeleton, pachanol I, in its aglycon. The aglycon of 3 was pachanol D, while those of 4 and 5 were both gummosogenin, which we have previously reported, but this is the first report of pachanol D and gummosogenin in their aglycon forms. Additionally, we evaluated the anti-type I allergy activity of the saponins with RBL-2H3 (Rat basophilic leukemia) cells by measuring the ß-hexosaminidase release inhibitory activity. As a result of these studies, gummososide A methyl ester (5) was found to show activity (IC(50)=99.5 µM) and thurberoside A exhibited mild activity (IC(50)=166.9 µM).

Hypoxylonols C-F, benzo[j]fluoranthenes from Hypoxylon truncatum.

Four novel benzo[j]fluoranthene derivatives, hypoxylonols C (3), D (4), E (5), and F (6), have been isolated from the mushroom Hypoxylon truncatum, together with two known benzo[j]fluoranthene derivatives, hypoxylonols A (1) and B (2). The structures were established by analysis of NMR spectroscopic data and X-ray diffraction data. Compounds 4 and 5 showed antiproliferative activity against HUVECs (human umbilical vein endothelial cells) and HUAECs (human umbilical artery endothelial cells).

Polymethoxyflavonoids from Kaempferia parviflora induce adipogenesis on 3T3-L1 preadipocytes by regulating transcription factors at an early stage of differentiation.

We previously reported that Kaempferia parviflora WALL. ex BAKER (KP) and its ethyl acetate extract (KPE) improve various metabolic disorders in obesity-model mice. However the mechanism is not certain, and, in this study, in order to elucidate the mechanism of the suppressive effect of KP on fat accumulation, we focused on adipocytes, which are closely linked to metabolic diseases. The finding was that KPE and its components, 3,5,7,4'-tetramethoxyflavone and 3,5,7,3',4'-pentamethoxyflavone, strongly induced differentiation of 3T3-L1 preadipocytes to adipocytes. The above two polymethoxyflavonoids (PMFs) also induced adiponectin mRNA levels, and release of adiponectin into the medium. In addition, these PMFs enhanced the expression of peroxisome proliferator-activated receptor γ (PPARγ), but did not show PPARγ ligand activity. We then investigated the expression of the differentiation-regulator located upstream of PPARγ. Expression of CCAAT/enhancer-binding protein (C/EBP) ß and -δ mRNA, a transcriptional regulator of PPARγ, was induced, and expression of GATA-2 mRNA, a down-regulator of adipogenesis, was suppressed by these PMFs. These functions of the KP PMFs that enhance adipogenesis and secretion of adiponectin are, to some extent at least, involved in the mechanisms of anti-metabolic disorders effects.

Chemical constituents of a marine fungus, Arthrinium sacchari.

Three new diterpenes, myrocin D (1), libertellenone E (2), and libertellenone F (3), and a new isocoumarin, decarboxyhydroxycitrinone (4), were isolated from the marine fungus Arthrinium sacchari, together with three known compounds (5-7). The structures of 1-4 were elucidated from spectroscopic data (NMR, MS, IR), and the absolute configurations of 1-3 were determined by X-ray diffraction analysis. The antiangiogenic activity of these compounds was evaluated by measuring their antiproliferation effects on human umbilical vein endothelial cells (HUVECs) and human umbilical artery endothelial cells (HUAECs). Compounds 4-7 showed inhibitory activity.

Amyloid ß aggregation inhibitory activity of triterpene saponins from the cactus Stenocereus pruinosus.

Six new triterpene saponins (1-5,7) and 3 known saponins (6,8,9) were isolated from MeOH extracts of the cactus Stenocereus pruinosus. The structures of the isolated saponins were elucidated using MS, IR, and comprehensive NMR measurements. To develop drugs for treating Alzheimer's disease (AD) on the basis of the amyloid cascade hypothesis, the isolated saponins were evaluated for inhibition of BACE1 activity and amyloid beta (Aß) aggregation using thioflavin-T assay, and triterpenes as an aglycone moiety and an alkaline hydrolysate of the saponins were also evaluated. One saponin, stenoside A (7), exhibited inhibitory activity related to Aß aggregation and its degree of Aß aggregation was 40.6% at 100 µM.

beta-Secretase (BACE-1) inhibitory effect of biflavonoids.

Here, we describe amentoflavone-type biflavonoids, which were isolated from natural sources and were found to inhibit beta-secretase (BACE-1). The structure-activity relationship was studied, and compounds 1-8, 10, 17, and 18 showed BACE-1 inhibitory activity. Among these compounds, 2,3-dihydroamentoflavone 17 and 2,3-dihydro-6-methylginkgetin 18 exhibited potent inhibitory effects with IC(50) values of 0.75 and 0.35 microM, respectively.