Effects of Roasting on the Quality of Moringa oleifera Leaf Powder and Loaf Volume of Moringa oleifera-Supplemented Bread.

Although a decrease in bread volume on adding nutrient-rich Moringa oleifera leaf powder (MLP) is known, to our knowledge, improving the swelling of MLP-added bread has not been attempted. This study aimed to investigate the effects of MLP and roasted MLP (RMLP) on bread quality. Bread was supplemented with MLP and RMLP treated at varying temperatures and times; the baked bread was then biochemically evaluated relative to the control. The specific volume of MLP-supplemented bread was 2.4 cm3/g, which increased to >4.0 cm3/g on using MLP roasted at 130 °C for ≥20 min, demonstrating remarkable swelling. The specific volume of bread supplemented with MLP roasted at 170 °C for 20 min was 4.6 cm3/g, similar to that of the control. Additionally, MLP interfered with carbon dioxide production in bread, thus decreasing the abundance of yeast cells; however, RMLP had no such effect and allowed normal fermentation. Scanning electron microscopy revealed gluten formation independent of MLP roasting. Thus, MLP-containing breads generally exhibit suppressed fermentation and expansion due to the bactericidal properties of raw MLP, but these effects are alleviated by heat treatment. These findings highlight the importance of heat treatment in mitigating the effects of MLP on bread fermentation and swelling.

Composition of Algerian Propolis, Plant Origin, and Its Antiangiogenic Activity In Vitro.

The antiangiogenic activity of the ethanol extract of propolis collected from different regions in western Algeria was investigated using in vitro human umbilical vein endothelial cells (HUVECs). The ethanol extract with the strongest activity, i.e., Algerian propolis 1 (EEPA1), inhibited the formation of capillary networks in a dose-dependent manner (6.25-50 µg/mL) within 12 h and induced cell fragmentation of HUVECs at 50 µg/mL after treatment for 24 h. To identify the active compounds in EEAP1, a high-performance liquid chromatography (HPLC) analysis was performed, revealing that EEAP1 contains two major compounds. Both compounds were isolated by repeated column chromatography and identified as ω-hydroxyferulenol (1) and ferulenol (2), which have a coumarin structure conjugated with a farnesyl group according to NMR, high-resolution electrospray ionization mass spectroscopy, and chemical modification. Compounds 1 and 2 inhibited the tube-forming activity of HUVECs, especially 2, which exhibited a stronger antiangiogenic effect even at a low concentration of 3.31 µg/mL. Moreover, 2 suppressed the elongation and induced cell fragmentation at the same dose. The molecular changes in tube-forming HUVECs induced by 2 were found to be related to the activation of the caspase signals. To confirm the plant origin of propolis, an HPLC comparative analysis of the ethanol extracts of some plants near beekeeping areas and that of Algerian propolis (EEAP1) was performed, and similar chromatographic patterns were observed. This result suggests that the plant origin of this Algerian propolis is the resin of Ferula communis.

Phytochemical and anti-inflammatory properties of Senegalese propolis and isolated compounds.

Propolis is a chemically complex resinous product collected from various plant sources by honeybees that has been used historically a traditional folk medicine in many parts of the world. The main constituents of propolis are beeswax and plant resins. We recently obtained Senegalese propolis, which, to our knowledge, has not been previously reported. The purpose of this study was to analyze the composition of Senegalese propolis and evaluate its anti-inflammatory activity. Ten known phenolic compounds with phenanthrene or stilbene skeletons were isolated. Nitric oxide (NO) production assay revealed that Senegalese propolis suppresses lipopolysaccharide (LPS)-stimulated production of NO in J774.1 cells in a dose-dependent manner. The anti-inflammatory potency of Senegalese propolis was higher than that of other previously reported propolis. Furthermore, the eight compounds isolated from Senegalese propolis showed high anti-inflammatory activity by inhibiting the LPS-induced expression of inducible NO synthase (iNOS). These results suggest that Senegalese propolis and its components have potential applications as anti-inflammatory agents.

Binding of Catechins to Staphylococcal Enterotoxin A.

Staphylococcal enterotoxin A (SEA) is a toxin protein, and is the most common cause of staphylococcal food poisoning. Polyphenols, such as catechins, are known to interact with proteins. In this study, we investigated the binding of catechins to SEA using SPR (Biacore), Fourier transform infrared spectroscopy (FT-IR), isothermal titration calorimetry (ITC), and protein-ligand docking. We found that (−)-epigallocatechin gallate (EGCG) could strongly bind to SEA. According to thermodynamic parameters, a negative ΔG indicated that the interaction between EGCG and SEA was spontaneous, and the electrostatic force accompanied by hydrophobic binding forces may play a major role in the binding. Data from Western blot analysis and docking simulation suggest that the hydroxyl group at position 3 of the galloyl group in the catechin structure was responsible for binding affinity with the Y91 of the A-6 region of SEA active sites. Our results provide further understanding of the binding interactions between catechins and SEA, and the inhibition of toxin activities by catechins.

5-Hydroxyferulic acid methyl ester isolated from wasabi leaves inhibits 3T3-L1 adipocyte differentiation.

To investigate the compounds present in wasabi leaves (Wasabia japonica Matsumura) that inhibit the adipocyte differentiation, activity-guided fractionation was performed on these leaves. 5-Hydroxyferulic acid methyl ester (1: 5-HFA ester), one of the phenylpropanoids, was isolated from wasabi leaves as a compound that inhibits the adipocyte differentiation. Compound 1 suppressed the intracellular lipid accumulation of 3T3-L1 cells without significant cytotoxicity. Gene expression analysis revealed that 1 suppressed the mRNA expression of 2 master regulators of adipocyte differentiation, PPARγ and C/EBPα. Furthermore, 1 downregulated the expression of adipogenesis-related genes, GLUT4, LPL, SREBP-1c, ACC, and FAS. Protein expression analysis revealed that 1 suppressed PPARγ protein expression. Moreover, to investigate the relationship between the structure and activity of inhibiting the adipocyte differentiation, we synthesized 12 kinds of phenylpropanoid analog. Comparison of the activity among 1 and its analogs suggested that the compound containing the substructure that possess a common functional group at the ortho position such as a catechol group exhibits the activity of inhibiting the adipocyte differentiation. Taken together, our findings suggest that 1 from wasabi leaves inhibits adipocyte differentiation via the downregulation of PPARγ.

Nymphaeol-C, a prenylflavonoid from Macaranga tanarius, suppresses the expression of fibroblast growth factor 18.

BACKGROUND: Fibroblast growth factor 18 (FGF18) is one of the key factors in human signaling pathways and has been reported to be associated with the formation of various tissues. Additionally, FGF18 has been reported to maintain the telogen stage of the hair cycle, and its over-expression has also been observed in cancer cells. HYPOTHESIS/PURPOSE: We searched for natural compounds that inhibit the expression of FGF18 expression in vitro and evaluated their inhibitory mechanisms. STUDY DESIGN: Various plant samples were screened using a luciferase assay targeting FGF18. One active compound was selected by the screening, isolated and identified. METHODS: The active compound was isolated using chromatographic techniques and identified by specific rotation measurements, LC-MS and NMR. Additionally, its inhibitory mechanism was evaluated using real-time RT-PCR and Western blotting. RESULTS: As a result of screening various plant leaf samples, Macaranga tanarius was identified as the most active plant and a prenylflavonoid nymphaeol-C was isolated as the active compound. Using real-time RT-PCR and Western blotting analysis, this compound was confirmed to strongly suppress the expression of FGF18. The compound lowered the ß-catenin level in the Wnt/ß-catenin pathway. Thus, it was suggested that nymphaeol-C suppresses the expression of FGF18 by suppressing ß-catenin expression. Additionally, the compound lowered the extracellular signal-regulated kinase1/2 (ERK1/2) phosphorylation level in the mitogen-activated protein kinase cascade (MAPK cascade). Therefore, nymphaeol-C suppressed downstream signals of FGF18 by suppressing the expression of FGF18. CONCLUSION: We isolated and identified prenylflavonoid nymphaeol-C from M. tanarius. The compound suppresses the expression of FGF18 and affects FGF18 related signals.

Effect of harvest time on some in vitro functional properties of hop polyphenols.

Health benefits of hop polyphenols are well studied. In the present investigation, we examined the antioxidant activity, anti-nitric oxide (NO) production, and anti-adipocyte differentiation of polyphenols extracted from hops (HPP), harvested at different times for over 4years. Saaz hop variety from the Czech Republic was used in the in vitro assays. Twenty-three polyphenols were quantitatively analyzed using Orbitrap liquid chromatography-mass spectrometry (LC-MS), and their health promoting effects were assayed individually. Strong effects of low concentrations of HPP were observed in the above three assays. A significant increase in anti-adipocyte differentiation activity per unit weight of HPP was obtained in the early harvested samples. A significant difference in anti-NO production activity per unit weight of HPP was observed among the different harvest years. HPP significantly increased in early harvested samples. Our results suggest that some in vitro functional properties of hops vary with harvest time and year and are dependent on different polyphenols.

In Vitro and In Vivo Antiangiogenic Activity of Crowberry (Empetrum nigrum var. japonicum).

Crowberry, Empetrum nigrum var. japonicum, is widely used in folk medicine and grows naturally in Korea. Although some constituents and biological activity of Korean crowberry have been examined, there is little detailed information available. In this study, we investigated the effects of ethanol extracts of crowberry (EECB) on the inhibition of angiogenesis, both in vitro and in vivo. The effects of EECB were tested on in vitro models of angiogenesis, that is, tube formation and proliferation of human umbilical vein endothelial cells (HUVECs). EECB exhibited significant inhibitory effects on tube formation of HUVECs in a concentration-dependent manner. In addition, crowberry significantly suppressed the proliferation of HUVECs in a concentration-dependent manner. Furthermore, strong antiangiogenic activity of EECB samples was observed in the in vivo assay using chick embryo chorioallantoic membrane (CAM). These results indicate that crowberry may have potential applications in the prevention and treatment of angiogenesis-dependent human diseases.

Antioxidant Activity of Phenolic Compounds from Fava Bean Sprouts.

Fava beans are eaten all over the world and recently, marketing for their sprouts began in Japan. Fava bean sprouts contain more polyphenols and l-3,4-dihydroxyphenylalanine (l-DOPA) than the bean itself. Our antioxidant screening program has shown that fava bean sprouts also possess a higher antioxidant activity than other commercially available sprouts and mature beans. However, the individual constituents of fava bean sprouts are not entirely known. In the present study, we investigated the phenolic compounds of fava bean sprouts and their antioxidant activity. Air-dried fava bean sprouts were treated with 80% methanol and the extract was partitioned in water with chloroform and ethyl acetate. HPLC analysis had shown that the ethyl acetate-soluble parts contained phenolic compounds, separated by preparative HPLC to yield 5 compounds (1-5). Structural analysis using NMR and MS revealed that the compounds isolated were kaempferol glycosides. All isolated compounds had an α-rhamnose at the C-7 position with different sugars attached at the C-3 position. Compounds 1-5 had ß-galactose, ß-glucose, α-rhamnose, 6-acetyl-ß-galactose and 6-acetyl-ß-glucose, respectively, at the C-3 position. The amount of l-DOPA in fava bean sprouts was determined by the quantitative (1) H NMR technique. The l-DOPA content was 550.45 mg ± 11.34 /100 g of the raw sprouts. The antioxidant activities of compounds 2-5 and l-DOPA were evaluated using the 2,2-diphenyl-1-picrylhydrazyl scavenging assay. l-DOPA showed high antioxidant activity, but the isolated kaempferol glycosides showed weak activity. Therefore, it can be suggested that l-DOPA contributed to the antioxidant activity of fava bean sprouts.

Principal component analysis of molecularly based signals from infant formula contaminations using LC-MS and NMR in foodomics.

BACKGROUND: The challenge in developing analytical assessment of unexpected excess contaminations in infant formula has been the most significant project to address the widespread issue of food safety and security. Foodomics based on metabolomics techniques provides powerful tools for the detection of tampering cases with intentional contaminations. However, the safety and risk assessments of infant formula to reveal not only the targeted presence of toxic chemicals, but also molecular changes involving unexpected contaminations, have not been reported. In this study, a huge amount of raw molecularly based signals from infant formula was analysed using reversed phase and hydrophilic interaction chromatography with time-of-flight MS (LC-MS) and (1) H nuclear magnetic resonance (NMR) and then processed by a principal component analysis (PCA). RESULTS: PCA plots visualised signature trends in the complex signal-data batches from each excess contamination of detectable chemicals by LC-MS and NMR. These trends in the different batches from a portion of excess chemical contaminations such as pesticides, melamine and heavy metals and out-of-date products can be visualised from spectrally discriminated infant formula samples. CONCLUSION: PCA plots provide possible attempts to maximise the covariance between the stable lot-to-lot uniformity and excess exogenous contaminations and/or degradation to discriminate against the molecularly based signals from infant formulas. © 2015 Society of Chemical Industry.

Anthocyanin Profile and Antioxidant Activity of Various Berries Cultivated in Korea.

This study examined the anthocyanin composition and antioxidant activity of various berries cultivated in Korea: blueberry, crowberry, Korean black raspberry, mulberry, and strawberry. The anthocyanins in berries were identified by high-performance liquid chromatography (HPLC) analysis, and each component was quantitatively analyzed. Furthermore, the antioxidant activity of berries was evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging, 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical cation decolorization, oxygen radical absorbance capacity (ORAC), and ferric ion reducing antioxidant (FRAP) assays. The results revealed that the total content of anthocyanins in crowberry was 35.1 mg/g of extract, which was higher than that in the other four major berry species (1.9-27.7 mg/g of extract). Nineteen anthocyanins were identified in the various berries. The major anthocyanins of crowberry were cyanidin-3-galactoside and delphinidin-3-galactoside, and those from Korean black raspberry were cyanidin-3-rutinoside and cyanidin-3-sambubioside-5-rhamnoside. These two berries also had relatively strong antioxidant activity accompanied by high total polyphenol contents. Thus, consumption of crowberry and Korean black raspberry may be beneficial in reducing the risk of developing lifestyle-related chronic diseases because of their strong antioxidant activity.

Scan-Free Absorbance Spectral Imaging A(x, y, λ) of Single Live Algal Cells for Quantifying Absorbance of Cell Suspensions.

Label-free, non-invasive, rapid absorbance spectral imaging A(x,y,λ) microscopy of single live cells at 1.2 µm × 1.2 µm resolution with an NA = 0.85 objective was developed and applied to unicellular green algae Chlamydomonas reinhardtii. By introducing the fiber assembly to rearrange a two-dimensional image to the one-dimensional array to fit the slit of an imaging spectrograph equipped with a CCD detector, scan-free acquisition of three-dimensional information of A(x,y,λ) was realized. The space-resolved absorbance spectra of the eyespot, an orange organelle about 1 µm, were extracted from the green-color background in a chlorophyll-rich single live cell absorbance image. Characteristic absorbance change in the cell suspension after hydrogen photoproduction in C. reinhardtii was investigated to find a single 715-nm absorption peak was locally distributed within single cells. The formula to calculate the absorbance of cell suspensions from that of single cells was presented to obtain a quantitative, parameter-free agreement with the experiment. It is quantitatively shown that the average number of chlorophylls per cell is significantly underestimated when it is evaluated from the absorbance of the cell suspensions due to the package effect.

Absorption and Metabolism of Luteolin and Its Glycosides from the Extract of Chrysanthemum morifolium Flowers in Rats and Caco-2 Cells.

To elucidate the bioavailability of luteolin and its glycosides in Chrysanthemum morifolium flowers, the absorption and metabolism of luteolin from them was investigated in rats and Caco-2 cells using HPLC and LC-MS. After oral administration of C. morifolium extract (1.7 g/kg body weight (bw), equivalent to 22.8 and 58.3 µmol/kg bw of luteolin and luteolin-7-O-glucoside, respectively) to rats, luteolin and its glycosides were quickly absorbed and luteolin, luteolin monoglucoside, and luteolin monoglucuronide were detected in the plasma. Their levels were highest at 1 h after administration (0.76 ± 0.27 µM). These compounds were also detected in media on the basolateral side from Caco-2 cells treated with the C. morifolium extract. These results suggest that luteolin and luteolin monoglucoside are rapidly absorbed after administration of C. morifolium flower extract and that luteolin, luteolin monoglucoside, and luteolin monoglucuronide may circulate in humans.

New acylated anthocyanins from purple yam and their antioxidant activity.

Purple yam (Dioscorea alata L.), which is widely distributed in tropical and subtropical regions, is characterized by its color and viscosity. Previous studies have shown that purple yams contain a variety of acylated anthocyanins that exhibit higher levels of antioxidant activity than the corresponding nonacylated compounds. In this study, the pigments found in purple yams from the Philippines (D. alata) were isolated and evaluated in terms of antioxidant activity. Four new acylated anthocyanins, alanins (1-4) were isolated from the MeOH extracts of purple yam, which were subsequently determined to be cyanidin (1, 2, and 4) and peonidin (3) type compounds, along with four known anthocyanins (5-8). The structures of 1-4 were determined by spectroscopic methods, including NMR and MS analyses. The antioxidant activities of anthocyanins 1-8 were investigated using oxygen radical absorbing capacity and ferric reducing antioxidant power assays.

Total Synthesis and Structure Determination of JBIR-108-A 2-Hydroxy-2-(1-hydroxyethyl)-2,3-dihydro-3(2H)-furanone Isolated from Streptomyces gramineus IR087Pi-4.

The planar and stereostructures of JBIR-108 isolated from Streptomyces gramineus IR087Pi-4 were determined partly by spectral analysis, and these structural assignments were confirmed and completed by the total synthesis of both 1-epimers. The key stereocenters in JBIR-108 were constructed via a Corey-Bakshi-Shibata (CBS) reduction (C-1), vinylogous Mukaiyama aldol reaction (C-7), and Brown crotylation (C-14 and C-15). Although it was difficult to determine the stereochemistries at the C-1 and C-7 positions in the natural product using the modified Mosher's method, the synthesis of two possible C-1 diastereomers enabled the identification of the configurations at the hitherto unknown stereocenters.

Sucutinirane-diterpene derivatives induce apoptosis via oxidative stress in HL-60 cells.

Previously, we reported the isolation of cassane-type diterpenes, sucutiniranes A-F, from the seeds of Bowdichia nitida. In this study, a series of sucutinirane derivatives was prepared, and their in vitro toxicity in the HL-60 cell line was evaluated. Then the action mechanism of a representative compound that induces cell death was investigated. Whereas C-6 or C-7 diol esters and ether decreased the activity against the HL-60 cell line, furan-oxidized derivatives 12 and 13 showed improvement or retention of the activity compared with those of the natural products sucutinirane A (11), E (1), and F (2). Treatment with sucutinirane derivative 13 elevated caspase 3/7 activity and also decreased expression of Bcl-2 family proteins, Mcl-1, and Bid. Derivative 13 generated reactive oxygen species in HL-60 cells, whose apoptotic effects were attenuated by the addition of an antioxidant, N-acetyl-L-cysteine. These results suggest that cassane butenolide 13 induces apoptosis in HL-60 via its oxidative effects.

Hawaiian propolis: comparative analysis and botanical origin.

Propolis is a resinous mixture of substances collected and processed from various botanical sources by honeybees (Apis mellifera). We recently obtained Hawaiian propolis, the study of which, to our knowledge, has not been reported. The purpose of this study was to analyze the composition of Hawaiian propolis and to identify its botanical origin. A comparative analysis of Hawaiian and Okinawan propolis and of the glandular trichomes on Macaranga tanarius fruit (the botanical origin of Okinawan propolis) was performed using reversed-phase high-performance liquid chromatography coupled with high resolution-electrospray mass spectrometry. Hawaiian propolis contained nine prenylflavonoids that were also isolated from Okinawan propolis. In conclusion, we suggest that the botanical origin of Hawaiian propolis is M. tanarius, the same as that of Okinawan propolis.

Epigallocatechin gallate decreases the micellar solubility of cholesterol via specific interaction with phosphatidylcholine.

The mechanisms underlying the effect of epigallocatechin gallate (EGCG) on the micellar solubility of cholesterol were examined. EGCG eliminated both cholesterol and phosphatidylcholine (PC) from bile salt micelles in a dose-dependent manner in vitro. When the bile salt micelles contained a phospholipid other than PC, neither cholesterol nor the phospholipid was eliminated following the addition of EGCG. When vesicles comprised of various phospholipids were prepared and, EGCG was added to the vesicles, EGCG effectively and exclusively eliminated only PC. An intermolecular nuclear Overhauser effect (NOE) was observed between PC and EGCG in bile salt micelles with EGCG added, but not between cholesterol and EGCG, by using a NOE-correlated spectroscopy nuclear magnetic resonance method. The results of binding analyses using surface plasmon resonance (SPR) showed that EGCG did not bind to cholesterol. These observations strongly suggest that EGCG decreases the micellar solubility of cholesterol via specific interaction with PC.

Identification of the phenolic compounds contributing to antibacterial activity in ethanol extracts of Brazilian red propolis.

The purpose of this study is to identify the quantity and antibacterial activity of the individual phenolic compounds in Brazilian red propolis. Quantitative analysis of the 12 phenolic compounds in Brazilian red propolis was carried out using reversed-phase high-performance liquid chromatography. The main phenolic compounds in Brazilian red propolis were found to be (3S)-vestitol (1), (3S)-neovestitol (2) and (6aS,11aS)-medicarpin (4) with quantities of 72.9, 66.9 and 30.8 mg g of ethanol extracts(- 1), respectively. Moreover, the antibacterial activities of each compound against Staphylococcus aureus, Bacillus subtilis and Pseudomonas aeruginosa were evaluated by measuring the minimum inhibitory concentrations. In particular, compound 4 exhibited the most potent antibacterial activity among all the assayed compounds against selected bacteria, indicating that 4 is the most active compound in Brazilian red propolis extracts. Thus, Brazilian red propolis may be used as food additives and pharmaceuticals to protect against bacteria.

Nymphaeol-A Isolated from Okinawan Propolis Suppresses Angiogenesis and Induces Caspase-Dependent Apoptosis via Inactivation of Survival Signals.

Propolis, a resinous substance that honeybees collect to protect their beehive from enemies, is reported to have various biological activities. In our screening program to search for antiangiogenic compounds from propolis, the ethanol extracts of Okinawan propolis (EEOP) showed significant antiangiogenic activities in a tube formation assay with human umbilical vein endothelial cells (HUVECs) in vitro at 3.13 µ g/mL and chorioallantoic membrane (CAM) assay in vivo at 25 µ g/egg. To elucidate the active compounds of EEOP and their mode of action, we isolated some prenylated flavonoids from EEOP and found that nymphaeol-A had the strongest antiangiogenic activity among them. Nymphaeol-A significantly reduced in vivo neovessel formation in the CAM assay at 25 µ g/egg. At the molecular level, nymphaeol-A markedly inactivated mitogen-activated protein kinase/ERK kinase 1/2 (MEK1/2) and extracellular signal-regulated kinase 1/2 (ERK1/2), whose molecular activations signal new vessel formation in HUVECs. In addition, nymphaeol-A dose- and time-dependently induced caspase-dependent apoptosis in tube-forming HUVECs. Taken together, nymphaeol-A was shown to inhibit angiogenesis at least in part via inactivation of MEK1/2-ERK1/2 signaling and induction of caspase-dependent apoptosis. Okinawan propolis and its major component, nymphaeol-A, may be useful agents for preventing tumor-induced angiogenesis.