Potent lipid peroxidation inhibitors from Withania somnifera fruits

A bioassay-guided purification of the methanolic extract of Withania somnifera fruits yielded novel withanamides A-I (1-9) and withanolides (10-13). Among the withanolides, compound 10 is novel. The structures of these compounds were determined by using FABMS, HRFABMS, 1D and 2D NMR spectral and chemical methods. The withanamides possess novel chemical structures and consisted of serotonin, glucose and long-chain hydroxyl fatty acid moieties. The stereochemistry of the hydroxyl group in the long-chain fatty acid moiety in compound 1 was determined by the modified Mosher's ester method. Compounds 1-13 were tested for their ability to inhibit lipid peroxidation in a model system using large unilamellar vesicles. Withanamides 1-5 and 9 inhibited lipid peroxidation by 98, 93, 79, 94, 81 and 86%, respectively, at 1 μg/mL. However, compounds 6-8 inhibited the lipid peroxidation by 85, 82 and 90%, respectively, at 0.5 μg/mL. Withanolides 10-13 were also tested and only compound 12 inhibited the lipid peroxidation by 82% at 10 μg/mL. To evaluate the structure activity relationships of withanamides A-I, compounds 14-16 were purchased and their lipid peroxidation activity determined as in the case of compounds 1-9. Commercial antioxidants, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and tert-butylhydroquinone (TBHQ), were also tested in this assay at 1 μg/mL and showed 80, 81 and 85% of inhibition, respectively. Our results suggest that the potent antioxidant activity exhibited by novel withanamides is probably due to the hydroxylated long-chain acyl group. This is the first report of withanamides, unique serotonin conjugates, from W. somnifera fruits.     

Dispersity of stratum corneum in the mixed aqueous solutions of binary mixtures between N,N-dimethyldodecylamine oxide and sodium dodecyl sulfate or two terpenes

Solubilization of cholesterol, differential scanning calorimetric (DSC), nuclear magnetic resonance (NMR) and dynamic light scattering (DLS) measurements were performed in order to reveal the dispersion mechanisms of stratum corneum (SC) into each intact corneocytes in the following systems: (1) in the aqueous mixed solutions of sodium dodecyl sulfate (SDS) and N,N-dimethyldodecylamine oxide (C₁₂DMAO); (2) in the aqueous micellar solutions of C₁₂DMAO containing solubilized α-terpineol (α-T); and (3) in the aqueous micellar solutions of C₁₂DMAO containing solubilized limonene. The intercellular lamellar structure of SC was revealed to be disrupted and/or removed in all these solutions. However, considering the micellar sizes and the interaction among molecules in micelle, the dispersion mechanisms in these three systems were different each other. The three dispersion mechanisms of SC were estimated and discussed on the basis of the results of solubilization, DSC, NMR and DLS, respectively.     

Influence of the level of cholesteryl sulfate in the solubilization of stratum corneum lipid liposomes by sodium dodecyl sulfate

 The role played by cholesteryl sulfate (Chol-sulf) in the solubilization of liposomes modeling the stratum corneum (SC) lipids by sodium dodecyl sulfate (SDS) was studied. We determined the surfactant-to-lipid molar ratios and the bilayer/aqueous phase surfactant partition coefficients of this interaction by varying the proportion of Chol-sulf, the relative proportions of the others lipids remaining constant. These parameters were determined by monitoring the changes in the static light scattering of the system during solubilization. The fact that the free surfactant concentration was always similar to its critical micelle concentration indicates that the liposome solubilization was mainly ruled by the formation of mixed micelles. The SDS ability to saturate and solubilize SC liposomes decreased as the proportion of Chol-sulf in the bilayers increased until a minimum was reached for a Chol-sulf proportion of about 15%. Inversely, the SDS partitioning into liposomes (or affinity with these bilayers) increased as the proportion of Chol-sulf increased until a maximum was reached at similar Chol-sulf proportions (10–15%). Hence, in these Chol-sulf proportions (similar to that existing in the intercellular lipids, which was 10%) the ability of SDS molecules to interact with liposomes exhibits a minimum despite their enhanced partitioning into liposomes. These effects may be related to the reported dependencies of the level of Chol-sulf on the abnormalities in the skin barrier function and on the SC intercellular cohesion. Received: 12 October 1999 Accepted: 20 January 2000     

Influence of the hydrophobic tail of alkyl glucosides on their ability to solubilize stratum corneum lipid liposomes

The lytic interactions of a series of alkyl glucosides (alkyl chain lengths ranging from C₈ to C₁₂) with liposomes formed by a mixture of lipids modeling the stratum corneum (SC) lipid composition were investigated. The surfactant-to-lipid molar ratios (Re) and the normalized bilayer/aqueous phase partition coefficients (K) were determined by monitoring the changes in the static light-scattering (SLS) of the system during solubilization. The fact that the free surfactant concentrations were always similar to their critical micelle concentrations indicates that the liposome solubilization was mainly ruled by the formation of mixed micelles. At the two interaction levels studied (100 and 0% SLS) the nonyl glucoside showed the highest ability to saturate and to solubilize liposomes (lowest Re values), whereas the dodecyl glucoside showed the highest degree of partitioning into liposomes or affinity with these structures (highest K values). Comparison of the data for octyl glucoside with that reported for the interaction of this surfactant with phosphatidylcholine (PC) liposomes shows that whereas the SC lipid liposomes were more resistant to the action of this surfactant (higher Re values), its degree of partitioning into SC bilayers was both in the saturation and solubilization of liposomes similar to that exhibited in PC vesicles (similar K values). Received: 27 November 2000/Accepted: 19 February 2001     

Evaluation of lipid peroxidation inhibition and free radical scavenging abilities of 5,6,7-trimethoxy dihydroflavonols

Four naturally rare 5,6,7-trimethoxy-2,3-cis-dihydroflavonols(3-6) and two 5,6,7-trimethoxy-2,3-trans-dihydroflavonols(7-8) were designed and synthesized.Their antioxidative properties were evaluated by way of examining their scavenging capacities towards DPPH and O_2~(·-) free radicals,as well as by measuring their inhibitory ability against LPO.Both the 2,3-trans and the 2,3- cis conformers exhibited certain quenching abilities to DPPH and O_2~(·-) radicals,while most of the synthetic dihydroflavonols ...     

FTIR and NMR study of poly(lactide-co-glycolide) and hydroxyapatite implant degradation under in vivo conditions

The influence of hydroxyapatite (HAP) addition on the rate and mechanism of lactide-co-glycolide copolymer (PGLA) degradation after implantation (in vivo study) was analyzed and compared with the process taking place during in vitro studies. Structural and phase changes of poly(lactide-co-glycolide) and its composite with hydroxyapatite were determined using IR and NMR spectroscopy.Degradation of PGLA and PGLA + HAP composite in biological environment proceeds faster than under in vitro condition. Concentration of glycolidyl units in the copolymer chain decreases and that of lactidyl units increases during in vivo degradation both, in PGLA and in PGLA + HAP composite. However, in the case of the composite the decrease of glycolidyl units concentration is slower and after 6 weeks of degradation the contents of lactidyl and glycolidyl units remain stable. On the other hand, PGLA + HAP composite degrades faster than pure PGLA. The addition of HAP nanoparticles distinctly accelerates degradation of PGLA copolymer which is probably connected with the increase of hydrophilicity of the composite and inhibition of semi-crystalline lactidyl domains formation during the degradation process. Observation of the bone tissue after implantation of PGLA + HAP allows to conclude that the degradation of the composite occurs simultaneously with the implant replacement by the bone cells.     

Reduction of the IgE-binding ability and maintenance of immunogenicity of gamma-irradiated Dermatophagoides farinae

House dust mites, Dermatophagoides farinae (DF) and Dermatophagoides pteronyssinus, are major allergens in the most common indoor allergen and are important risk factor for asthma. The modified antigen has been studied to treat allergic disorder. This study was carried out to measure possibility of modified allergen using gamma irradiation to treat allergy such as asthma. DF solutions (2 mg/ml) as target allergen were irradiated with Co-60 at 50 and 100 kGy. Conformational alternation of irradiated DF was observed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Levels of anti-irradiated DF mouse IgGs (sub-isotypes) against intact DF were measured similar to that of anti-intact DF IgGs. The binding abilities of house dust mite-allergic patients’ IgE were reduced depending on radiation dose, and irradiation could inhibit the binding ability of patients’ IgE more than 40%. This study has shown that the binding ability of IgE was reduced by conformational alteration by irradiation and the irradiated DF had epitopes capable to induce immunogeniciy.     

五味子中木脂素类成分的高效液相色谱-电喷雾质谱研究

采用液相色谱-电喷雾质谱联用(HPLC-ESI-MSⁿ)技术, 对北五味子与南五味子中木脂素类成分进行了系统研究. 通过HPLC-ESI-MS技术, 获得了相应化合物的保留时间、紫外光谱和分子量等信息, 利用电喷雾多级串联质谱技术(ESI-MSⁿ), 获得了相应化合物的结构信息. 研究结果表明, 北五味子与南五味子的主要木脂素成分除5个共有成分外其它成分差异较大, 并且其共有成分含量差别较大. 在此基础上, 建立了简便、快速的北五味子与南五味子药材分析鉴定的新方法.     

Improved postharvest quality in patagonian squash (Cucurbita moschata) coated with radiation depolymerized chitosan

Different molecular weight chitosans were evaluated on the decay of coated Anquito squashes (Cucurbita moschata) as well as the maintenance of the fruit quality along five storage months. The original chitosan (Mw=391 kDa, 83% DD), was depolymerized by gamma radiation. Apart from chain scission, other chemical changes were not detected by FTIR or UV–vis analyses. The molecular weight characterization of chitosans was done by size exclusion chromatography with dual light scattering and concentration detection (SEC-MALLS-RI). The coating effectiveness was evaluated on the following parameters: fungal decay incidence, weight loss, firmness, total reducing sugar, soluble solid, flesh color, carotene content, pH and titratable acidity. No sign of fungal decay was observed in squashes coated with 122 and 56 kDa chitosans, which were also the most effective treatments in reducing the weight loss. The chitosan with Mw=122 kDa was also the best treatment considering firmness, internal aspect, sugar and carotene content. Then, radiation degraded chitosan was better in C. moschata preservation than the original chitosan.     

Detection of Phakopsora pachyrhizi by polymerase chain reaction (PCR) and use of germination test and DNA comet assay after e-beam processing in soybean

Soybean harvest is the main agribusiness in Brazil, which is the second largest exporter in the world and has a revenue of billions of dollars. Asian dust is caused by the fungus Phakopsora pachyrhizi and its dissemination is difficult to control, since it occurs through wind dispersion. Actually P. pachyrhizi is found in different parts of the world. Electron beam treatment could be an alternative process to minimize these losses, especially for the grains exportation industry. Besides the possibility of being disconnected when not in use, this source does not need to be reloaded, is easily available and, streamlines the process and reduces logistics costs. The present work aims to identify, by the polymerase chain reaction technique (PCR), the P. pachyrhizi fungus presence in the irradiated soybeans and the possibility to use radiation treatment as a sanitary alternative. Doses 0, 1.0, 2.0, 5.0, 6.0, 7.0, 8.0, 9.0 and 10.0 kGy (IPEN-CNEN/SP Electron Accelerator) were applied and two fast-screening methods were performed: DNA comet assay (for the detection of DNA damage) and germination test (for the measurement of roots inhibition). These tests are very easy to carry out and measure damage response depending on radiation dose.     

Characterization of heat-induced interaction of neutral liposome with lipid membrane of Streptomyces griseus cell

The interaction between the neutral 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) liposomes and cell membrane of Streptomyces griseus induced by the heat treatment at specific temperature was investigated, focusing on the internalization of the neutral POPC liposomes with S. griseus cells. In an attempt to clarify the modes of liposome internalization, various kinds of inhibitors of endocytotic pathways were used to treat S. griseus cells. The efficiency of the heat treatment on liposome–cell membrane interactions was finally characterized based on the hydrophobic, electrostatic interactions and hydration effect. In fact, the internalization of the neutral liposomes into these cells was found to show higher rate and greater amount at higher temperatures. The kinetic study showed that the maximum amount of the internalized liposomes was, respectively, 469 × 10⁵ and 643 × 10⁵ liposomes/cell at 37 and 41 °C. The internalization of the neutral liposomes induced by the heat treatment was characterized, implying that the endocytosis occurred. The interactions involving the internalization, adsorption, and fusion of these liposomes with S. griseus cells were mainly contributed by the hydrophobic interaction and the unstable hydrogen bonds caused by the loss of water of surface hydration of cell membrane rather than the electrostatic interaction under the specific heat condition.     

Microsensor in vivo monitoring of oxidative burst in oilseed rape (Brassica napus L.) leaves infected by Sclerotinia sclerotiorum

Oxidative burst is the rapid and transient production of large amounts of reactive oxygen species, including superoxide anion, hydrogen peroxide (H₂O₂), and hydroxyl radical. A rapid and simple technique was employed for in vivo detection of oxidative burst in oilseed rape (Brassica napus L.) leaves, using a modified electrode. Platinum (Pt) micro-particles were dispersed on a Pt electrode, coated with a poly (o-phenylenediamine) film. This exhibited high sensitivity, selectivity and stability in H₂O₂ detection. Amperometry was used to obtain satisfactory linear relationships between reductive current intensities and H₂O₂ concentrations at −0.1 V potential in different electrolytes. This electrode was used in vivo to detect oxidative burst in oilseed rape following fungal infection. Oxidative bursts induced by infection of the fungal pathogen Sclerotinia sclerotiorum (Lib.) de Bary exhibited notably different mechanisms between a susceptible and a resistant glucose oxidase-transgenic genotype.     

Calorimetric study of the effect of protoberberine alkaloids in Coptis chinensis Franch on Staphylococcus aureus growth

The effects of five protoberberine alkaloids (PBAs) from rhizoma of Coptis chinensis Franch on Staphylococcus aureus growth were investigated by calorimetry. The power-time curves of S. aureus with and without PBA were measured in closed glass ampoules in a TAM Air isothermal calorimeter. And, the extent and duration of inhibitory effects on the metabolism were evaluated by growth rate constant (k), half inhibitory ratio (IC₅₀), maximum heat-output (P_max) and peak time (t_p). The obtained calorimetric data showed that the inhibitory action varied for different protoberberine alkaloid. The results also revealed that the sequence of antimicrobial activity of five PBAs was: berberine>coptisine>palmatine>epiberberine>jatrorrhizine. One explanation could be substitutions at several positions in the core structure of berberine possess different antimicrobial activity. In conclusion, it can be proposed that this technique should be as a useful analytical method for determining the bioactivity of PBAs.     

Determination of volatile compounds in foliage of Fraser fir (Abies fraseri) and balsam fir (Abies balsamea)

The Fraser fir (Abies fraseri) and balsam fir (Abies balsamea) are eastern North American conifers which have been infested by an exotic insect, the balsam woolly adelgid (BWA). BWA infestation has had particularly severe effects on Fraser fir, with up to 95% mortality rates at some sites, and is characterized by attack on mature trees only. The purpose of this research was to perform a chemosystematic study to evaluate whether differences in volatile chemical concentrations of various stands of fir were observed as a function of resistance to BWA infestation. The concentrations of volatiles were determined by a methylene chloride extraction procedure, followed by analysis by gas chromatography. First, comparisons were made of concentration levels of volatiles in Fraser and balsam fir foliage of seedlings, saplings, and mature trees. If a chemical provided resistance, one would expect higher volatile levels in the balsam foliage because of its greater resistance to BWA. Second, the volatile levels in Fraser fir saplings and mature trees at uninfested sites were compared to the levels in surviving Fraser fir saplings and mature trees at infested sites. For a compound that provided BWA-resistance, higher volatile levels would be expected at the infested site because of the greater resistance of the surviving trees. Lastly, the concentrations of volatiles in sapling foliage were compared to those in mature foliage, where higher levels of resistance-providing chemicals would be expected in the saplings. 3-Carene was shown to consistently follow the expected pattern for a compound that provides resistance against BWA and β-pinene followed the pattern for the majority of the comparisons. These results indicate that while maltol and total volatiles did not correlate with the expected pattern, 3-carene, and possibly β-pinene and sesquiterpenes may provide fir with defense against BWA infestation.     

Novel megastigmanes with lipid accumulation inhibitory and lipid metabolism-promoting activities in HepG2 cells from Sedum sarmentosum

Four novel megastigmanes, neosedumosides I (1), II (2), III (3), and IV (4) were isolated from the whole plant of Sedum sarmentosum (Crassulaceae). Absolute stereostructures of these constituents were determined on the basis of chemical and physicochemical evidence. Among them, 1-3 were found to show lipid accumulation inhibitory activity in HepG2 cells. Furthermore, 2 and 3 were found to also show lipid metabolism-promoting activity.     

Applications of in vivo and in vitro solid-phase microextraction techniques in plant analysis: A review

As a very popular sample preparation technique, solid-phase microextraction (SPME) coupled with various analytical instrumentation, has been widely used for the determination of trace levels of different plant compounds, such as volatile organic compounds (VOCs) emitted from the different plant organs, and environmental contaminants in plants. In this review, recent applications of in vitro and in vivo SPME in plant analysis are discussed and summarized according to the different organs of plants, including fruits, flowers, leaves, stems, roots and seeds, and the whole plant as well. Future developments and applications of SPME in plant analysis, especially in vivo sampling approaches, are also prospected.     

Qualitative and quantitative analyses of three bioactive compounds in different parts of Forsythia suspensa by high-performance liquid chromatography-electrospray ionization-mass spectrometry

A high-performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) analytical method was developed to simultaneously detect and quantify three main distinctive compounds (forsythiaside, rutin and forsythin) in different parts of Forsythia suspensa (F. suspensa), an herbal medicine. This was the first report on the quantification of bioactive constituents in different parts of F. suspensa by HPLC-ESI-MS analytical method. The calibration curves of the three compounds showed good linearity (R² > 0.9994). The method was reproducible with intra- and inter-day variation less than 1.35% and 2.00%, respectively. The recovery of the assay was in the range of 98.27–101.07%. The results indicated that the developed assay could be considered as a suitable quality control method for this commonly used herbal medicine.     

A correlation between the virulence and the adhesion of Listeria monocytogenes to silicon nitride: An atomic force microscopy study

Listeria monocytogenes is a facultative intracellular Gram-positive bacterium that is widely distributed in the environment. Despite being pathogenic at the species level, L. monocytogenes in fact comprises a diversity of strains from pathogenic ones that can result in disease and/or mortality to others that are relatively avirulent. The main goal of the current study was to answer the question on whether enhanced binding or attachment of L. monocytogenes to inert surfaces bears any relationship to pathogenicity in food-borne isolates. To answer this question, the nanoscale adhesion forces of eight L. monocytogenes strains that vary in their pathogenicity levels to a model surface of silicon nitride were quantified using atomic force microscopy. The strains used were the highly pathogenic (EGDe, 874, 1002, ATCC 19115), the intermediate pathogenic (ATCC 19112, ATCC 19118), and the non pathogenic (ATCC 15313 and HCC25). Our results indicate that the average nanoscale adhesion (in nN) and the 50% lethal dose (LD50) of strain virulence quantified in mice are logarithmically correlated according to: (nN) = −0.032 ln (LD50) + 1.040, r² = 0.96. Such correlation indicates that nanoscale adhesion could potentially be used as a design criterion to distinguish between virulent and avirulent L. monocytogenes strains. Finally, stronger adhesion of virulent strains to inert surfaces modeled by silicon nitride might be a way for pathogenic strains to survive better in the environment and thus increase their likelihood of infecting animals or humans.     

纳米钙钛矿型LaFe1-xNixO3(x=0~1.0)的制备及其在紫外光激发下的电化学性能

本文采用溶胶-凝胶法制备了LaFe1-xNixO3(x=0,0.2,0.4,0.6,0.8,1.0)纳米晶粉末,利用XRD、TEM和电化学测试方法对LaFe1-xNixO3材料的相结构、形貌、成分组成和其在碱液中的充放电性能以及电化学动力学性能等方面进行了表征和分析,同时对电极受紫外光激发前后的电化学行为进行了对比研究。XRD和TEM分析表明,用硝酸盐作为原材料和溶胶-凝胶方法可制备出单一相结构的纳米晶钙钛矿型LaFe1-xNixO3复合氧化物,随Ni替代量x的增大,LaFe1-xNixO3的相结构由正交结构向菱面体结构转变,其分子体积和晶粒尺寸呈现减小的趋势。电化学研究结果表明,紫外光激发前,LaFe1-xNixO3电极的放电容量随x的增加而逐渐增大;光激发后,电极的放电容量和交换电流Io与未激发前相比显著提高,当x=0.4时其放电容量具有最大值483.1mAh·g-1,Io值由光激发前的3.54～11.58 mA·g-1大幅增加至激发后的8.37～40.11 mA·g-1。