Gas-liquid Chromatography-mass Spectroscopy Research Articles

Bioprospection of Antiviral and Antitumor Compounds from Some Marine Algae from Egyptian Shores.

The marine algae are considered a diverse source of bioactive compounds. Many active compounds have been isolated from algae and show good biological activities. The aim of this study is to detect the antiviral and anticancer activities in some extracts of marine algae. Extraction, purification and identification of some marine algae common in Egypt were conducted. Extraction of Ulva lactuca, Sargassum dentifolium, and Cystoseiara myrica was conducted. A sequence of extractions, including extraction by ethanol, boiling water, hydrochloric acid and sodium hydroxide were carried out. The obtained extracts were evaluated for their antitumour and antiviral activities against liver tumour cells, brain tumour cell lines, measles virus, mumps virus and hepatitis B virus (HBV). The extracts of the best activities were subjected for purification by size exclusion chromatography and anion exchange chromatography for ethanolic extracts or precipitation by cetylpyridinium chloride (CPC) then by size exclusion chromatography and anion exchange chromatography for aqueous extracts. Separation by GLS/MS was performed. The structures of the active compounds have been identified through different chemical analyses, including sugar analysis, configurational analysis, High-Performance Liquid Chromatography (HPLC), infrared spectroscopy (IR), Gas-Liquid Chromatography-Mass Spectroscopy (GLC-MS) and 1H,13C nuclear magnetic resonance (NMR) at ZV. The active compounds from the water extracts have been identified mainly as polysaccharides and sulphated polysaccharides. The antitumour and the antiviral activities of ethanolic extracts are attributable to compound identified as Ethyl Palmitate. These natural compounds did not show cytotoxic effect. These outputs could be preliminary for further biological studies aiming to therapeutic application.

Phytochemical Biological and Cytotoxic Activities of Calothamnus quadrifidus Leaves Cultivated in Egypt

The hydro distilled essential oil obtained from Calothamnus quadrifidus leaves was analyzed by gas liquid chromatography-mass spectroscopy. Quantification of the oil constituents was carried out using the gas liquid chromatography-flame ionization detection device where 1,8-cineole was quantified as 525.62 μg/ml. The oil showed potential antioxidant activity when tested by 1,1-diphenyl-2-picrylhydrazine radical scavenging assay where the concentration required to scavenge 1,1-diphenyl-2-picrylhydrazine by 50 %=37.3 μg. The cytotoxic activity of the essential oil was evaluated using 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide assay compared with 1,8-cineole and doxorubicin. Calothamnus quadrifidus leaves essential oil cytotoxic activity was significant against human lung cell line A-549 (2.37 μg), human colon cell line HCT-116 (2.39 μg), human hepato-cellular cell line HepG-2 (2.55 μg) and intestinal epithelium carcinoma Caco-2 (5.80 μg) cell lines, respectively. The oil cytotoxicity exceeded 1,8-cineole except for intestinal epithelium carcinoma Caco-2. Additionally, the antimicrobial activity was evaluated using agar disc diffusion and minimum inhibitory concentration where the oil exhibited the best activity against Penicillium italicum and Geotrichum candidum with minimum inhibitory concentration=0.49 and 0.98 μg, respectively. Structure elucidation of the pure compounds isolated from Calothamnus quadrifidus leaves was done according to their chromatographic behavior, chemical and spectroscopic data. These compounds were isolated for the first time from Calothamnus quadrifidus leaves and identified as gallic acid, gallic acid methyl ester, 7,4`-dimethoxy-apigenin, 6,7-dimethoxy-apigenin, (Bis[3(2-methyl propan-1- one)-2, 6 dihydroxy-4- methoxy phenyl] methane, quercetin 3-O-beta-D-arbinopyranoside and quercetin 3-O-beta-D-4C1-xylopyranoside. In addition to the molecular docking of novel compound 3 was done in peroxiredoxins and the binding mode was found. Conformation 2 was found to be the most stable and may be responsible for the antioxidant effect as it showed the best free binding energy and affinity when compared to conformations.

Antibacterial activities and bioactive compounds of the ethyl acetate extract of the sea cucumberHolothuria forskalifrom Tunisian coasts

AbstractPrevious scientific studies have listed chemical compounds from sea cucumber species. The ability to produce bioactive metabolites derives from their response to various ecological stresses and avoidance of predation. In view of this, the present work aimed to evaluate the antibacterial activity and quantitatively assess the active components of different organs of the sea cucumberHolothuria foskalicollected from the north-east of Tunisia. The antibacterial activity of ethyl acetate extracts was tested againstBacillus subtilis,Bacillus cereus,Escherichia coliandPseudomonas aeruginosa. It was revealed thatEscherichia coliandBacillus subtiliswere inhibited by the sea cucumber extracts. To identify the composition of these extracts, Gas Chromatography-Mass Spectroscopy (GC-MS) and Liquid Chromatography-Mass Spectroscopy (LC-MS) methods were used. The GC-MS revealed the presence of 25 major components, the most concentrated ones were characterized as C26H27N5O6Morphinan-4,5-epoxy-3,6-diol, 6 amino;C17H31F3O23-trifluoroacetoxypentadecane; C31H50O2Stigmasta-5,22-dien-3-ol,acetate; C11H14O21-Phthalanol,1,3,3-trimethyl 1-phenyl; C30H48O2Ergosta-5,22-dien-3-ol,acetate and C25H40O2(9E)-8-Phenyl-9-octadecenoic acid methyl ester; while the LC-MS revealed the presence of eight molecules, identified as phenolic acid, phytosterol and fat-soluble vitamins. Some molecules are interesting and deserve to be explored for their potential uses as ingredients in functional foods or as raw materials for new drug forms.

Chemical profiling of Phlomis thapsoides (Lamiaceae) and in vitro testing of its biological activities

A phytochemical study of Phlomis thapsoides (Lamiaceae) resulted in the isolation of one new compound, 6,10,17-trimethyl-2-octadecanone, and three known compounds (sitosterol-3-O-β-glucoside, and the iridoid glucosides ipolamiide and lamiide). The structures of the isolated compounds were elucidated using mass spectrometry, 1D/2D NMR spectroscopy experiments in comparison with published data. The chemical composition of the essential oil obtained from aerial parts of P. thapsoides was determined by gas liquid chromatography and gas liquid chromatography-mass spectroscopy. The main volatile constituents were phenylethyl alcohol (6.81 %), trans-3-hexenol (5.55 %), 1-octen-3-ol (5.10 %), α-cadinol (4.92 %), and α-muurolol (4.67 %). The antioxidant activity of the extracts was evaluated by three methods: 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activity, 2,2′-azinobis-[3-ethylbenzthiazoline-6-sulfonic acid] and ferric reducing antioxidant power assay along with the determination of total phenolic content. Although the ethyl acetate extract contained 362.06 mg of gallic acid equivalents, the antioxidant activities, as revealed by 1,1-diphenyl-2-picrylhydrazyl, 2,2′-azinobis-[3-ethylbenzthiazoline-6-sulfonic acid] and ferric reducing antioxidant power assays, were of medium strength (IC50 of 73.08 ± 3.18, 9.48 ± 0.71 µg/mL and 9.33 ± 0.17 mM of FeSO4 equivalents, respectively). Only lamiide inhibited soybean 5-lipoxygenase with an IC50 value of 72.92 µg/mL in vitro. In-silico molecular modeling studies on 5-lipoxygenase and human 5-lipoxygenase-activating protein were used to evaluate the potential anti-inflammatory activity. Sitosterol-3-O-β-glucoside followed by the iridoid glucoside lamiide exhibited the highest inhibition of 5-lipoxygenase whereas the new compound 6,10,17-trimethyl-2-octadecanone and sitosterol-3-O-β-glucoside exhibited the highest inhibition of 5-lipoxygenase-activating protein as evidenced from their higher fitting scores. The cytotoxicity of the plant extracts and lamiide against Caco2 and HepG-2 cancer cells resulted in IC50 values of >100 µg/mL indicating a low cytotoxicity.

Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars.

BackgroundGrape cultivars and wines are distinguishable by their color, flavor and aroma profiles. Omic analyses (transcripts, proteins and metabolites) are powerful tools for assessing biochemical differences in biological systems.ResultsBerry skins of red- (Cabernet Sauvignon, Merlot, Pinot Noir) and white-skinned (Chardonnay, Semillon) wine grapes were harvested near optimum maturity (°Brix-to-titratable acidity ratio) from the same experimental vineyard. The cultivars were exposed to a mild, seasonal water-deficit treatment from fruit set until harvest in 2011. Identical sample aliquots were analyzed for transcripts by grapevine whole-genome oligonucleotide microarray and RNAseq technologies, proteins by nano-liquid chromatography-mass spectroscopy, and metabolites by gas chromatography-mass spectroscopy and liquid chromatography-mass spectroscopy. Principal components analysis of each of five Omic technologies showed similar results across cultivars in all Omic datasets. Comparison of the processed data of genes mapped in RNAseq and microarray data revealed a strong Pearson’s correlation (0.80). The exclusion of probesets associated with genes with potential for cross-hybridization on the microarray improved the correlation to 0.93. The overall concordance of protein with transcript data was low with a Pearson’s correlation of 0.27 and 0.24 for the RNAseq and microarray data, respectively. Integration of metabolite with protein and transcript data produced an expected model of phenylpropanoid biosynthesis, which distinguished red from white grapes, yet provided detail of individual cultivar differences. The mild water deficit treatment did not significantly alter the abundance of proteins or metabolites measured in the five cultivars, but did have a small effect on gene expression.ConclusionsThe five Omic technologies were consistent in distinguishing cultivar variation. There was high concordance between transcriptomic technologies, but generally protein abundance did not correlate well with transcript abundance. The integration of multiple high-throughput Omic datasets revealed complex biochemical variation amongst five cultivars of an ancient and economically important crop species.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2115-y) contains supplementary material, which is available to authorized users.

Chemical Composition of the Essential Oils of Variegated Pink-Fleshed Lemon (Citrus x limon L. Burm. f.) and their Anti-Inflammatory and Antimicrobial Activities

The volatile secondary metabolites of essential oils from fruit peel and leaves of variegated pink-fleshed lemon (Citrus x limon) were investigated using GLC and GLC-MS (gas-liquid chromatography-mass spectroscopy). Altogether 141 compounds were identified and quantified, accounting for 99.59% and 96.33% of the total hydrodistilled peel and leaf oil, respectively. Limonene occurred in higher amounts in fruit peel (52.73%) than in leaf oil (29.13%). Neral (12.72%), neryl acetate (8.53%), p-menth-1-en-7-al (4.63%), beta-pinene (6.35%), and nerol (4.42%) were the most abundant constituents in leaf oil, whereas gamma-terpinene (9.88%), beta-pinene (7.67%), geranial (4.44%), and neral (3.64%) dominated in the fruit peel oil. The antioxidant, anti-inflammatory, antitrypanosomal, and antimicrobial activities of the fruit peel essential oil were evaluated. The oil had a low antioxidant activity with an IC50 value of (26.66 +/- 2.07) mg/ml as compared to the efficient antioxidant ascorbic acid [IC50 (16.32 +/- 0.16) microg/ml]. The oil moderately inhibited soybean 5-lipoxygenase (5-LOX) with an IC50 value of (32.05 +/- 3.91) microg/ml and had moderate antitrypanosomal activity [IC50 (60.90 +/- 0.91) microg/ml]. In addition, moderate antimicrobial activities were detected against Gram-positive bacteria (Bacillus subtilis, Staphylococcus capitis, Micrococcus luteus), one Gram-negative bacterium (Pseudomonas fluorescens), and yeasts (Saccharomyces cerevisiae, Candida parapsilosis).

Biodesel Production from Pseudomonas Fluorescens Lp1 Lipase Immobilized on Amino-silane Modified Super Paramagnetic Fe3O4 Nanoparticles

An extracellular lipase from Pseudomonas fluorescens Lp1 isolated from oil contaminated soil was immobilized onto amino silane modified superparamagnetic Fe3O4 nanoparticles. The magnetic nanoparticles, magnetite was synthesized chemically by co-precipitation and characterized by Scanning Electron Microscopy (SEM), Fourier Transformed Infrared Spectroscopy (FT-IR) and Powder X-ray diffraction studies (XRD). The structure of the synthesized magnetic nanoparticles was uniform, spherical and the size was determined around 31 nm by powder XRD. The biodiesel production mixture was prepared by addition of waste cooking oil, lipase immobilized magnetite and methanol. The transesterified products were analyzed by Gas Liquid chromatography-Mass spectroscopy (GC-MS). The methyl esters such as Oxiraneundecanoic acid, 3-pentyl-methyl ester, Hexadecanoic acid, methyl ester and 10-Octadecenoic acid, methyl ester were obtained. The study experimentally proved the use of amino silane modified superparamagnetic Fe3O4 nanoparticles in biodiesel production from waste cooking oil.

Chemical, chromatographic, and thermal analysis of rosemary (Rosmarinus officinalis)

AbstractRosemary (Rosmarinus officinalis) is mainly known for its cooking qualities, but it possesses an essential oil with pharmacological properties. Moreover, this plant was the start of forest fires in Provence, France. The presence of biopolymers can contribute to electrical energy production by biomass combustion. The aim of this work was to study a stalk of rosemary during its biological cycle (November 1998 to June 1999). The chemical analysis (water, mineral ashes, cellulose, lignin, holocellulose, and extractives) showed few variations during this period. Leaves and branches differentiated themselves according to their chemical composition. Analyzed by chromatography gas–liquid chromatography/mass spectroscopy (GC‐MS) and gas–liquid chromatography/flam ionization detector (GC‐FID), the essential oil of this rosemary had a high level of camphor (30–45%), which could be used in new therapeutics (cardiac and respiratory analeptic). Finally, the simultaneous thermal analysis (DTA‐TG) was run on the fresh plants and the dried powdered samples. Several kinetic constants of the biopolymers had been computed: activation energies of cellulose and holocellulose. During heating, the rosemary's lignin decomposed more than others plants, thereby increasing the fire risks. The chemical, chromatographic, and thermal analysis run on the rosemary could be applied on other species in the Mediterranean basin. © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 83: 747–756, 2002

Fatty acid characteristics in two symbiont-bearing mussels from deep-sea hydrothermal vents of the south-western Pacific

Two bacteria-bearing mussels,Bathymodiolus breviorandB. elongatus, collected at five deep-sea hydrothermal sites from Lau and Fiji back arc basins (south-west Pacific), were analysed for polar and neutral lipids using gas-liquid chromatography mass spectroscopy (GLC-MS). The respective fatty acid distributions found in gills and mantles were compared to data from a Mediterranean littoral filter-feeding mussel,Mytilus galloprovincialis. The main characteristic common to the gills of the two vent mussels was the high level of (n-7) monounsaturated phospholipid fatty acids, which are supposed to originate from sulphur-oxidizing symbionts. Using palmitoleic, 16:1 (n-7), and vaccenic, 18:1 (n-7), phospholipid fatty acids as bacterial markers, endobacterial biomass was estimated to account for 2–10% of the vent mussel gill volume. Vent mussel tissues were poor in plant-derived polyunsaturated fatty acids, indicating the limited importance of the photosynthesis based material in the metabolism of these hydrothermal vent bivalves. It is suggested that some compounds, such as nonmethylene-interrupted dienoic fatty acids and specific trienoic fatty acids originating from elongation and desaturation of oleic and palmitoleic acids, respectively, were synthesized in order to substitute exogenous plant-derived polyunsaturated fatty acids considered as essential in marine invertebrates. The high level of neutral lipids in vent mussel gills suggests that these compounds are used as a major energy storage material. The neutral lipid/phospholipid ratio in gills emphasized the preponderance of the endosymbionts in vent mussel nutrition. Using this ratio as an index of the physiological state, the symbiosis patterns in different hydrothermal fields were compared.

Fatty Acid Composition of Symbiotic Cyanobacteria from Different Host Plant (Azolla) Species: Evidence for Coevolution of Host and Symbiont

The total cellular fatty acid contents of 40 recently isolated cyanobacterial symbionts obtained from seven species of Azolla host plants were determined by gas-liquid chromatography-mass spectroscopy. A total of 63 fatty acids belonging to seven distinct chemical classes were identified. Fatty acid compositions varied among the cyanobacteria depending on the hosts species. Parameters that differed significantly (at the 99% level of probability) included the concentrations of the 16:0 and 18:3 fatty acids, the total concentrations of the polyunsaturated acids, the total concentrations of the 16-carbon and 18-carbon fatty acids, the ratios of unsaturated fatty acids to saturated fatty acids, and the total percentages of straight-chain even-carbonnumber fatty acids, unsaturated fatty acids, and branched-chain unsaturated fatty acids. The results of an analysis of variance suggested statistical regression for the total percentages of these fatty acids and chemical classes according to the following linear alignment of cyanobacteria by host: Azolla filiculoides, Azolla microphylla, Azolla caroliniana, Azolla mexicana, Azolla rubra, Azolla nilotica, and Azolla pinnata (including Azolla pinnata subsp. pinnata and Azolla pinnata subsp. imbricata). The seven groups could be divided into two distinct clusters on the basis of the results of a dendrogram analysis of Euclidian distances. The symbionts obtained from A. filiculoides, A. microphylla, A. mexicana, and A. caroliniana constituted one cluster, and the symbionts obtained from A. rubra, A. nilotica, and A. pinnata constituted a second cluster. A minor dichotomy separated the A. filiculoides symbionts from the other members of the first cluster. The clustering of Azolla cyanobacterial symbionts based on the results of our fatty acid analysis correlates remarkably well with the taxonomic grouping of the American Azolla species. This correlation suggests that the cyanobacterial symbionts of Azolla spp. coevolved into distinct genetic groups with their hosts.

Oligosaccharides derived from the xyloglucan isolated from the seeds of Hymenaea courbaril var. stilbocarpa

On aqueous extraction, Hymenaea courbaril var. stilbocarpa, known in Brazil as jatobá, furnishes a high yield of viscous xyloglucan (45%) from its seeds. The crude polysaccharide (B1) was hydrolysed and the products, analysed as alditol acetates, were glucose, xylose, galactose and arabinose in the ratio 50:35:13:2. After further fractionation on a DEAE-cellulose column (chloride form), the main fraction (70% yield, B2) was obtained. The basic structure of the xyloglucan was determined as a cellulose-type (1 → 4)-linked β- d-glucan backbone partially substituted with side chains at 06 of α- d-xylopyranose, some of which were themselves substituted at 02 by the units of β- d-galactopyranose. Treatment of the xyloglucan (B2) with commercial cellulase from Trichoderma sp. yielded six oligosaccharides. These oligosaccharides were isolated by preparative paper chromatography, and their structures were determined by gas-liquid chromatography-mass spectroscopy of the derived partially O-methylated alditol acetates. These results confirm the structure proposed for jatobá seed xyloglucan.

Differentiation of free-living Anabaena and Nostoc cyanobacteria on the basis of fatty acid composition.

The cellular fatty acids of free-living, nitrogen-fixing cyanobacteria belonging to the genera Anabaena and Nostoc were analyzed to differentiate the genera. The fatty acid compositions of 10 Anabaena strains and 10 Nostoc strains that were grown for 12 days on BG-11o medium were determined by gas-liquid chromatography-mass spectroscopy. Of the 53 fatty acids detected, 17 were major components; the average level for each of these 17 fatty acids was at least 0.9% of the total fatty acids (in at least one of the genera). These fatty acids included (with mean percentages in the Anabaena and Nostoc strains, respectively) the saturated fatty acids 16:0 (30.55 and 23.23%) and 18:0 (0.77 and 1.27%); several unsaturated fatty acids, including 14:1 cis-7 (2.50 and 0.11%), 14:1 cis-9 (3.10 and 3.41%), a polyunsaturated 16-carbon (sites undetermined) fatty acid with an equivalent chain length of 15.30 (1.20 and 1.03%), 16:4 cis-4 (0.95 and 0.87%), 16:3 cis-6 (2.16 and 1.51%), 16:1 cis-7 (1.44 and 0.36%), 16:1 cis-9 (6.53 and 18.76%), 16:1 trans-9 (4.02 and 1.35%), 16:1 cis-11 (1.62 and 0.42%), 18:2 cis-9 (10.16 and 12.44%), 18:3 cis-9 (18.19 and 17.25%), 18:1 cis-9 (4.01 and 5.10%), and 18:1 trans-9 (0.92 and 1.94%); and the branched-chain fatty acids iso-16:0 (2.50 and 1.14%) and iso-15:1 (0.34 and 2.05%).(ABSTRACT TRUNCATED AT 250 WORDS)

Galactoxylomannans of Cryptococcus neoformans

Galactoxylomannans (GalXMs) from single isolates of Cryptococcus neoformans serotypes A, B, and D were isolated from culture supernatants and then purified by affinity, ion-exchange, and gel-filtration chromatography. GalXMs are a group of closely related complex polysaccharides. GalXMs from serotypes A (9759 A) and C (3183 C) and an acapsular mutant of serotype D (Cap67 D) have similar galactose, xylose, and mannose molar ratios, but each has some unique structural features. GalXM9759 A and GalXM 3183 C were associated with a starchlike glucan that was removed during purification. Only a trace of glucose was detected in the Cap67 D GalXM. Gas-liquid chromatography-mass spectroscopy of per-O-methylated polysaccharides and 13C nuclear magnetic resonance spectroscopy showed that GalXM is a complex branched polysaccharide. The main chain consists of mannose or galactose or alternating mannose and galactose residues. Xylose is present only as nonreducing termini. Galactofuranose occurs only in 3183 C and Cap67 D, and it is always present as nonreducing termini.

Gas-Liquid Chromatography-Mass Spectroscopy Determination of Clopamide in Plasma

A gas-liquid chromatography-mass spectroscopy(GLC-MS) method for the determination of clopamide (1) in human plasma was developed to evaluate the pharmacokinetics and bioavailability of 1 in humans. The method is specific, sensitive, and rapid and allows routine analysis as required for extensive pharmacokinetic studies. The assay procedure involves addition of furosemide (2) as an internal standard to plasma, separation on Sep Pack-C18 cartridges, elution by ether: methanol (1:1), evaporation, and subsequent derivatization with trime-thylanilinlum hydroxide in methanol (Methelute). Then, 5 μL of the reaction mixture is injected into a GLC-MS system which consists of a 1% SE-30 on Gas Chrom Q (100–120 mesh) glass column. The MS information was obtained under the following conditions: ionization beam energy 70 eV, ion source 200°; and m/e 372 for single ion monitoring. The retention times for 1 and 2 were 0.6 and 1.0 min, respectively. The limit of detection is 10 ng/mL of 1 in plasma and the calibration curve was shown to be linear between 10 and 500 ng/mL of 1. After repeated analysis of spiked plasma samples, the coefficient of variation ranged from 5.1 to 8.3%. The recovery from the extraction procedure was 92 ± 2.2%. Spiked samples frozen at —20°C were stable for at least 8 weeks. The method has been successfully used in a pharmacokinetic study with po dosing of 5mg of 1 to eight healthy volunteers. Peak plasma concentrations of 197 ± 56 ng/mL were observed after 1.1 ± 0.34h. No measurable concentration of 1 beyond 12h after dosing was observed.

Purified native haptens of Brucella abortus B19 and B. melitensis 16M reveal the lipopolysaccharide origin of the antigens

Purification of the Brucella polysaccharide referred to as native hapten (NH) and extracted from cells by the autoclaving procedure, was accomplished by ultrafiltration, followed by repetitive gel filtration using high-performance liquid chromatography on a ⪡TSK-G2000-SW⪢ column. The purified NH was analysed by SDS-PAGE, gas-liquid chromatography mass spectroscopy, and 13 C and 1 H NMR spectroscopy. NH from B. abortus B19 (NH-A) was shown to have a structure identical to that of A polysaccharide from B. abortus 1119-3, a linear homopolymer of α-1,2-linked-4,6-dideoxy-4-formamido-D-mannopyrannosyl residues. The structure of the NH from B. melitensis 16M (NH-M) was identified as a linear homopolysaccharide of the same sugar but composed of a pentasaccharide repeating unit in which four α1,2-linked-4,6-dideoxy-4-formamido-D-mannopyrannosyl residues are linked α-1,3 to the last monosaccharide of the sequence. This structure is similar to that determined for the Brucella M polysaccharide from B. melitensis 16M. The discovery in highly purified NH preparations of covalently bound monosaccharides characteristics of lipopolysaccharide inner core regions e.g. , quinovosamine, mannose and 3-deoxy-D-manno-octulosonate (KDO), indicates that this polysaccharide is derived from lipopolysaccharides (LPS) by hydrolytic conditions fortuitously generated during the extraction protocol. The antigenically important polysaccharides of Brucelle are now established to be either A or M antigens. Polysaccharide B is a cyclic glucan with no structural or serological relationship were to A for M polysaccharides, its apparent activity in diagnostic tests of infected cattle resuls from O polysaccharide B.

Purified native haptens of Brucella abortus B19 and B. melitensis 16M reveal the lipopolysaccharide origin of the antigens

Purification of the Brucella polysaccharide referred to as native hapten (NH) and extracted from cells by the autoclaving procedure, was accomplished by ultrafiltration, followed by repetitive gel filtration using high-performance liquid chromatography on a ⪡TSK-G2000-SW⪢ column. The purified NH was analysed by SDS-PAGE, gas-liquid chromatography mass spectroscopy, and 13C and 1H NMR spectroscopy. NH from B. abortus B19 (NH-A) was shown to have a structure identical to that of A polysaccharide from B. abortus 1119-3, a linear homopolymer of α-1,2-linked-4,6-dideoxy-4-formamido-D-mannopyrannosyl residues. The structure of the NH from B. melitensis 16M (NH-M) was identified as a linear homopolysaccharide of the same sugar but composed of a pentasaccharide repeating unit in which four α1,2-linked-4,6-dideoxy-4-formamido-D-mannopyrannosyl residues are linked α-1,3 to the last monosaccharide of the sequence. This structure is similar to that determined for the Brucella M polysaccharide from B. melitensis 16M. The discovery in highly purified NH preparations of covalently bound monosaccharides characteristics of lipopolysaccharide inner core regions e.g., quinovosamine, mannose and 3-deoxy-D-manno-octulosonate (KDO), indicates that this polysaccharide is derived from lipopolysaccharides (LPS) by hydrolytic conditions fortuitously generated during the extraction protocol. The antigenically important polysaccharides of Brucelle are now established to be either A or M antigens. Polysaccharide B is a cyclic glucan with no structural or serological relationship were to A for M polysaccharides, its apparent activity in diagnostic tests of infected cattle resuls from O polysaccharide B.

Chemical characterization and immunogenicity of capsular polysaccharide isolated from mucoid Staphylococcus aureus.

In this study we report the isolation and purification of the capsular polysaccharide elaborated by Staphylococcus aureus SA1 mucoid. The capsule was isolated from bacterial extracts and culture supernatants by a series of ethanol precipitations and enzyme digestions, followed by ion-exchange chromatography. Teichoic acid contamination was eliminated by oxidation with sodium metaperiodate, and the final product eluted in the void volume of a Sephacryl S-300 column. The purified capsular polysaccharide was analyzed by gas-liquid chromatography-mass spectroscopy, 13C and 1H nuclear magnetic resonance, amino acid analysis, immunelectrophoresis, and numerous biochemical assays. The major constituents of the capsule were 2-acetamido-2-deoxy-alpha-galacturonic acid (4-O linked), 2-acetamido-2-deoxy-alpha-fucose (3-O linked), and taurine. The polysaccharide also contained O-acetyl groups which were removed by mild alkaline hydrolysis. Serologically and biochemically, the capsule from strain SA1 mucoid appeared very similar to that produced by strain M. Purified capsular polysaccharide was immunogenic in both rabbits and mice. The optimal immunizing dose in mice was 0.1 microgram of purified capsular polysaccharide administered intraperitoneally. SA1 mucoid resisted opsonophagocytic killing by human leukocytes and complement. However, antibodies raised to the purified capsular polysaccharide neutralized the antiphagocytic effect of the capsule.

Formation of lipoxin A by granulocytes from eosinophilic donors

The formation of arachidonic acid-derived lipoxygenase products was examined with human granulocytes obtained from eosinophilic donors. These eosinophil-enriched leukocyte populations, challenged in vitro with the ionophore of divalent cations A23187, transformed both exogenous and endogenous sources of arachidonic acid to several lipoxygenase-derived products, including 5( S),6( R),15( S)-trihydroxy-7,9, 13- trans-11- cis-eicosatetraenoic acid (lipoxin A). Lipoxin A was detected and characterized by high-pressure liquid chromatography (HPLC), ultraviolet absorbance, and gas-liquid chromatography-mass spectroscopy. Neither lipoxin B nor 6( S)-LXA was consistently detected in extracts from these incubations. The amounts of lipoxin A formed were proportional to the percentage of eosinophils present in the suspension. The results indicate that granulocytes from eosinophilic donors can generate lipoxin A.

Stimulatory effect of certain plant sphingolipids on fruiting of Schizophyllum commune.

We reported previously that certain cerebrosides and ceramides from fungi were active upon fruiting of Schizophyllum commune (Kawai, G., and Ikeda, Y. (1982) Biochim. Biophys. Acta 719, 612-618; Kawai, G., and Ikeda, Y. (1983) (Biochim. Biophys. Acta 754, 243-248). This work was undertaken to extend our study to sphingolipids in wheat grain. The cerebrosides from wheat grain were fractionated by high-performance liquid chromatography into at least 40 components with and without the fruiting-inducing activity. Four major active fractions were characterized by thin-layer chromatography, infrared spectroscopy, gas-liquid chromatography, gas-liquid chromatography-mass spectroscopy, and 1H and 13C nuclear magnetic resonance spectroscopy. The active cerebrosides consist of glucose, 2-hydroxyhexadecanoic acid or 2-hydroxyoctadecanoic acid acid, and (4E,8Z)-sphingadienine or (8Z)-sphingenine. The cerebroside with (8Z)-sphingenine became inactive when the double bond was hydrogenated. Diglycosylceramides were as active as the monoglycosylceramides, but triglycosylceramides were only about 10% as active. The relationship between the structure and the activity is discussed.

Measurement of Δ9-Tetrahydrocannabinol (THC) in Whole Blood Samples from Impaired Motorists

The major psychoactive cannabinoid in marihuana, delta 9-tetrahydrocannabinol (THC) was measured in 1792 randomly selected blood specimens from erratic motorists arrested for impairment who submitted to blood alcohol sampling. Of these specimens, 14.4% were positive for THC (greater than or equal to 5.5 ng/mL). In those erratic driver specimens negative for alcohol THC positives rose to 23%. Drivers who used marihuana covered a broad age range. Aliquots of hemolyzed blood (10 microL) were analyzed by a sensitive radioimmunoassay (RIA) not requiring extraction. RIA accuracy and specificity were validated by gas liquid chromatography/mass spectroscopy (GLC/MS) split pair analysis (correlation coefficient = 0.93). This initial experience should facilitate and amplify a program designed to set forth the epidemiology of marihuana use in motorists and possible behavioral correlates.