The system of sulfated galactans from the red seaweed Gymnogongrus torulosus (Phyllophoraceae, Rhodophyta): Location and structural analysis

Sulfated polysaccharides were localized in the cuticle, cortex and medulla of the gametophyte thallus, being more concentrated in the intercellular matrix than in the cell walls. During the water extraction sequence, a small percentage of galactan sulfates (5.1% of dry seaweed) with average low M_r (6–11.4 kDa) were extracted at room temperature without disturbing the cellular arrangement, while sulfated galactans of average medium M_r (18–45 kDa) were obtained by further hot-water extractions (52.4% of dry seaweed), with diorganization of the tissue. The residue (40.0% of dry seaweed) still contained carrageenan-type (major) and agaran-type (minor) galactans. Part of these galactans was extracted with 8.4% LiCl solution in DMSO, from which “pure” κ/ι-carrageenans were isolated.Carrageenans and agarans were extracted in a ratio 1:0.5, showing the highest amount of agaran-structures for a carrageenophyte. The galactans comprise alternating 4-sulfated (major) and non-sulfated (minor) 3-linked β-d-galactopyranose units, and 4-linked α-galactopyranose units with the following substitutions: (i) non-sulfated and 2-sulfated 3,6-anhydro-α-d-galactopyranose residues in the carrageenan-structures, which belong to the κ-family (κ/ι-carrageenans); (ii) 3-sulfated α-l-galactopyranose units and 2-sulfated 3,6-anhydro-α-l-galactopyranose residues in the agaran-structures.Alkaline treatment and alkaline dialysis of the main extracts gave “pure” κ/ι-carrageenans, showing that carrageenan molecules are extracted together with low M_r agarans or agaran-dl-hybrids.     

Proximate composition and biological activity of food plants gathered by chilean Amerindians

The proximate composition and biological activity of food plants and mushrooms gathered by Chilean Amerindians were assessed. The gathered plants served primarily as sources of carbohydrates with highest values for Dioscorea tubers, Prosopis alba pods meal and Bromus catharticus seeds. The mushrooms Clavaria coralloides and Boletus loyus proved to be the best protein sources in our survey, but deficient in the amino acids methionine and cysteine. Some extracts of the plants and mushrooms under study showed biological activity as free radical scavengers, enzyme inhibitors, hypotensive or DNA binding effect. Free radical scavenging activity was detected in Cryptocarya alba fruit extract, while Typha angustifolia showed a strong DNA binding effect at 0.50 mg/ml. Methanolic extracts of the Apiaceae species Sanicula graveolens and Apium australe were moderately active as β-glucuronidase inhibitors at 50 fig/ml.     

neo-Clerodane diterpenoids from Ajuga: structural elucidation and biological activity

The presence of several types of allelochemicals has been reported from Ajuga, a Labiatae genus comprising more than 40 species of wide distribution in extratropical regions of both hemispheres. The genus is of great medicinal and economic importance and among the biological properties of the secondary metabolites, the antifeedant activity against pest insects appears to be related to the presence of neo-clerodane type diterpenes. This review focuses on the isolation and structural elucidation of this type of compounds from Ajuga species and the hemisynthetic compounds of closely related structure obtained. The reported biological activity of crude extracts and isolated diterpenes will be briefly commented.

Short-term changes in activity rhythms in an intertidal arthropod (Acarina: Bdella interrupta Evans)

Summary On a marine saltmarsh, the intertidal mite, Bdella interrupta Evans, is exposed to periods of regular tidal submergence which alternate with periods of tidal emergence. The mite shows well defined day and night peaks of locomotory activity on the soil surface. During periods of tidal submergence the activity peaks showed an apparent periodicity, of around 12.5 h, characteristic of a tidal rhythm. In the presence of non-submerging tides the periodicity of the activity peaks changed to around 11.5 h. It is suggested that this shorter periodicity results either from free-running of the circatidal clock in the absence of entraining tides or from a sequence of transients that are phaseshifting towards a stable relationship with a second zeitgeber. It is further suggested that the alternation between a ca. 12.5 h and a ca. 11.5 h rhythm ensures that an activity peak is maintained in daylight and, also, reduces the possibility of inundation during the first critical floodings during sequences of rising spring tides.     

Characterization of nativeFrankia strains isolated from chilean shrubs (Rhamnaceae)

Nine nativeFrankia strains were isolated from root nodules of four chilean actinorhizal plants (Rhamnaceae). The strains were designated as ChI1, ChI2, ChI3 and ChI4 fromColletia hystrix; ReI4 and ReI6 fromRetanilla ephedra; TqI12 and TqI15 fromTalguenea quinquinervis and TtI42 fromTrevoa trinervis. By scanning electron microscopy, all the strains exhibited similar actinomycetal structures: hyphae, sporangia and vesicles. The growth patterns of the isolates in BAP medium were similar. All showed a lag phase of approximately 6–7 days, then exhibited a logarithmic phase, except the ReI4 strain which seems to follow a linear growth pattern. A common feature of all the strains was a rapid loss of biomass at the end of the growth phase. All native strains grew on BAP medium supplemented with glucose. In six out of nine strains, the glucose was the best of the carbon sources tested. However, the strains differed in their ability to use other carbon sources such as arabinose, mannitol, maltose, succinate, sucrose, pyruvate, propionate and galactose. The isolates were sensitive to six antibiotics assayed (ampicillin, penicillin G, rifampicin, chloramphenicol, erythromycin and kanamycin). Using the acetylene reduction assay, the nitrogenase activity of the strains was determined. All strains grown in BAP medium lacking a combined nitrogen source were able to reduce acetylene ‘in vitro’.     

Summer biomass of a population of Phyllophora antarctica (Phyllophoraceae,Rhodophyta) from Antarctica

Results of a study on summer biomass in aninfralittoral population of Phyllophora antarctica A.et E. S. Gepp from Terra Nova Bay (Ross Sea,Antarctica) are reported. The population studied grewat depths of 5 to 12 m. The highest value of biomass(1548 wet g m⁻²) was found at the end of January at6 m depth. Data showed that biomass depended mainly onthe presence or absence of large heavy specimens, eventhough these were always few in number. Moreover, thevery high number of specimens in weight classes 1 (upto 0.03 wet g) and 2 (> 0.03 to 0.06 wet g) recordedfrom December to January showed a relevant productionof new thalli in that period; the decrease of thetotal number of thalli from 8832 counted on 30December to 4384 counted on 10 February, showed someself-thinning in the population; the occurrence ofnumerous thalli belonging to weight classes 3 (>0.06 to 0.125 wet g), 4 (> 0.125 to 0.25 wet g) and5 (> 0.25 to 0.5 wet g) just at the end of December,allows to conclude that P. antarctica continues togrow, although at low rate, during the period ofwinter darkness. Finally, both the low number ofthalli of weight classes 6 (> 0.5 to 1 wet g) and 7(> 1 to 2 wet g) and the absence of thalli of theweight class 8 (> 2 wet g), recorded at the end ofDecember, indicate that most of specimens reachingduring the winter such weight classes (due to theirability to dark growth), died before summer. This revised version was published online in July 2006 with corrections to the Cover Date.     

Mallomonas corymbosa var. interrupta nov. var. (Synurophyceae) from Argentina

A new variety of Mallomonas corymbosa is described from a pond in Nahuel-Huapi National Park (Argentina): M. corymbosa var. interrupt nov. var. Fine structure of scales and bristles has been studied by scanning and transmission electron microscopy.     

Pollen analysis of honeys of Melipona (Michmelia) seminigra merrillae and Melipona (Melikerria) interrupta (Hymenoptera: Apidae) bred in Central Amazon,Brazil

The pollen present in honey in colonies of Melipona seminigra merrillae and Melipona interrupta bred in Manaus was analysed. Between August and October 2012, honey samples were collected from the Sucupira meliponary, located in Manaus, Amazonas, Brazil. We identified a total of 70 pollen types belonging to 35 botanical families. In the samples from Melipona seminigra merrillae, the most represented pollen types were: Miconia-type (Melastomataceae) was the dominant pollen (DP) in August (51.19%), September (76.83%) and October (58.33%); Triplaris-type (Polygonaceae) was an accessory pollen (AP) in August (20.5%); and the remaining pollen types were classified as isolated pollen (IP), with Talisia macrophylla (Sapindaceae) exhibiting the highest percentage in August (12.92%). For Melipona interrupta, the most frequent pollen types were as follows: Miconia-type was the DP in August (59.33%) and October (61.33%) and an AP in September (37.5%); and Triplaris-type was an AP in August (35.83%), September (34.16%) and October (23.33%). The diversity of pollen types was not significantly different between the bees in the months evaluated. However, there was a large significant niche overlap in the months studied, August (O_ik = 0.95), October (O_ik = 0.89) and September (O_ik = 0.69), revealing that of the 70 pollen types found in the samples, 22 were shared by the two bee species in large proportions, 28 were exploited exclusively by Melipona seminigra merrillae and 20 were collected only by Melipona interrupta.     

Poliovirus RNA-dependent RNA polymerase (3Dpol): structural, biochemical, and biological analysis of conserved structural motifs A and B

We have constructed a structural model for poliovirus RNA-dependent RNA polymerase (3D(pol)) in complex with a primer-template (sym/sub) and ATP. Residues found in conserved structural motifs A (Asp-238) and B (Asn-297) are involved in nucleotide selection. Asp-238 appears to couple binding of nucleotides with the correct sugar configuration to catalytic efficiency at the active site of the enzyme. Asn-297 is involved in selection of ribonucleoside triphosphates over 2'-dNTPs, a role mediated most likely via a hydrogen bond between the side chain of this residue and the 2'-OH of the ribonucleoside triphosphate. Substitutions at position 238 or 297 of 3D(pol) produced derivatives exhibiting a range of catalytic efficiencies when assayed in vitro for poly(rU) polymerase activity or sym/sub elongation activity. A direct correlation existed between activity on sym/sub and biological phenotypes; a 2.5-fold reduction in polymerase elongation rate produced virus with a temperature-sensitive growth phenotype. These data permit us to propose a detailed, structural model for nucleotide selection by 3D(pol), confirm the biological relevance of the sym/sub system, and provide additional evidence for kinetic coupling between RNA synthesis and subsequent steps in the virus life cycle.     

Ethanol analysis from biological samples by dual rail robotic autosampler

Detection, identification, and quantitation of ethanol and other low molecular weight volatile compounds in liquid matrices by headspace gas chromatography-flame ionization detection (HS-GC-FID) and headspace gas chromatography-mass spectrometry (HS-GC-MS) are becoming commonly used practices in forensic laboratories. Although it is one of the most frequently utilized procedures, sample preparation is usually done manually. Implementing the use of a dual-rail, programmable autosampler can minimize many of the manual steps in sample preparation. The autosampler is configured so that one rail is used for sample preparation and the other rail is used as a traditional autosampler for sample introduction into the gas chromatograph inlet. The sample preparation rail draws up and sequentially adds a saturated sodium chloride solution and internal standard (0.08%, w/v acetonitrile) to a headspace vial containing a biological sample, a calibrator, or a control. Then, the analytical rail moves the sample to the agitator for incubation, followed by sampling of the headspace for analysis. Using DB-624 capillary columns, the method was validated on a GC-FID and confirmed with a GC-MS. The analytes (ethanol, acetonitrile) and possible interferences (acetaldehyde, methanol, pentane, diethyl ether, acetone, isopropanol, methylene chloride, n-propanol, and isovaleraldehyde) were baseline resolved for both the GC-FID and GC-MS methods. This method demonstrated acceptable linearity from 0 to 1500 mg/dL. The lower limit of quantitation (LOQ) was determined to be 17 mg/dL and the limit of detection was 5 mg/dL.     

Instrumental neutron activation analysis of biological samples

The elemental compositions of 18 biological reference materials have been processed, for 14 stepped combinations of irradiation/decay/counting times, by the INAA Advance Prediction Computer Program. The 18 materials studied include 11 plant materials, 5 animal materials, and 2 other biological materials. Of these 18 materials, 14 are NBS Standard Reference Materials and four are IAEA reference materials. Overall, the results show that a mean of 52% of the input elements can be determined to a relative standard deviation of ±10% or better by reactor flux (thermal plus epithermal) INAA.

     

Isolation of lipids from biological samples

Abstract

Isolation of the lipid fraction from biological samples has been a crucial part of countless studies over the last century. This considerable research interest has led to the development of a number of methods for isolating a range of molecular species that fall under the umbrella term “lipid”. Such methods vary in popularity, complexity, specificity and even toxicity. In this review, we explore examples of published methods (1952–2014) for isolating lipids from biological samples and attempt to assess the limits of techniques both from a chemical and biological perspective. We also suggest how a suitable method might be chosen for a novel application.     

Patterns of reproductive isolation in nolana (chilean bellflower)

We examined reproductive isolating barriers at four postmating stages among 11 species from the morphologically diverse genus Nolana (Solanaceae). At least one stage was positively correlated with both genetic and geographic distance between species. Postzygotic isolation was generally stronger and faster evolving than postmating prezygotic isolation. In addition, there was no evidence for mechanical isolation, or for reproductive character displacement in floral traits that can influence pollinator isolation. In general, among the potential isolating stages examined here, postzygotic barriers appear to be more effective contributors to reducing gene flow, including between sympatric species.     

A novel fluorometric assay for quantitative analysis of dihydrofolate reductase activity in biological samples

In an effort to study the level of dihydrofolate reductase (DHFR), the main molecular target of antifolate drugs, in healthy and malignant tissues of human origin, a new and convenient fluorometric enzymatic assay has been developed. The technique measures the overall decrease in fluorescence emission at 454 nm (lambda ex = 342 nm) due to the contributions from coenzyme oxidation and substrate reduction. This technique was developed by using an enzyme purified from beef liver. All criteria of quality were checked: sensitivity, reproducibility and specificity made it suitable for low activity measurements. It was successfully applied to human tissue crude extracts.     

Extraction and purification of prostaglandins and thromboxane from biological samples for gas chromatographic analysis

An efficient extraction procedure for the isolation of prostaglandins (PGs) from biological samples for their subsequent quantification by gas chromatography-electron capture detection (GC-ECD) is described. PGs were extracted from lung, kidney, spleen and stomach fundus into ethyl acetate at different pHs. The highest recovery and least extraction of contaminating pigments was obtained at pH 4.5. Pigments and other contaminants are removed by thin-layer chromatography using a solvent system chloroform-isopropyl alcohol-ethanol-formic acid (45:5:0.5:0.3). The isolated PGs were determined by GC-ECD after appropriate derivatization. The overall recovery of PGs using this procedure is 60%.     

Ruthenium(III) maltolato-nitroimidazole complexes: synthesis and biological activity

The Ru(III) metronidazole-maltolato and -ethylmaltolato complexes, trans-[RuL₂(metro)₂]CF₃SO₃ (L = ma (1a) or etma (1b)), have been synthesized and tested for potential anti-tumour activity against the human breast cancer cell line MDA-MB-435S using a so-called MTT assay in phosphate-buffered saline; ma = 3-hydroxy-2-methylpyran-4-onato, etma = 2-ethyl-3-hydroxypyran-4-onato, metro = 2-methyl-5-nitro-1H-imidazole-1-ethanol (metronidazole); MTT = 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. The complexes exhibit lower IC₅₀ values than our previously reported Ru(III) tris-maltolato and -ethylmaltolato complexes [D.C. Kennedy, A. Wu, B.O. Patrick, B.R. James, Inorg. Chem. 44 (2005) 6529–6535]. An improved synthetic route to the 2-nitroimidazole EF5 (2-(2-nitro-1-H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide) is reported, as well as a related synthesis of a 3-nitro-1,2,4-triazole derivative of EF5, triF5 (2-(3-nitro-1-H-triazol-1-yl)-N-(2,2,3,3,3- pentafluoropropyl)acetamide). The complexes [RuL₂(EF5)₂]CF₃SO₃ (4a and 4b) and [Ru(ma)₂(triF5)₂]CF₃SO₃ (5) were prepared from the [RuL₂(EtOH)₂]CF₃SO₃ complexes (3a and 3b); IC₅₀ values for 3–5 are high. Data on the uptake of Ru by the cells are also reported. The complexes were characterized generally by all or some of the following methods: elemental analyses, NMR, IR and mass spectroscopies, conductivity, and cyclic voltammetry; complexes 1a and 1b were also analyzed by X-ray crystallography.