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Background: For hepatocellular carcinoma (HCC), effective therapeutic approaches are lacking. As aberrant gene methylation is a major contributor to HCC development, demethylating drugs such as 5-azacytidine (5-Aza) have been proposed. As most 5-Aza mechanisms of action are unknown, we investigated its phenotypic/molecular effects. Methods: 5-Aza effects were examined in the human HCC cell lines JHH-6/HuH-7 and in the rat cell-line N1-S1. We also employed a xenograft mouse model (HuH-7), a zebrafish model (JHH-6), and an orthotopic syngeneic rat model (N1-S1) of HCC. Results: 5-Aza downregulated cell viability/growth/migration/adhesion by upregulating miR-139-5p, which in turn downregulated ROCK2/cyclin D1/E2F1 and increased p27kip1, resulting in G1/G0 cell accumulation. Moreover, a decrease in cyclin B1 and an increase in p27kip1 led to G2/M accumulation. Finally, we observed a decrease in MMP-2 levels, a stimulator of HCC cell migration. Aza effects were confirmed in the mouse model; in the zebrafish model, we also demonstrated the downregulation of tumor neo-angiogenesis, and in the orthotopic rat model, we observed impaired N1-S1 grafting in a healthy liver. Conclusion: We demonstrate for the first time that 5-Aza can impair HCC development via upregulation of miR-139-5p, which in turn impairs the ROCK2/cyclin D1/E2F1/cyclin B1 pro-proliferative pathway and the ROCK2/MMP-2 pro-migratory pathway. Thus, we provide novel information about 5-Aza mechanisms of action and deepen the knowledge about the crosstalk among ROCK2/cyclin D1/E2F1/cyclin B1/p27kip1/MMP-2 in HCC.

PURPOSE To develop a novel approach to monitor lung ventilation/inflammation in cystic fibrosis (CF) patients. Lung assessment in CF patients is relevant given that most patients succumb to respiratory failure. Respiratory functional tests (forced expiratory volume in the first second; FEV1 ) and inflammatory markers are used to test pulmonary ventilation/inflammation, respectively. However, FEV1 is effort dependent and might be uncomfortable for CF patients. Furthermore, inflammatory marker detection is costly and not rapid. To overcome these limitations, we propose the measurement, by means of low field nuclear magnetic resonance, of the spin-spin relaxation time (T2m ) of water hydrogens present in CF patient sputum. In CF sputum, different biological components are pathologically increased and inversely related to lung functionality. Moreover, we showed that these components alter in a dose-dependent manner the T2m in synthetic CF sputum. METHODS Sputum samples were obtained from 42 CF subjects by voluntary expectoration; FEV1 , C-reactive protein (CRP), blood neutrophil counts together with cytokine (tumor necrosis factor alpha [TNFα], interleukin [IL]-1β, IL-4, and vascular endothelial growth factor) quantifications were then evaluated. RESULTS In sputum samples, we observe that T2m directly correlates (rFEV1 = 0.44; P < 10-4 ; 169 samples) with FEV1 . Moreover, T2m inversely correlates with the circulating inflammation markers CRP/neutrophil number (rCRP = -0.44, P < 10-4 ; rNC = -0.37, P < 2 * 10-4 ; 103 and 86 samples, respectively) and with the sputum inflammatory cytokines TNFα/IL-β1 (rTNFα = -0.72, P < 10-4 ; rIL-1β = -0.685, P < 10-4 ; 27 samples). T2m variations also correspond to FEV1 values over time in defined patients. CONCLUSION These findings, together with the fast, reliable, and simple determination of T2m , make our approach a novel tool potentially usable in the real world of CF patients.

International audience

Carbon nanotube (CNTs) derivatives are nowadays under thorough investigation as biomedically interesting materials. In this paper we describe a method for the preparation of water-soluble CNTs by condensation of the carboxylic groups introduced onto the carbon framework and the primary amine moieties inserted in the naturally occurring biopolymer hyaluronan (HA). The covalent conjugation between CNTs and HA should merge the biocompatibility and further processability of the HA chains with the well-known cellular penetration properties of the CNT derivatives to produce novel drug delivery platforms. In fact, thanks to the primary amino groups introduced in the HA chains, HA-CNT derivatives can be further covalently modified with model drugs like ibuprofen and methotrexate. We describe also the monitoring of all the CNT derivatization steps by diffusion-ordered NMR spectroscopy (DOSY), a technique that allows fast and reliable characterization of these novel derivatives.

The effects of feeding 1,3/1,6 β-glucans on the innate and the adaptive immune responses of European sea bass (Dicentrarchus labrax) was investigated. Two experiments were carried out during the study. In the first, a number of non-specific immune parameters were examined at 4, 7, 10, 14, 21 and 25 days of feeding fish with a semipurified diet containing Macrogard ©, a commercially available form of 1,3/1,6 β-glucans. The respiratory burst activity of head kidney macrophages isolated from the different groups of fish fed the immunostimulant peaked and subsequently decreased at different times during the experiment. Head kidney macrophages from fish fed 250 ppm β-glucans had a statistically higher level of respiratory burst activity at Day 21 of feeding compared with fish fed no immunostimulant. No statistical differences were observed in lyzozyme activity during this trial. In the second experiment, the effect of feeding 1,3/1,6 β-glucans on the immune response of fish to an alginate-encapsulated Vibrio vaccine administered orally was examined. Respiratory burst of head kidney macrophages and serum lysozyme activity decreased in all fish over the course of the trial, while serum lysozyme activity was considerably lower than values obtained in the first experiment. Fish vaccinated orally had significant increases in antibody response by Week 2 post-vaccination, but β-glucans did not appear to affect these levels. Vaccination may have resulted in activating the immune system as a whole, thus masking any difference in immunostimulation by the β-glucans. It may be that the optimal doses and timing of β-glucans administration is different when the immunostimulant is administered alone or in combination with the vaccine. In conclusion, European sea bass can be immunomodulated with oral administration of β-glucan. Optimal doses and administration times have been established when β-glucans are fed alone, although further studies are needed to establish ideal feeding regimes for glucans administrated in combination with vaccination. Questo studio è stato condotto per determinare l’effetto che i 1,3/1,6 β-glucani possono esercitare sul sistema immunitario specifico e aspecifico del branzino Dicentrarchus labrax. A questo scopo sono stati effettuati due esperimenti. Nel primo, alcuni parametri relativi al sistema immunitario aspecifico sono stati esaminati dopo 4, 7 10, 14, 21 e 25 giorni di alimentazione con diete semi-purificate contenenti il prodotto commerciale a base di β-glucani Macrogard®. L’attività relativa allo scoppio respiratorio dei macrofagi del rene craniale isolati dagli animali di tutti i gruppi sperimentali ha mostrato un picco di attività seguito da un decremento, ma con temporalità differenti a seconda del dosaggio. I macrofagi dei pesci alimentati con la dieta contenente 250 ppm di glucani hanno mostrato differenze statisticamente significative rispetto agli altri gruppi al giorno 21 di somministrazione. Nessuna differenza è stata riscontrata per quanto riguarda l’attività del lisozima. Nel secondo esperimento, gli effetti della somministrazione orale di β-glucani sono stati valutati quando somministrati insieme ad un vaccino orale contro la vibriosi. L’attività relativa allo scoppio respiratorio dei macrofagi e l’attività del lisozima sono calati lungo tutto il periodo di osservazione, con quest’ultimo valore considerevolmente più basso rispetto a quello ottenuto nel primo esperimento. I pesci vaccinati oralmente hanno mostrato un aumento della risposta anticorpale dopo 2 settimane dalla somministrazione di vaccino anche se i β-glucani non hanno mostrato effetti sul sistema immunitario specifico dei pesci. La vaccinazione, stimolando l’attività del sistema immunitario, può aver mascherato l’effetto dei β-glucani. La dose ottimale di β-glucani potrebbe essere differente quando questi vengono somministrati insieme ad un vaccino. In conclusione, il branzino sembra poter essere immunomodulato mediante la somministrazione oral dei β-glucani. È stato possibile stabilire la dose e il tempo di somministrazione ottimali quando i β-glucani vengono utilizzati singolarmen-te; al contrario, sono necessari ulteriori studi per stabilire la dose ottimale quando questi vengono somministrati insieme ad un vaccino.

A novel methodology for making drug conjugates using hyaluronan as a carrier was developed. This strategy involves a completely regioselective two-step synthesis of 6-amino-6-deoxyhyaluronan, which is then easily functionalized with drugs through a suitable linker. The case of hyaluronan–camptothecin conjugates is described, making use of a simple succinate linker. The antitumor activity of new hyaluronan derivatives prepared is at present under evaluation.

International audience; The study of the interactions of drugs with plasmatic proteins is of fundamental importance for the understanding of their biological applications and effects. The classical analytical techniques employed for the characterization of the degree of binding in such ligand/receptor systems are mainly based on separation method and spectroscopic characterizations. In the present study, we show that diffusion-based NMR spectroscopy is a useful tool for the fast and reliable determination of the quantitative binding between a polymer-drug bioconjugate and a model protein in aqueous solution. The dissociation constant (Kd) related to the binding event was calculated and compared with the results obtained from a classical fluorescence-based assessment, showing good match between these two methods.

Starch is the main source of carbohydrates in human diet. It is widely used in food processing and non-food industrial applications. The effects on starch digestion and absorption in humans are reviewed in relation to the starch composition, sources, plant genetic variation, food processing and cooking. The impact of food industrial processing and starch modification on the digestibility of starch containing foods and on gut microbiota are discussed. Considering that the resistant starch (RS) fraction escaped from the small intestine is fermented in large intestine, all the variables that influence starch digestibility and absorption must be taken into account when discussing about healthy properties of fibers. Future trends in food industries are aimed to increase the RS fraction in processed foods in order to improve nutritional quality as well as to clarify the influence of RS3 and RS4 on gut microbiota.

Magnetic resonance imaging (MRI) has already been successively used to investigate polysaccharide matrices. In particular, MRI at microscopic resolution (MR microscopy) is now one of the most powerful techniques for studying the physical properties of natural hydrogels. To contribute to a better understanding of the correlation between chemical and physical properties of agar gels, we report here the measurement of the water magnetic parameters for agar gels extracted from different species of Gelidium: T1 and T2 relaxation times, magnetisation transfer (Ms /M0) and diffusion (D) were measured to evaluate their use for studying the gel characteristics. MR microscopic images were acquired at 7.05 Tesla using various pulse sequences. The results obtained confirmed the possibility to use quantitative MRI for the characterisation of physical parameters correlated with the type of agar chemical structure. In particular, T2 data obtained for gels at different concentrations indicate that this magnetic parameter is very sensitive to the agar concentration and hence particularly useful for the gel strength determination.

Dried samples of the red algae Solieria filiformis and Agardhiella subulata from Mar Piccolo, Taranto, were extracted by stirring in water at 25 (Cold), 85 (Hot) and 120 (pressure) °C. For each species, six successive cold, three hot and two pressure extractions were performed and analyzed separately. Water-soluble polysaccharides were isolated after treatment with amylase, alcohol precipitation, dialysis and freeze drying. The total, starch-free yields were 27 and 24% of the dry weight for S. filiformis (SF) and A. subulata (AS), respectively. FTIR and NMR spectroscopic analysis showed that the different cold and hot water extracts had a similar structural backbone, which can be characteristic of gelling type carrageenans with a dominant iota repeating structure (DA2S-G4S). However, some irregularities were detected by NMR, which may be attributed to 6-sulphated 4-linked precursor units. The occurrence of such and other irregularities was more frequent in the SF samples, giving a possible explanation for the enhanced extractability in cold water of carrageenan polymers from this species.

Hyaluronan (hyaluronic acid) is a naturally occurring polysaccharide of a linear repeating disaccharide unit consisting of β-(1→4)-linked D-glucopyranuronic acid and β-(1→3)-linked 2-acetamido-2-deoxy-D-glucopyranose, which is present in extracellular matrices, the synovial fluid of joints, and scaffolding that comprises cartilage.In its mechanism of synthesis, its size, and its physico-chemical properties, hyaluronan is unique amongst other glycosaminoglycans. The network-forming, viscoelastic and its charge characteristics are important to many biochemical properties of living tissues. It is an important pericellular and cell surface constituent; its interaction with other macromolecules such as proteins, participates in regulating cell behavior during numerous morphogenic, restorative, and pathological processes in the body. The knowledge of HA in diseases such as various forms of cancers, arthritis and osteoporosis has led to new impetus in research and development in the preparation of biomaterials for surgical implants and drug conjugates for targeted delivery.A concise and focused review on hyaluronan is timely. This review will cover the following important aspects of hyaluronan: (i) biological functions and synthesis in nature; (ii) current industrial production and potential biosynthetic processes of hyaluronan; (iii) chemical modifications of hyaluronan leading to products of commercial significance; and (iv) and the global market position and manufacturers of hyaluronan.

Steam explosion was investigated as a pretreatment step in the isolation of agar from the macroalgaGracilaria dura. As compared to conventional procedures, the yield of agar obtained using this method on alkali (Na2CO3) conditioned algal material was higher. Extractions performed first at 95 °C and then at 121 °C showed that the major fraction of the agar was extracted at 95 °C, independently of the pretreatment. The efficiency of sulphate hydrolysis during steam explosion ofG. dura previously conditioned in Na2CO3, was similar to that of a NaOH based alkali pretreatment. Except for a lower nitrogen content of the sample obtained after NaOH based alkali pretreatment and a higher 6-O-methyl-β-d-galactose content in the agar after steam explosion, the chemical composition of the agars showed no significant difference. Agars extracted after steam explosion had melting temperature, gel strength and apparent modulus of elasticity lower than those of corresponding ‘native’ and alkali (NaOH) pretreated samples, but comparable to those of a commercial sample.

International audience; Functionalized carbon nanotube (CNT) derivatives are currently under thorough investigation in different biomedical investigations. In this field of research, the composition of sample either in terms of covalently attached or physisorbed moieties can greatly affect the observed results and hamper the comparison between different studies. Therefore, the availability of a fast and reliable analytical technique to assess both the type of interaction (covalent vs noncovalent) and the composition of CNT conjugates is of great importance. Here we describe that the two-dimensional diffusion-ordered (DOSY) NMR spectroscopy is extremely useful to discriminate between conjugated and unconjugated polyethylene glycol groups in samples obtained by condensation with oxidized single-walled carbon nanotubes (SWNTs). This fast and nondestructive technique allows us to follow the removal of unconjugated polyethylene glycol chains during the purification. In particular, DOSY analysis reveal that about 1/3 (wt %) of the polyethylene glycol used for the condensation remained physisorbed to functionalized SWNTs after dialysis. Complete elimination of physisorbed polyethylene glycol was achieved using diafiltration.

The FT-IR and FT-Raman spectral studies of the Methotrexate (MTX) were carried out. The equilibrium geometry, various bonding features and harmonic vibrational frequencies of MTX have been investigated with the help of B3LYP density functional theory (DFT) using 6-31G(d) as basis set. Detailed analysis of the vibrational spectra has been made with the aid of theoretically predicted vibrational frequencies. The vibrational analysis confirms the differently acting ring modes, steric repulsion, conjugation and back-donation. The energy and oscillator strength calculated by Time-Dependent Density Functional Theory (TD-DFT) results complement with the experimental findings. The calculated HOMO and LUMO energies show that charge transfer occur within the molecule. Good correlations between the experimental 1H and 13C NMR chemical shifts in DMSO solution and calculated GIAO shielding tensors were found.

International audience; The emerging applications of functionalized carbon nanotubes (CNTs) in various research domains necessitate the use of many different analytical techniques to confirm their structural modifications in a fast and reliable manner. Thus far, NMR spectroscopy has not been among the main tools for characterization of organically modified carbon nanostructures. (1)H analysis is limited because the signals in these derivatives are typically weak and broad, resulting in uncertainties of a few parts per million, and because of the strong interference of residual solvent signals. To overcome these limitations, we investigated the applicability of proton NMR spectroscopy based on gradient-edited diffusion pulse sequences (1D diffusion-ordered spectroscopy, DOSY) in the characterization of CNT derivatives. In general, diffusion NMR experiments allow the separation of NMR signals of different species present in a mixture, according to their own diffusion coefficients, merging spectroscopy information with size analysis. In the present study, a selected set of CNT derivatives was synthesized and analyzed using 1D DOSY experiments by applying strong magnetic field gradients (up to 42.6 G cm(-1)). Colorimetric tests (i.e., Kaiser test) and TGA analysis support the NMR findings, which are related to isolated and/or bundled short SWNTs, on the basis of TEM and AFM characterization. The overall results show that the diffusion-based NMR spectroscopy is a fast and promising approach for the characterization of covalently modified CNT derivatives.

Selective halogenation of hyaluronan and partial halogen substitution by methotrexate led to 6-chloro-6-deoxy-6- O -methotrexylhyaluronan, a potential antitumor drug. The remaining halogen could be further substituted by a second organic carboxylate, leading to mixed esters. 6- O -Acetyl-6- O -methotrexylhyaluronan and 6- O -butyryl-6- O -methotrexylhyaluronan were thus synthesized and characterized by NMR spectroscopy.

Polysaccharides are widely used as carriers in the field of drug delivery. We present a methodology to obtain water soluble drug-conjugates based on scleroglucan. Selective C-6 halogenation gives access to C-6 esters; conjugates between methotrexate and scleroglucan are described, potentially useful for antitumour therapy or in rheumatoid arthritis treatment.

In this study, alginate polymers are used to get homogeneous cylindrical or spherical gels. MRI techniques are employed to study homogeneity of these gels. Four dif- ferent alginates are used and, for each one, five different con- centrations for mechanical tests and three different concen- trations for release tests are studied. Mechanical tests are performed to get gels' linear viscoelasticity region and then to evaluate their crosslink density in relation to polymer con- centration. Afterwards, three model molecules (theophylline, vitamin B12, and myoglobin) are loaded within gels to study the release kinetics in water from both cylindrical and spher- ical gels. Diffusion coefficients calculated from these experi- ments are then used to estimate the polymeric network mesh wideness. This work shows how crosslink density increases with polymer concentration regardless of the alginate type considered. In addition, while vitamin B12 diffusion coeffi- cient is inversely proportional to crosslink density, myoglo- bin is too large to diffuse through the polymeric network, whatever the alginate type and polymer concentration. At the same time, theophylline is too small to be sensibly affected by increasing the polymeric network crosslink den- sity. Finally, MRI analysis and vitamin B12 diffusion coeffi- cient values prove that, structurally speaking, cylinders and spheres are similar and homogeneous. 2008 Wiley Periodi- cals, Inc. J Biomed Mater Res 87A: 808-818, 2008

Investigation of light conditions suitable for cultivation of Gelidium pulchellum (Turner) Kurtz was performed under controlled laboratory conditions at 20 °C and in the range of irradiance of 10–430 μmol photons m−2 s−1. Growth, measured as fresh weight increment, increased with irradiance up to 130 μmol m−2 s−1 and no significant photoinhibition was observed up to 430 μmol m−2 s−1. Maximum growth rate (10.0% day−1) was obtained at 130–240 μmol m−2 s−1 under continuous light and aeration. The effect of irradiance on agar yield and quality was assessed. Agar yield varied from 31 to 38.6% of the algal dry weight, and variation was not related to irradiance. However, the yield of agar molecules soluble at 95 °C increased with increasing irradiance. A similar trend was found for sulphate content in both series o f extracts, at 95 and 121 °C. On the contrary, the molecular weight and the degree of methylation of agar molecules in the 95 °C extracts decreased with increasing light intensity. As a consequence of the variations in sulphate content, molecular weight and molecular weight distribution, the gel strength was considerably lower at high light intensity. Starch content varied from 0.9 to 7.7% of the algal dry weight, and apparently was not related with irradiance.

The phenotypic and agarolytic features of an unidentified marine bacteria that was isolated from the southern Pacific coast was investigated. The strain was gram negative, obligately aerobic, and polarly flagellated. On the basis of several phenotypic characters and a phylogenetic analysis of the genes coding for the 16S rRNA, this strain was identified as Pseudoalteromonas antarctica strain N-1. In solid agar, this isolate produced a diffusible agarase that caused agar softening around the colonies. An extracellular agarase was purified by ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography on DEAE-cellulose. The purified protein was determined to be homogeneous on the basis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and it had a molecular mass of 33 kDa. The enzyme hydrolyzed the β-1,4-glycosydic linkages of agar, yielding neoagarotetraose and neoagarohexaose as the main products, and exhibited maximal activity at pH 7. The enzyme was stable at temperatures up to 30°C, and its activity was not affected by salt concentrations up to 0.5 M NaCl.

The conformation of agarose in dimethyl sulfoxide (Me 2 SO) or in aqueous solutions has been studied by 1 H and 13 C NMR spectroscopy. Partial assignment of the 1 H NMR spectrum of agarose in Me 2 SO- d 6 has been obtained using 1D and 2D techniques. Coupling constant, chemical shift temperature dependence and exchange rate data for the hydroxyl protons have been used to establish the presence of intramolecular hydrogen bonds. The reported data show that the protons of HO-2 in the 3,6- anhydro-α- l -Gal p residue (A) and HO-4 in the β- d -Gal p residue (G) have a less rotational freedom and a higher thermal stability than the other hydroxyl protons. On the basis of the exchange rate data, obtained from 1D NOESY experiments with selective excitation of the residual water signal, an inter-residue hydrogen bond is proposed only for G HO-4 in Me 2 SO, while the involvement of A HO-2 in the agarose-Me 2 SO complex, recently reported, is suggested. The latter hypothesis is supported by the 13 C NMR data obtained in Me 2 SO- d 6 , in a Me 2 SO- d 6 -D 2 O mixture and in D 2 O, which also show that a relatively small conformational rearrangement of the chain segments occurs in aqueous solution. Deuterium isotope effect data indicate the persistence in water of the inter-residue G HO-4 mediated hydrogen bond with the additional involvement of G HO-6. In addition, the negligible isotope effect on the C-2 atom of A would indicate that the corresponding OH group, although free from the agarose-Me 2 SO complex, is not available for further strong interactions. The hydrogen bonding and conformation of agarose in methyl sulfoxide or in aqueous solutions were studied by NMR spectroscopy. Based on 1 H NMR data, an inter-residue H-bond is proposed for HO-4 of β- d -Gal p in Me 2 SO, while the involvement of HO-2 of 3,6- anhydro -α- l -Gal p in an agarose-Me 2 SO complex is suggested. These hypotheses are supported by the 13 C NMR data.

The red seaweeds Gelidiella acerosa and Gracilaria mammillaris growing along the coast of Venezuela were investigated as potential economic sources of agar. Agar extracted from Gracilaria mammillaris accounted for 27% of the algal dry weight but had quite a poor gelling ability because of the presence of alkali-stable sulphate groups on the D-galactose residue. However, the gel strength of its aqueous solutions was considerably enhanced by the addition of potassium, sodium and calcium ions (up to 1N). On the contrary, the galactan from Gelidiella acerosa (yield of about 20% w/w) exhibited quite good properties. In particular, the gel strength was comparable to that of commercial agaroses. Sulphate esters were not detectable by chemical methods and NMR spectroscopy revealed an agarose backbone with a high degree of methylation on both D and L-galactose residues.

Agar polymers synthesized by species of the genus Gracilaria constitute a complex mixture of molecules, containing several extremes in structure. Sulphate hemi-esters, methyl ethers and pyruvic ketals can alter in a number of ways the structural regularity of agar based on strictly 3-O-linked β-l-galactopyranose and 4-O-linked α-l-galactopyranose residues. In comparison with agars from Gelidium and Pterocladia, agars from Gracilaria can have higher degrees of sulphation, methoxylation and pyruvylation. The gelling ability of agars from most of Gracilaria species is considerably improved by adopting, before extraction, an alkali pretreatment which converts α-l-galactose 6 sulphate into 3,6-anhydro-α-l-galactose. Native agars obtained from Gracilaria cannot be classified, with few exceptions, as bacteriological grade agar as they have a high content of methoxyls and consequently high gelling temperatures. On the contrary, the genus Gracilaria is considered the most important source of food and sugar-reactive grade agars. Among techniques which can be used to study algal polysaccharides, combined 1H and 13C nuclear magnetic resonance spectroscopy represent the most effective and powerful method for the investigation of the chemical structure of agarocolloids.

One- and two-dimensional1H- and13C-NMR spectra of lipid extracts fromUlva rigida, Gracilaria longa, Fucus virsoides andCodium tomentosum collected in the northern Adriatic Sea allowed screening of the content of fatty acid chains, carotenoids, free and acylated cholesterol and chlorophylls. The carotenoid-to-polyunsaturated fatty acid molar ratio was taken as a comparison parameter in samples ofUlva rigida collected in differentloci and seasons; the value was markedly higher in samples from the Lagoon of Venice than from marine coastal waters. The total cholesterol concentration was evaluated by1H-NMR spectroscopy and similar values were found for all species. Two-dimensional heterocorrelated NMR spectroscopy was shown to give characteristic fingerprints of the lipid extracts from algal samples as regards the content in chlorophylls, unsaturated fatty acids and carotenoids.

Lipid extracts of the red algaGracilaria longa were studied by1H- and13C-NMR spectroscopy. Peaks in the13C-NMR spectra attributable to sterols, chlorophylls and carotenoids allowed free and acylated cholesterol, chlorophylla and lutein to be identified as the most abundant components of these classes. A content of 0.5 ± 0.1 μmoles of total cholesterol/g wet alga was estimated from the1H-NMR spectrum, which also allowed the determination of the phosphatidylcholine/total lipid molar ratio (9.5 ± 0.5%). The13C-NMR spectroscopic experiments provided information on the position of the double bonds on the fatty acid residues. A comparison between NMR spectra of lipid extracts obtained for wet and dried alga showed that the alga undergoes both a dramatic peroxidation and some glycolipid degradation during the drying process.

This paper presents optical and chirooptical data on the interaction of the microscopy-staining agent ruthenium red with carboxylated polysaccharides in dilute aqueous solution. The polysaccharides used are both natural (alginate and pectate) and semisynthetic (C6-oxidized cellulose and C6-oxidized amylose). A preliminary discussion of the molecular structure and conformational features which control the interaction is presented.

The evaluation of biopolymers of biotechnological interest has stressed the importance of renewable sources from the northern Adriatic Sea. In the frame of this research program, the red alga Gracilaria cf. dura, has been studied. Agar was obtained from Gracilaria cf. dura by means of alkali treatment and hot water extraction. In order to purify the extracts, amylase and isopropyl alcohol were used. Analysis of sugars was carried out by means of gas chromatography on alditol acetate derivatives; sulfate content was evaluated by means of infrared spectroscopy. An analysis of molecular weight distribution was carried out by means of high performance gel permeation chromatography coupled with a low angle laser light scattering device as detector. Analytical data were examined in terms of specificity of the algal source utilized, and the agar fractions extracted by means of this procedure were compared with a commercial sample.

Alginates, both commercial and laboratory made, are strongly fluorescent due to small amounts of polyphenolic materials. These contaminants can be detected by fluorescence spectroscopy in concentrations lower than 1 ppm. This technique has been used to measure polyphenols in a wide range of alginates and various procedures for preparation of biotechnological-grade alginates have been evaluated.

Agar polymers were extracted with and without alkali pre-treatment from the red seaweed Gracilaria dura , growing in the northern Adriatic Sea. Chemical analysis combined with 1 H and 13 C NMR spectroscopy showed that agar polysaccharides characterised by low methoxyl and fairly high pyruvate contents were obtained. Alkali pre-treatment markedly reduced the molecular weight and significantly increased the polydispersity of the extracted polymers. The native and alkali-treated agars exhibited comparable gelling and melting temperatures; however, the alkali-treated sample had higher gel strength than that of the native one. Chemical structure, molecular weight and rheological properties of the extracted galactans are discussed with regard to the algal source.

International audience; The purpose of the present study was to isolate and characterize ailanthone-rich materials from the bark of the deciduous tree Ailanthus altissima (Mill.) Swingle and to assess their herbicide activity on selected herbaceous species. Ailanthone-rich fractions were obtained from A. altissima bark by extraction with dichloromethane and ethyl acetate and subsequent purification of these crude extracts, and of the remaining water mixture after solvent extraction, by means of gel permeation chromatography. A number of fractions were isolated and characterized for ailanthone content. A dichloromethane fraction was shown to contain 92% w/w of ailanthone, as demonstrated by HPLC and NMR analysis. A significant pre-emergence herbicide activity was found for most of the extracts which was directly correlated to ailanthone concentration. A remarkable combined pre-and post-emergence herbicide activity was found for a specific fraction. These results indicate that the bark of A. altissima may represent an interesting source for the production of natural herbicides for use in agriculture.