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The 1 H- and 13 C-NMR spectra of the ethyl and benzyl esters and the tetrabutylammonium and tetraethylammonium salts of hyaluronic acid { 2)-β- d -Glc p A-(1 → 3)-β- d -Glc p NAc-(1 n } in Me 2 SO- d 6 have been assigned using 1D and 2D techniques. The chemical shifts of the resonance of GlcNAc C-3 suggest that the relative orientations of the monosaccharides at the (1 → 3) linkage in the esters and salts are different. Small differences in the chemical shifts of the resonance GlcA C-4 suggest only a slight conformational variation around the (1 → 4) linkage. The 13 C-NMR data also suggest similarities in conformation between the esters in Me 2 SO- d 6 and the salts in water. The chemical shifts of the 1 H resonances for NH and OH groups and their temperature dependence for the esters and salts in Me 2 SO reveal markedly stronger inter-residue hydrogen bonds between the carboxyl and NH groups and between HO-4 of GlcA and O-5 of GlcNAc for the salts. The 3 J 2,NH values indicate a slightly different orientation for the acetamido group. For solutions in Me 2 SO, the higher segmental flexibility of the esters is supported by the line widths, whereas the reduced viscosity for the tetrabutylammonium salt showed a sigmoidal concentration dependence and suggests association of chains which could contribute to the segmental rigidity. The linear concentration dependence for the benzyl ester suggests a higher overall flexibility without chain association.

Some aspects of the physiology of chondrocytes from horse articular cartilage were studied, since this animal model can be helpful in understanding arthritic processes. The replicative ability of articular chondrocytes, measured by the incorporation of [ 3 H]thymidine, and their capacity of proteoglycan production, evaluated from the incorporation of [ 35 S]sulfate, are very low. In addition, these cells do not differentiate in vitro as shown by the constant specific activity of alkaline phosphatase measured at different times in culture. Two types of potassium channels were identified by patch clamp experiments in the cell-attached configuration, one characterized by a conductance of 40 pS and the other of 100 pS. No active K + channels were found at V pip = 0. It was shown by Furs-2 experiments that the low replicative ability is paralleled by a modest variation of the intracellular calcium concentration after a mitogenic stimulus. 31 P NMR experiments, both on slices of whole articular cartilage and on isolated cells, demonstrate that chondrocytes derive their energy mainly from the glycolytic pathway.

A series of oligosaccharides was prepared from hyaluronate by depolymerisation with bovine testicular hyaluronidase. Complete assignment of the 1 H and 13 C NMR spectra was obtained for the disaccharide, the tetrasaccharide, and the NaBH 4 -treated tetrasaccharide, by using various 1D and 2D NMR methods. The 1 H assignments for the tetrasaccharide differ from the incomplete data reported recently (ref. 11). The 13 C NMR spectra of the aqueous di, tetra-, hexa-, and octa-saccharides of this series show that all resonances, apart from those subject to obvious end effects, have chemical shifts comparable to those of the corresponding resonances of hyaluronate in D 2 O. The observed 13 C chemical shifts suggests that cooperative intramolecular hydrogen bonds probably play a minor role in determining the conformation of hyaluronate in water.

The Klebsiella pneumoniae K40 capsular polysaccharide has been isolated and investigated by use of methylation analysis, specific degradations and NMR spectroscopy. The polysaccharide was depolymerised by a bacteriophage-associated endogalactosidase, and the resulting oligosaccharide was characterised by one-dimensional and two-dimensional NMR spectroscopy and direct chemical ionisation MS. The repeating unit of the K40 capsular polysaccharide was shown to be a linear hexasaccharide with the composition-->3)- alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->4)-alpha-D-GlcpA++ +-(1-->2-)- alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->3)-alpha-D-Galp-(1--> (Rha, rhamnose).

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.