First application of mass spectrometry and gas chromatography in investigation of α-cellulose hydrolysates: the influence of climate changes on glucose molecules in pine tree-rings.

We present the first results of the quantitative and qualitative gas chromatographic and isotope ratio mass spectrometric analysis of monosaccharides derived from acid hydrolysis of α-cellulose extracted from annual pine tree-rings. The conifers investigated in this study grew in the Niepolomice Forest in Poland, and the annual rings covered the time span from 1940 to 2000 AD. The main components of the α-cellulose samples were two saccharides: glucose and mannose. The amount of glucose in the annual rings varied between 17 and 44%. The δ(13)C of glucose was found to be less negative than that of α-cellulose and the δ(18)O values in glucose were less positive than those in α-cellulose. The content of monosaccharides in the α-cellulose samples has an influence on the isotope fractionation factors. The values of the carbon isotope fractionation factor increase with an increase in the monosaccharides concentration in α-cellulose, while the values of the oxygen isotope fractionation factor decrease with an increase in monosaccharides concentration in α-cellulose. The challenge is to establish, with respect to climate changes and environmental conditions, the significance of the interannual variations in the observed monosaccharide concentration.

Enthalpic studies of xyloglucan-cellulose interactions.

We report a study of xyloglucan (XG)-cellulose interactions made possible by the preparation of various well-defined cellulosic and xyloglucosidic substrates. Bacterial microcrystalline cellulose (BMCC) as well as cellulose whiskers (CellWhisk) were used as cellulosic substrates. Xyloglucosidic substrates were obtained from Rubus cells and Tamarindus indica seeds. Different primary structure characteristics of XGs such as the backbone length and the nature of the side chains, as well as their repartition, were considered in order to examine the influence of the primary structure on their interaction capacity. Two complementary approaches were carried out: first, the determination of adsorption isotherms and its associated models, and second, an enthalpic study using isothermal titration calorimetry (ITC). This study highlighted that an increase of XG interaction capacity occurred with increasing XG molecular weight. Furthermore, we determined that a minimum of 12 glucosyl residues on the backbone is required to observe significant interactions. Moreover, both the presence of trisaccharidic side chains with fucosyl residues and an increase of unsubstituted glucosyl residues enhanced XG-cellulose interactions. The evolution of adsorption isotherms with temperature and ITC measurements showed that two different processes were occurring, one exothermic and one endothermic, respectively. Although the presence of an exothermic interaction mechanism has long been established, the presence of an endothermic interaction mechanism has never been reported.

A novel 1,3-ß-glucan synthase from the oomycete Saprolegnia monoica.

An apparently novel 1,3-ß-glucan synthase from the oomycete Saprolegnia monoica has been characterized. The enzyme exhibits properties that differ markedly from those of the enzyme previously described [Fèvre, M. & Dumas, C. (1977). J Gen Microbiol 103, 297-306] as it is active at alkaline pH, stimulated by the divalent cations Ca2+, Mg2+ and Mn2+, and appears to be located mainly in the apical part of the hypha. Taking into consideration the differences in pH optimum and effect of divalent ions, each enzyme activity could be assayed in the presence of the other. The insoluble polymeric product of the enzyme with alkaline pH optimum was characterized as a linear 1,3-ß-glucan. Comparisons of the general properties of 1,3-ß-glucan synthases suggest that enzymes from the oomycetes are more closely related to enzymes from higher plants than to those of true fungi, reflecting the fact that the oomycetes are highly divergent from chitinous fungi.

An arabinogalactan from the skin of Opuntia ficus-indica prickly pear fruits.

The cold-water extract from the skin of Opuntia ficus-indica fruits was fractionated by anion-exchange chromatography. The major fraction, which was purified by size exclusion chromatography, consisted of a polysaccharide composed of galactose and arabinose residues in the ratio 6.3:3.3, with traces of rhamnose, xylose and glucose, but no uronic acid. The results of methylation analysis, supported by (13)C NMR spectroscopy, indicated that this polysaccharide corresponded to an arabinogalactan having a backbone of (1-->4)-linked beta-D-galactopyranosyl residues with 39.5% of these units branched at O-3. The side-groups consisted either of single L-arabinofuranosyl units or L-arabinofuranosyl alpha-(1-->5)-linked disaccharides. This polysaccharide is thus an arabinogalactan that can be classified in the type I of the arabinogalactan family.

Conversion of a non-water soluble potato starch waste into reducing sugars under non-conventional technologies.

In this exploratory work, the comparison of the utilisation of different non-conventional technologies (ultrasound and microwave irradiations) for the depolymerisation of a complex industrial starch-based waste into reducing sugars was investigated. Reducing sugars could then be converted into higher value-added compounds such as higher alcohols. The experiments were performed on three different starting materials named as 'Potato flour', 'Wet potato sludge' and 'Dry potato sludge'. The conversion of 'Potato flour' into reducing sugars reached in acidic conditions 61% within an hour under microwave irradiation, 70% and 84% within 120 min under low and high frequency ultrasonic irradiation, respectively.

The molecular structure and solution conformation of an acidic heteropolysaccharide from Auricularia auricula-judae.

A water soluble acidic heteropolysaccharide named WAF was isolated from Auricularia auricula-judae by extracting with 0.9% NaCl solution. By using gas chromatography, gas chromatography-mass spectrometry, and NMR, its chemical structure was determined to be composed of a backbone of α-(1→3)-linked D-mannopyranose residues with pendant side groups of ß-D-xylose, ß-D-glucose, or ß-D-glucuronic acid at position O6 or O2. Six fractions prepared from WAF with a weight-average molecular mass (M(w)) between 5.9 × 104 and 64.7 × 104 g/mol were characterized with laser light scattering and viscometry in 0.1M NaCl at 25°C. The dependence of intrinsic viscosity ([η]) and radius of gyration (R(g)) on M(w) for this polysaccharide were found to be [η] = 1.79 × 10⁻³ M(w) °.96 cm³ g⁻¹ and R(g) = 6.99 × 10⁻² M(w) (0.54) nm. The molar mass per unit contour length (M(L)) and the persistence length (L(p)) were estimated to be 1124 nm⁻¹ and 11 nm, respectively. The WAF exhibited a semirigid character typical of linear polysaccharides. Molecular modeling was then used to predict the ordered and disordered states of WAF; the simulated M(L) and L(p) were however much smaller than the experimental values. Taken altogether, the results suggested that WAF formed a duplex in solution.

Identification of cellulolytic bacteria in soil by stable isotope probing.

Plant residues, mainly made up of cellulose, are the largest fraction of organic carbon material in terrestrial ecosystems. Soil microorganisms are mainly responsible for the transfer of this carbon to the atmosphere, but their contribution is not accurately known. The aim of the present study was to identify bacterial populations that are actively involved in cellulose degradation, using the DNA-stable isotope probing (DNA-SIP) technique. (13)C-cellulose was produced by Acetobacter xylinus and incubated in soil for 7, 14, 30 and 90 days. Total DNA was extracted from the soil, the (13)C-labelled (heavy) and unlabelled (light) DNA fractions were separated by ultracentrifugation, and the structure of active bacterial communities was analysed by bacterial-automated ribosomal intergenic spacer analysis (B-ARISA) and characterized with denaturing gradient gel electrophoresis (DGGE). Cellulose degradation was associated with significant changes in bacterial community structure issued from heavy DNA, leading to the appearance of new bands and increase in relative intensities of other bands until day 30. The majority of bands decreased in relative intensity at day 90. Sequencing and phylogenetic analysis of 10 of these bands in DGGE profiles indicated that most sequences were closely related to sequences from organisms known for their ability to degrade cellulose or to uncultured soil bacteria.