Debranched cassava starch crystallinity determination by Raman spectroscopy: Correlation of features in Raman spectra with X-ray diffraction and C CP/MAS NMR spectroscopy

Because starch crystallinity influences the physical, mechanical, and technological aspects of numerous starch-based products during production and storage, rapid techniques for its assessment are vital. Samples of different levels of crystallinity were obtained by debranching gelatinized cassava starch, followed by subjection to various hydrothermal treatments. The recrystallized products were further subjected to partial hydrolysis with a mixture of α-amylase and glucoamylase prior to freeze-drying. Crystallinities were determined using X-ray diffraction (XRD) and ¹³C CP/MAS NMR spectroscopy, and correlated with FT-Raman spectra features. XRD crystallinities ranged between 0 and 58%, and agreed with crystalline-phase fractions (R² = 0.99) derived from the respective ¹³C CP/MAS NMR spectra. A strong linear correlation was found between crystallinities and integrated areas of the skeletal mode Raman band at 480 cm⁻¹ (R² = 0.99). With appropriate calibration, FT-Raman spectroscopy is a promising tool for rapid determination of starch crystallinity.     

Differences in crystalline cellulose modification due to degradation by brown and white rot fungi

Wood-decaying basidiomycetes are some of the most effective bioconverters of lignocellulose in nature, however the way they alter wood crystalline cellulose on a molecular level is still not well understood. To address this, we examined and compared changes in wood undergoing decay by two species of brown rot fungi, Gloeophyllum trabeum and Meruliporia incrassata, and two species of white rot fungi, Irpex lacteus and Pycnoporus sanguineus, using X-ray diffraction (XRD) and ¹³C solid-state nuclear magnetic resonance (NMR) spectroscopy. The overall percent crystallinity in wood undergoing decay by M. incrassata, G. trabeum, and I. lacteus appeared to decrease according to the stage of decay, while in wood decayed by P. sanguineus the crystallinity was found to increase during some stages of degradation. This result is suggested to be potentially due to the different decay strategies employed by these fungi. The average spacing between the 200 cellulose crystal planes was significantly decreased in wood degraded by brown rot, whereas changes observed in wood degraded by the two white rot fungi examined varied according to the selectivity for lignin. The conclusions were supported by a quantitative analysis of the structural components in the wood before and during decay confirming the distinct differences observed for brown and white rot fungi. The results from this study were consistent with differences in degradation methods previously reported among fungal species, specifically more non-enzymatic degradation in brown rot versus more enzymatic degradation in white rot.     

The surface structure of well-ordered native cellulose fibrils in contact with water

CP/MAS ¹³C NMR spectroscopy was used in combination with spectral fitting to examine the surface structure of hydrated cellulose I fibrils from Halocynthia and Gluconoacetobacter xylinus. To increase the spectral intensities and minimize signal overlap, G. xylinus celluloses site-specifically enriched in ¹³C either on C4 or on both C1 and C6 were examined. The experimental data showed multiple C4 and C6 signals for the water accessible fibril surfaces in the highly crystalline celluloses. These signal multiplicities were attributed to structural features in the surface layers induced by the fibril interior, and could not be extracted by spectral fitting in celluloses with a lower degree of crystallinity such as cellulose from cotton.     

Origin of the regioselectivity in an intramolecular nucleophilic addition to arene chromium tricarbonyl complexes

The source of the regioselectivity in the intramolecular nucleophilic addition of nitrile-stabilized carbanions to arene chromium tricarbonyl complexes was investigated for seven different substitution patterns on the arenes. All of the arenes are 1,4-dioxygenated and the substitution varies in the oxygen substituent and in the substituents of the arene carbons (hydrogen and alkyl). The regioselectivity is correlated with the preferred conformations of the chromium tricarbonyl group which in turn was determined by solution and solid-state ¹³C NMR spectroscopy, ¹H NMR spectroscopy in solution as well as X-ray diffraction. In the four complexes analyzed by X-ray diffraction and the three complexes analyzed by solid-state ¹³C NMR spectroscopy, there was only one complex where it was found that the preferred conformation of the -Cr(CO)₃ is different in solution than it is in the solid-state.     

Novel properties of cholesterol-dioleoylphosphatidylcholine mixtures

We have studied the properties of mixtures of cholesterol with dioleoylphosphatidylcholine (DOPC), and with several other phospholipids, including 1-stearoyl-2-oleoylphosphatidylcholine (SOPC) and dioleoleoylphosphatidylserine (DOPS), as a function of cholesterol molar fraction and of temperature. Mixtures of DOPC with a cholesterol molar fraction of 0.4 or greater display polymorphic behavior. This polymorphism includes the formation of structures that give rise to isotropic peaks in ³¹P NMR at cholesterol molar fractions between 0.4 and 0.6, dependent on the thermal history of the sample. Cryo-electron microscopy studies demonstrate the formation of small globular aggregates that would contribute to a narrowing of the ³¹P NMR powder pattern.At molar fraction cholesterol 0.6 and higher and at temperatures above 70 °C, the mixtures with DOPC convert to the hexagonal phase. Lipid polymorphism is accompanied by the phase separation of cholesterol crystals in the anhydrous form and/or the monohydrate form. The crystals that are formed have substantially altered kinetics of hydration and dehydration, compared with both pure cholesterol monohydrate crystals and with crystals formed in the presence of the other phospholipids that do not form the hexagonal phase in the presence of cholesterol. This fact demonstrates that these cholesterol crystals are in intimate contact with the DOPC phospholipid and are not present as morphologically separate structures.     

Crosslinked chitosan--preparation and characterization

Chitosan undergoes radical-induced depolymerization in the presence of potassium persulfate at 60 degrees C, leading to extensive crosslinking of the fragmented chains on subsequent cooling at 4 degrees C. As a result, a possible conformational change leading to higher crystallinity, as evidenced by IR, X-ray and 13C NMR was observed.     

Preparation and structural organisation of heteroleptic tetraphenylantimony(V) complexes comprising unidentately and bidentately coordinated O,O′-dialkyldithiophosphate groups: Multinuclear (C, P) CP/MAS NMR and single-crystal X-ray diffraction studies

O,O′-dipropyldithiophosphate and O,O′-di-iso-butyldithiophosphate (Dtph) tetraphenylantimony(V) complexes of the general formula [Sb(C₆H₅)₄{S₂P(OR)₂}] (R = C₃H₇, i-C₄H₉) were prepared and studied by means of ¹³C, ³¹P CP/MAS NMR spectroscopy and single-crystal X-ray diffraction. Distorted octahedral and trigonal bipyramidal molecular structures have been established for prepared complexes. These unexpected structural distinctions between chemically related compounds are defined by the principally different coordination modes of O,O′-dipropyldithiophosphate and O,O′-di-iso-butyldithiophosphate ligands in their molecular structures (i.e., S,S′-bidentate chelating and S-unidentately coordinated, respectively). To characterise quantitatively phosphorus sites in both species of dithiophosphate ligands, ³¹P chemical shift anisotropy parameters (δ_aniso and η) were calculated from spinning sideband manifolds in MAS NMR spectra. The ³¹P chemical shift tensors for the bidentate chelating and unidentately coordinated dithiophosphate ligands display a profoundly rhombic and nearly axially symmetric characters, respectively.     

The heterogeneous nature of microbial products as shown by solid-state13C CP/MAS NMR spectroscopy

Homoionic Na-, Ca-, and Al-clays were prepared from the <2 m fractions of Georgia kaolinite and Wyoming bentonite and mixed with sand to give artificial soils with 5, and 25% clay. The artificial soils were inoculated with microbes from a natural soil before incubation. Unlabelled and uniformly¹³C-labelled (99.9% atom) glucose were incorporated into the artificial soils to study the effects of clay types, exchangeable cations and clay contents on the mineralization of glucose-carbon and glucose-derived organic materials. Chemical transformation of glucose-carbon upon incorporation into microbial products and metabolites, was followed using solid-state¹³C CP/MAS NMR spectroscopy.There was a significant influence of exchangeable cations on the mineralization of glucose-carbon over a period of 33 days. At 25% clay content, mineralization of glucose-carbon was highest in Ca-soils and lowest in Al-soils. The influence of exchangeable cations on mineralization of glucose-carbon was more pronounced in soils with bentonite clay than those with kaolinite clay. Statistical analysis of data showed no overall effect of clay type on mineralization of glucose-carbon. However, the interactions of clay type with clay content and clay type with clay content and exchangeable cations were highly significant. At 25% clay content, the mineralization of glucose-carbon was significantly lower in Na- and Al-soils with Wyoming bentonite compared with Na- and Al-soils with Georgia kaolinite. For Ca-soils this difference was not significant. Due to the increased osmotic tension induced by the added glucose, mineralization of glucose-carbon was slower in soils with 5% clay than soils with 25% clay.Despite the differences in the chemical and physical characteristics of soils with Ca-, Na- and Al-clays, the chemical composition of organic materials synthesised in these soils were similar in nature. Assuming CP/MAS is quantitative, incorporation of uniformly¹³C-labelled glucose (99.9% atom) in these soils resulted in distribution of carbon in alkyl (24–25%), O-alkyl (56–63%), carbonyl (11–15%) and small amounts of aromatic and olefinic carbon (2–4%). However, as decomposition proceeded, the chemistry of synthesised material showed some changes with time. In the Ca- and Na-soils, the proportions of alkyl and carbonyl carbon decreased and that of O-alkyl carbon increased with time of incubation. However, the opposite trend was found for the Al-soil.Proton-spin relaxation editing (PSRE) subspectra clearly showed heterogeneity within the microbial products. Subspectra of the slowly-relaxing (long T₁(H)) domains were dominated by alkyl carbon in long- and short-chain structures. The signals due to N-alkyl (55 ppm) and carbonyl carbon were also strong in these subspectra. These subspectra were very similar to those obtained for microbial and fungal materials and were probably microbial tissues attached to clay surfaces by polysaccharide extracellular mucilage. Subspectra of fast-relaxing (short T₁(H)) domains comprised mostly O-alkyl and carbonyl carbon and were probably microbial metabolites released as neutral and acidic sugars into the extracellular environment, and strongly sorbed by clay surfaces.     

A study of ordered structure in acid-modified tapioca starch by C CP/MAS solid-state NMR

Acid modification of tapioca starch earlier reported to increase the mechanical strength of tablets. The development of ordered structure (double helices) of these starches was monitored after equilibrating at 0.90 a_w (25 °C) using ¹³C CP/MAS NMR and X-ray diffraction. As the hydrolysis time increased, the intensity of the resonance for C1 and C4 amorphous fractions decreased while that for C1 and C4 double helix fractions increased. Relative crystallinity (%) obtained from ¹³C CP/MAS NMR and X-ray diffraction methods both increased sharply initially and then levelled off with hydrolysis time. The initial increase in relative double helix content and crystallinity was due to a hydrolytic destruction in the amorphous domain, retrogradation of the partially hydrolyzed amylose and crystallization of free amylopectin double helices. After 192 h, these two parameters were not significantly different (=0.05) indicating that the double helices that were not arranged into crystalline regions either had been hydrolyzed or crystallized.     

Effects of organosolv pretreatment and enzymatic hydrolysis on cellulose structure and crystallinity in Loblolly pine

Ethanol organosolv pretreatment was performed on Loblolly pine to enhance the efficiency of enzymatic hydrolysis of cellulose to glucose. Solid-state ¹³C NMR spectroscopy coupled with line shape analysis was used to determine the structure and crystallinity of cellulose isolated from pretreated and enzyme-hydrolyzed Loblolly pine. The results indicate reduced crystallinity of the cellulose following the organosolv pretreatment, which renders the substrate easily hydrolyzable by cellulase. The degree of crystallinity increases and the relative proportion of para-crystalline and amorphous cellulose decreases after enzymatic hydrolysis, indicating preferential hydrolysis of these regions by cellulase. The structural and compositional changes in this material resulting from the organosolv pretreatment and cellulase enzyme hydrolysis of the pretreated wood were studied with solid-state CP/MAS ¹³C NMR spectroscopy. NMR spectra of the solid material before and after the treatments show that hemicelluloses and lignin are degraded during the organosolv pretreatment.     

Characterization of photosynthetic reaction centers with specific isotope labels

In this minireview the information at the atomic level that has been obtained by studying photosynthetic reaction centers with site-directed isotope labelling is discussed. The required isotopically labelled RCs are prepared using isotopically labelled amino acids and cofactors that have been prepared via organic total synthetic chemistry. In some cases the results of isotopically labelled RCs that are prepared via other methods are also discussed.     

Clean conversion of cellulose into fermentable glucose

We studied the process of conversion of microcrystalline-cellulose into fermentable glucose in the formic acid reaction system using cross polarization/magic angle spinning ¹³C-nuclear magnetic resonance, X-ray diffraction and Fourier transform infrared spectroscopy. The results indicated that formic acid as an active agent was able to effectively penetrate into the interior space of the cellulose molecules, thus collapsing the rigid crystalline structure and allowing hydrolysis to occur easily in the amorphous zone as well as in the crystalline zone. The microcrystalline-cellulose was hydrolyzed using formic acid and 4% hydrochloric acid under mild conditions. The effects of hydrochloric acid concentration, the ratio of solid to liquid, temperature (55–75 °C) and retention time (0–9 h), and the concentration of glucose were analyzed. The hydrolysis velocities of microcrystalline-cellulose were 6.14 × 10^− 3 h^− 1 at 55 °C, 2.94 × 10^− 2 h^− 1 at 65 °C, and 6.84 × 10^− 2 h^− 1 at 75 °C. The degradation velocities of glucose were 0.01 h^− 1 at 55 °C, 0.14 h^− 1 at 65 °C, 0.34 h^− 1 at 75 °C. The activation energy of microcrystalline-cellulose hydrolysis was 105.61 kJ/mol, and the activation energy of glucose degradation was 131.37 kJ/mol.     

Synthesis of a novel N[bond]O-interglycosidic disaccharide

The carbohydrate subunits carrying an N-O-interglycosidic bond play a very important role in the biological activity of the enediyne antibiotics. Condensation of O-(alpha- and beta-D-glucopyranosyl)hydroxylamine (5a and 5b) with the hex-3-ulopyranoside (6) furnished methyl 4,6-O-benzylidene-2,3-dideoxy-3-(2,3,4,6-tetra-O-benzyl-alpha-D-glucopyranosyloxy)imino-alpha- and beta-D-erythro-hexopyranoside (7a and 7b). Stereoselective reduction of the Cz.dbnd6;N bond of 7a and 7b with sodium cyanoborohydride resulted in the formation of the required protected N-O-interglycosidic disaccharides (8a and 8b). Finally, catalytic hydrogenation of 8a afforded methyl 2,3-dideoxy-3-(alpha-D-glucopyranosyloxy)amino-alpha-D-ribo-hexopyranoside (9a). Under similar conditions the beta anomer 8b underwent decomposition.     

Cultivation effects on the nature of organic matter in soils and water extracts using CP/MAS 13C NMR spectroscopy

The objective of this study was to examine the chemical structure of the organic matter (SOM) of Oxisols soils in slash and burn agriculture, in relation to its biological properties and soil fertility. The CP/MAS ¹³C technique was used to identify the main structural groups in litter and fine roots as SOM precursors; to identify the changes on the nature of the SOM upon cultivation and the proportion of labile and stable components; and to identify the nature of the organics present in water extracts (DOC). Carbohydrates were the main structural components in litter whereas components such as carbonyl C, carboxyl C,O-alkyl C and alkyl C were more common in SOM. Phenolic C and the degree of aromaticity were similar in litter and SOM. Cultivation resulted in a small decrease in the relative proportion of carbohydrates in SOM, little change in the levels of O-alkyl C and carbonyl C, but an increase in carboxyl C, phenolic C and aromaticity of the SOM. The level of alkyl C in soil was higher than the level of O-alkyl C, indicating the importance of long-chain aliphatics along with lignins in the stabilization of the SOM in Oxisols. The SOM of Mollisols from the Canadian Prairies differed from the Oxisol, with a generally stronger expression of aromatic structures, particularly in a cultivated soil in relation to a native equivalent. Carbohydrate components were the predominant structures in the DOC, indicating their importance in nutrient cycling and vertical translocations in the Oxisol.     

13C NMR chemical shifts can predict disulfide bond formation

The presence of disulfide bonds can be detected unambiguously only by X-ray crystallography, and otherwise must be inferred by chemical methods. In this study we demonstrate that ¹³C NMR chemical shifts are diagnostic of disulfide bond formation, and can discriminate between cysteine in the reduced (free) and oxidized (disulfide bonded) state. A database of cysteine ¹³C C and C chemical shifts was constructed from the BMRB and Sheffield databases, and published journals. Statistical analysis indicated that the C shift is extremely sensitive to the redox state, and can predict the disulfide-bonded state. Further, chemical shifts in both states occupy distinct clusters as a function of secondary structure in the C/C chemical shift map. On the basis of these results, we provide simple ground rules for predicting the redox state of cysteines; these rules could be used effectively in NMR structure determination, predicting new folds, and in protein folding studies.     

Synthesis and characterization of gold(III) complexes with alkyldiamine ligands

A series of gold(III) metalacycle of five-, six- and seven-membered ring was prepared by reacting Auric acid (HAuCl₄ · 3H₂O) with 1 equiv. unsubstituted ethylenediamine (en), propylene diamine (pn) and butylenediamine (bn) ligands and with some N-mono-substituted as well as N,N′-disubstituted ethylenediamine ligands. The general formula of these complexes is [Au(alkyldiamine)Cl₂]Cl. These complexes are characterized by melting point and elemental analysis, while structural analysis was done by spectroscopic techniques such as UV-Vis, Far-IR, IR spectroscopy, ¹H and ¹³C solution as well as ¹³C and ¹⁵ N solid-state NMR. The solid-state ¹⁵ N NMR shows that the chemical shift difference between free and bound ligand decreases as bn > pn > en, indicating stronger Au-N bond for bn complex compared to pn and en. UV-Vis shows relative stability of the Au(III) complexes of unsubstituted ethylenediamine with respect to N,N′-di-substituted ethylenediamine. Far-IR data show the six-membered metalacycle gold(III) alkanediamine complexes to be more stable. Spectroscopic data are evaluated by comparisons with calculated data of the built and optimized structure by gaussian03 at the RB3LYP level with LanL2DZ bases set.     

An NMR investigation of putative interresidue H-bonding in methyl α-cellobioside in solution

Methyl α-cellobioside (methyl β-d-glucopyranosyl-(1→4)-α-d-glucopyranoside) was labeled with ¹³C at C4′ for use in NMR studies in DMSO-d₆ solvent to attempt the detection of a trans-H-bond J-coupling (^3hJ_CCOH) between C4′ and OH3. Analysis of the OH3 signal at 600 MHz revealed only the presence of two homonuclear J-couplings: ³J_H3,OH3 and a smaller, longer range J_HH. No evidence for ^3hJ_C4′,OH3 was found. The longer range J_HH was traced to ⁴J_H4,OH3 based on 2D ¹H–¹H COSY data and inspection of the H2 and H4 signal lineshapes. A limited set of DFT calculations was performed on a methyl cellobioside mimic to evaluate the structural dependencies of ⁴J_H2,O3H and ⁴J_H4,O3H on the H3–C3–O3–H torsion angle. Computed couplings range from about −0.7 to about +1.1 Hz, with maximal values observed when the C–H and O–H bonds are roughly diaxial.     

Biotransformation of lignin polymers derived from beech wood pulping by Sporobolomyces roseus isolated from leafy material

The ability of the yeast, Sporobolomyces roseus, isolated from leafy material, to modify lignin derived from beechwood pulping was examined by FTIR and 13C NMR spectroscopy, which revealed oxidative cleavage of the Calpha-Cbeta linkages between lignin units. Using veratryl alcohol as a model substrate confirmed that Sp. roseus could oxidize veratryl alcohol into veratric acid. This yeast might be suitable for the pretreatment of lignocellulosic materials and/or for biotransformation of technical lignins.     

Physicochemical characterization of carrageenan fromGigartina teedii (Rooth) Lamouroux (Gigartinales,Rhodophyta)

The phycocolloids of female gametophytes ofGigartina teedii (Roth) Lamouroux harvested in Roscoff (Brittany, France) are a hybrid carrageenan resulting from juxtaposition of fragments of kappa-, iota-and nu-carrageenan. They represent 70% of the dry matter of the alga in summer. After alkaline transformation the proportion of iota-carrageenan increased to 76%, demonstrating the presence of nu-carrageenan. Absence of mu-carrageenan, the precursor of kappa-carrageenan, suggests that iota-carrageenan is desulfated enzymically to kappa-carrageenan.