Carboxylic OH Groups Research Articles

Structural features, kinetics and SAR study of radical scavenging and antioxidant activities of phenolic and anilinic compounds

BackgroundPhenolic compounds are widely distributed in plant kingdom and constitute one of the most important classes of natural and synthetic antioxidants. In the present study fifty one natural and synthetic structurally variant phenolic, enolic and anilinic compounds were examined as antioxidants and radical scavengers against DPPH, hydroxyl and peroxyl radicals. The structural diversity of the used phenolic compounds includes monophenols with substituents frequently present in natural phenols e.g. alkyl, alkoxy, ester and carboxyl groups, besides many other electron donating and withdrawing groups, in addition to polyphenols with 1–3 hydroxyl groups and aminophenols. Some common groups e.g. alkyl, carboxyl, amino and second OH groups were incorporated in ortho, meta and para positions.ResultsSAR study indicates that the most important structural feature of phenolic compounds required to possess good antiradical and antioxidant activities is the presence of a second hydroxyl or an amino group in o- or p-position because of their strong electron donating effect in these positions and the formation of a stable quinone-like products upon two hydrogen-atom transfer process; otherwise, the presence of a number of alkoxy (in o or p-position) and /or alkyl groups (in o, m or p-position) should be present to stabilize the resulted phenoxyl radical and reach good activity. Anilines showed also similar structural feature requirements as phenols to achieve good activities, except o-diamines which gave low activity because of the high energy of the resulted 1,2-dimine product upon the 2H-transfer process. Enols with ene-1,2-diol structure undergo the same process and give good activity. Good correlations were obtained between DPPH inhibition and inhibition of both OH and peroxyl radicals. In addition, good correlations were obtained between DPPH inhibition and antioxidant activities in sunflower oil and liver homogenate systems.ConclusionsIn conclusion, the structures of good anti radical and antioxidant phenols and anilines are defined. The obtained good correlations imply that measuring anti DPPH activity can be used as a simple predictive test for the anti hydroxyl and peroxyl radical, and antioxidant activities. Kinetic measurements showed that strong antioxidants with high activity have also high reaction rates indicating that factors stabilizing the phenoxyl radicals lower also the activation energy of the hydrogen transfer process.

Selective Amine Recognition Driven by Host–Guest Proton Transfer and Salt Bridge Formation

The stepwise synthesis of ionizable p-tert-butylcalix[5]arenes 1a·H and 1b·H, featuring a fixed cone cavity endowed with a carboxyl moiety at the narrow rim, is described. Single-crystal X-ray analyses have shown that in the solid state 1a·H and 1b·H adopt a cone-out conformation with the carboxylic OH group pointing in, toward the bottom of the aromatic cavity, as a result of a three- or two-center hydrogen-bonding pattern between the carboxyl group and the phenolic oxygen atom(s). The affinity of amines for calix[5]arene derivatives 1a·H and 1b·H was probed by (1)H NMR spectroscopy and single-crystal X-ray diffraction studies. These carboxylcalix[5]arenes are shown to selectively recognize linear primary amines--over branched, secondary, and tertiary amines--by a two-step process involving a proton transfer from the carboxyl to the amino group to provide the corresponding alkylammonium ion, followed by binding of the latter inside the cavity of the ionized calixarene. Proton transfer occurs only with linear primary amines, that is, when the best size and shape fit between host and substrate is achieved, while the other amines remain in their noncompeting unprotonated form. The role of the solvent in the ionization/complexation process is discussed. Structural studies on the n-BuNH(2) complexes with 1a·H and 1b·H provide evidence that binding of the in situ formed n-BuNH(3)(+) substrate to the cavity of the ionized macrocycle is ultimately secured, in the case of 1a·H, by the formation of an unprecedented salt-bridge interaction.

3-(6-Aminopyridinium-3-yl)benzoate monohydrate

The title compound, C12H10N2O2·H2O, crystallizes as a zwitterion in which the pyridine N atom is protonated and the carboxyl OH group is deprotonated. The benzene and pyridinium rings are inclined at a dihedral angle of 54.93 (1)°. In the crystal, O—H⋯O and N—H⋯O hydrogen bonds link the mol­ecules into a three-dimensional supra­molecular network.

Surprisingly complex supramolecular behaviour in the crystal structures of a family of mono-substituted salicylic acids

Crystal structure packing analyses of a family of monosubstituted salicylic acids, including some hydrates, show a surprisingly varied choice of supramolecular arrangements, even though the expected carboxyl–carboxyl dimer occurs in 17 of the 24 structures studied, and the 2-hydroxyl group hydrogen bonds with the carboxyl carbonyl oxygen in all molecules. In three of the four hydrates (3-MeO, 3NO2 and 5-Cl), the water molecules disrupt the carboxyl–carboxyl dimer to form a centrosymmetric [carboxyl–water]2 cluster. In the fourth hydrate, of the 5-acetamido (ACM) derivative, one water molecule forms an (ACM)CO⋯H–O(H)⋯H–O(carbox) single bridge. In the 4-ACM structure, the acetamido and carboxyl groups interact via (ACM)CO⋯H–O(carbox) and (ACM–Me)C–H⋯OC(carbox) double bridges, whilst the 6-MeO molecules, in which the carboxyl OH group is H-bonding intramolecularly to the MeO oxygen, forming catemers via bifurcating carboxyl OH and CO functions. The remaining outlier is the 5-amino (5-NH2) derivative, which exists in a zwitterionic form, with one carboxyl oxygen forming a short O⋯H–N intermolecular bond to an –NH3+ group, whilst the second oxygen forms three longer O⋯H–N bonds to three different –NH3+ groups. The larger, “normal” group of 17 derivatives adopt 16 different structures, generally comprising stacks of molecules with varying relative shifts of the molecular planes, and interactions involving the substituents. Surprisingly, the family contains only two polymorphic pairs, 5-Br α, β and 5-I α, β, in each of which the forms show low similarities, but two isostructures, the 5-Br α and 5-I β forms.

3-Carboxy-5-(pyridinium-4-yl)benzoate: a redetermination

The title compound, C13H9NO4, crystallizes in a zwitterionic form with the pyridine N atom protonated and the carboxyl OH group deprotonated. The benzene and pyridinium rings are inclined with a dihedral angle of 31.42 (14)° between them. A previous report of this stucture claims, we believe incorrectly, that neither of the carboxyl­ate groups is deprotonated [Zhang et al. (2010 ▶). Acta Cryst. E66, o2928–o2928]. In the crystal, inter­molecular O—H⋯O, N—H⋯O and weak C—H⋯O hydrogen-bonding inter­actions link adjacent mol­ecules into a three-dimensional supra­molecular network.

4-Oxo-1,4-dihydrobenzo[h][1,3]thiazeto[3,2-a]quinoline-1,3-dicarboxylic acid

In the title mol­ecule, C16H9NO5S, there is an intra­molecular O—H⋯O hydrogen bond involving the quinolone carbonyl O atom and a carboxyl OH group. In the crystal, inter­molecular O—H⋯O hydrogen bonds between the carbonyl group of the quinolone carboxyl group, and a second carboxyl group on the thia­zeto moiety lead to the formation of chains propagating along [201] and perpendicular to the π-stacks of mol­ecules.

Nickel(II) and zinc(II) complexes of N-substituted di(2-picolyl)amine derivatives: Synthetic and structural studies

The interaction of di(2-picolyl)amine (1) and its secondary N-substituted derivatives, N-(4-pyridylmethyl)-di(2-picolyl)amine (2), N-(4-carboxymethyl-benzyl)-di(2-picolyl)amine (3), N-(4-carboxybenzyl)-di(2-picolyl)amine (4), N-(1-naphthylmethyl)-di(2-picolyl)amine (5), N-(9-anthracenylmethyl)-di(2-picolyl)amine (6), 1,4-bis[di(2-picolyl)aminomethyl]benzene (7), 1,3-bis[di(2-picolyl)aminomethyl]benzene (8) and 2,4,6-tris[di(2-picolyl)amino]triazine (9) with Ni(II) and/or Zn(II) nitrate has resulted in the isolation of [Ni(1)(NO3)2], [Ni(2)(NO3)2], [Ni(3)(NO3)2], [Ni(4)(NO3)2]·CH3CN, [Ni(5)(NO3)2], [Ni(6)(NO3)2], [Ni2(7)(NO3)4], [Ni2(8)(NO3)4], [Ni3(9)(NO3)6]·3H2O, [Zn(3)(NO3)2]·0.5CH3OH, [Zn(5)(NO3)2], [Zn(6)(NO3)2], [Zn(8)(NO3)2] and [Zn2(9)(NO3)4]·0.5H2O. X-ray structures of [Ni(4)(NO3)2]·CH3CN, [Ni(6)(NO3)2] and [Zn(5)(NO3)2] have been obtained. Both nickel complexes exhibit related distorted octahedral coordination geometries in which 4 and 6 are tridentate and bound meridionally via their respective N3-donor sets, with the remaining coordination positions in each complex occupied by a monodentate and a bidentate nitrato ligand. For [Ni(4)(NO3)2]·CH3CN, intramolecular hydrogen bond interactions are present between the carboxylic OH group on one complex and the oxygen of a monodentate nitrate on an adjacent complex such that the complexes are linked in chains which are in turn crosslinked by intermolecular offset π–π stacking between pyridyl rings in adjacent chains. In the case of [Ni(6)(NO3)2], two weak CH⋯O hydrogen bonds are present between the axial methylene hydrogen atoms on one complex and the oxygen of a monodentate nitrate ligand on a second unit such that four hydrogen bonds link pairs of complexes; in addition, an extensive series of π–π stacking interactions link individual complex units throughout the crystal lattice. The X-ray structure of [Zn(5)(NO3)2] shows that the metal centre once again has a distorted six-coordinated geometry, with the N3-donor set of N-(1-naphthylmethyl)-di(2-picolyl)amine (5) coordinating in a meridional fashion and the remaining coordination positions occupied by a monodentate and a bidentate nitrato ligand. The crystal lattice is stabilized by weak intermolecular interactions between oxygens on the bound nitrato ligands and aromatic CH hydrogens on adjacent complexes; intermolecular π–π stacking between aromatic rings is also present.

Serine–Ca2+versus serine–Cu2+complexes — A theoretical perspective

The association of Ca2+and Cu2+to serine was investigated by means of B3LYP DFT calculations. The [serine–M]2+(M = Ca, Cu) potential energy surfaces include, as does the neutral serine, a large number of conformers, in which a drastic reorganization of the electron density of the serine moiety is observed. This leads to significant changes in the number and strength of the intramolecular hydrogen bonds existing in the neutral serine tautomers. In some cases, a proton is transferred from the carboxylic OH group to the amino group and accordingly, some of the more stable [serine–M]2+complexes can be viewed as the result of the interaction of the zwiterionic form of serine with the doubly charged metal ion. Whereas the interaction between Ca2+and serine is essentially electrostatic, that between Cu2+and serine has a non-negligible covalent character, reflected in larger electron densities at the bond critical points between the metal and the base, in the negative values of the electron density between the two interacting systems, and in much larger Cu2+than Ca2+binding energies. More importantly, the interaction with Cu2+is followed by a partial oxidation of the base, which is not observed when the metal ion is Ca2+. The main consequence is that in Cu2+complexes a significant acidity enhancement of the serine moiety takes place, which strongly favors the deprotonation of the [serine–Cu]2+complexes. This is not the case for Ca2+complexes. Thus, [serine–Ca]2+complexes, like those formed by urea, thiourea, selenourea, or glycine, should be detected in the gas phase. Conversely, the complexes with Cu2+should deprotonate spontaneously and therefore only [(serine–H)–Cu]+monocations should be experimentally accessible.

Thermal stability and structure of a new co-crystal of theophylline formed with phthalic acid

A new co-crystal of theophylline and phthalic acid with 1:1 molar ratio has been prepared. It crystallises in the monoclinic crystal system, space group P21/c, a=11.5258(9), b=10.1405(6), c=13.9066(12) A, β=106.827(4)°. The structure of the co-crystal has been revealed by single crystal X-ray diffraction. An infinite helical polymeric chain is formed by intermolecular hydrogen bonds of the two neutral constituents. The hydroxyl group and carbonyl oxygen atom in one of the carboxyl groups of phthalic acid form hydrogen bonds to O6 and to N(7)H atoms of theophylline, respectively, while the other carboxyl OH group of phthalic acid is in hydrogen bond to N9 atom of theophylline by very strong intermolecular interactions proven by 1883 cm−1 centred peak in FTIR spectrum. Thermal degradation of this new supramolecular compound is a two-step process in air. At first phthalic acid (47.4%) released up to 230°C, meanwhile it loses water and transforms into phthalic anhydride. In EGA-MS spectra, the characteristic fragments of water (m/z=17, 18) appear from about 180°C, while absorption bands of phthalic anhydride are shown in EGA-FTIR spectrum at about 210°C. In the second step theophylline begins to sublime, melts at 276°C, and then evaporates up to 315°C with minute residues.

4-(Diphenylphosphinoyl)benzoic acid

Mol­ecules of the title compound, C19H15O3P, are connected by O—H⋯O hydrogen bonds between the carboxylic acid OH group and the phosphinoyl O atom, forming chains running along the crystallographic b axis.

Solid state molecular assemblies of five bile acid derivatives

Abstract The crystal structures of five bile acid derivatives are reported: 1: 6α,7α-Dihydroxy-5β-cholan-24-oic acid; 2: 3α,6α-Dihydroxy-5β-cholan-24-oic acid (Hyodeoxycholic acid); 3: 3α,7α,12α-Trihydroxy-5β-cholan-24-hydrazide hemihydrate; 4: 3-Dimethyl ketal,7,12-dioxo-5β-cholan-24-oic acid and 5: 3α,7α-Di-O-acetyl-12-oxo-5β-cholan-24-oic acid methyl ester. In all the structures the four saturated cycles, forming the common alicyclic steroidal skeleton, have the same conformation, while the flexible side chain adopts different conformations. Whereas all the crystal structures of Cholic Acid inclusion compounds are characterized by the formation of a bilayer-type structure with channels within the lipophilic layers, the chemical variations of the substituents on the cholanic framework induce remarkable changes in the H-bond scheme and in the crystal packing arrangements. For instance, the structures 1, 2, having a different number and position of the hydroxyl groups on the rigid skeleton, display a variety of supramolecular architectures dominated by networks of cooperative …O—H…O… H-bonds which do not produce typical molecular layers and channels of Cholic Acid inclusion compounds. Cholic hydrazide hemihydrate, 3, where both the hydrazide group and water molecule are strongly involved in the H-bond system, forms an overall structural motif similar to that observed in CA inclusion compounds containing hydrophilic and lipophilic layers and small channels. Compound 4, containing only the carboxylic OH group as H-bond donor, makes simple antiparallel chains of molecules, while compound 5, lacking of H-bond donors, forms a crystal aggregation dominated by weak C—H…O contacts and van der Waals interactions leading to a crystal packing where neither molecular layers nor channels are recognizable.

Transdermal Delivery of Ketoprofen: The Influence of Drug–Dioleylphosphatidylcholine Interactions

Considering that most inflammatory diseases occur locally and near the body surface, transdermal delivery of non-steroidal anti-inflammatory drugs (NSAIDs) may be an interesting strategy for delivering these drugs directly to the diseased site. To optimize ketoprofen (KP) transdermal delivery we investigated the influence of dioleylphosphatidylcholine (DOPC) on skin permeation. The formulations studied were: i) a physical mixture of KP and DOPC and ii) DOPC and KP complex, in a molar ratio of 1:3, obtained by dissolution of the components in chloroform followed by drying under a N2 atmosphere. Both systems were dispersed in mineral oil and the in vitro percutaneous was assayed by absorption using a flow through diffusion cell. Differential Scanning Calorimetry (DSC) and 1H NMR studies were carried out to characterize KP and DOPC interactions. Geometry optimizations using Density Functional Theory and semiempirical methods, as well as a flexible docking procedure were carried out to obtain a binding model for KP with DOPC. KP solubility and partition studies in the formulations, as well as skin irritation and hypersensitivity assays were also carried out. DSC determinations in the complex showed enthalpy and temperature depressions, indicating KP and DOPC interaction. In addition, dipole-dipole interactions between the KP carboxylic acid and OH groups in phospholipids were shown by 1H NMR studies. Based on the NMR studies, a KP-DOPC binding model is proposed, in which KP is involved by the two long aliphatic chains of the phospholipid. Solubility studies indicated that DOPC improved drug solubility. KP permeation was enhanced by both formulations tested, but the complex also increased its skin uptake. Such behavior could be attributed to the solubilizing, melting and enhancing effects of DOPC. Skin irritation and hypersensitivity were not significantly changed compared to control, suggesting that the formulation may be therapeutically explored for KP transdermal delivery.

Why are α-hydroxycarboxylic acids poor chiral modifiers for Pt in the hydrogenation of ketones?

Mandelic acid ( 1) and its derivatives and 1-naphthylglycolic acid ( 8) were used as modifiers for the asymmetric hydrogenation of ketopantolactone to pantolactone on supported Pt, Ru, and Rh catalysts. A systematic variation of the modifier structure showed that 8 was the best modifier, affording up to 28% ee on Pt; the other metals were barely efficient. The catalytic studies were completed with FTIR spectroscopic analysis of the modifier–modifier and modifier–substrate interactions in solution. Attenuated total reflection infrared spectroscopy was applied to study the adsorption of the α-hydroxy-carboxylic acid type modifiers on a Pt/Al 2O 3 model catalyst in the presence of hydrogen. The study indicates that the modifiers are present on the metal surface as monomers with an internal hydrogen bond between the hydroxyl group and the carboxyl carbonyl group, and the phenyl or naphthalene rings are in a tilted position relative to the surface. The moderate enantioselection is attributed to a hydrogen bond interaction between the carboxylic OH group of the modifier and the carbonyl O atom of the substrate. The poor enantioselectivities attained in ketopantolactone hydrogenation may be explained by the overly weak adsorption of the modifiers on Pt and by the weakness of the O H O-type H bond between the substrate and the modifiers. It seems that the carboxyl group has a double role, allowing interaction with the substrate and the Pt surface.

First experimental evidence of the third conformer of glycolic acid: combined matrix isolation, FTIR and theoretical study

The FT-IR spectra of monomeric glycolic acid, GA, isolated in argon and xenon matrices (at 10 and 20 K) were studied, enabling identification of three different conformers of GA. The most stable SSC conformer involves a hydrogen bond between the carbonyl group and the hydrogen atom of the alcohol hydroxyl group. The minor GAC and AAT forms involve intramolecular hydrogen bonds between alcohol and carboxyl OH groups, with alcohol and carboxylic groups serving as proton donors, respectively. The GAC conformer, predicted by theoretical calculations (at the DFT/B3LYP and MP2 levels with the 6-31++G(d,p) and aug-cc-pVDZ basis sets) as the second most stable form, was now experimentally identified for the first time.

Crystal Structure of 4-Dimethylaminoazobenzene-4'-sulfonyl-L-tryptophan Dimethylsulfoxide Solvate

Crystals of 4-dimethylaminoazobenzene-4′-sulfonyl-L-tryptophan dimethylsulfoxide solvate (Dabs-Trp·DMSO) are monoclinic, space group P21 with a = 12.941(6)A, b = 7.760(3)A, c = 14.131(7)A, β = 103.161(8)° and Z = 2. The structure was solved by direct methods (SIR88) and refined to a final R value of 0.058 for 3152 reflections (I > 2.00σ(I)). The molecular geometries and conformations are comparable to the typical values of azobenzene and L-tryptophan. The DMSO molecule is involved in the intermolecular hydrogen bonds with the sulfonamide NH and the carboxyl OH groups of different Dabs-Trp molecules. The hydrogen-bonded chains propagate along the b axis, and are constructed from molecules related by a two-fold screw axis. The sulfonamide S=O and the carboxyl C=O groups do not participate in hydrogen bonding.

Alpha-gamma hybrid peptides that contain the conformationally constrained gabapentin residue: characterization of mimetics of chain reversals.

The crystal structures of four dipeptides that contain the stereochemically constrained gamma-amino acid residue gabapentin (1-(aminomethyl)cyclohexaneacetic acid Gpn) are described. The molecular conformation of Piv-Pro-Gpn-OH (1), reveals a beta-turn mimetic conformation, stabilized by a ten atom C[bond]H...O hydrogen bond between the Piv CO group and the pro S hydrogen of the Gpn CH(2)[bond]CO group. The peptides Boc-Gly-Gpn-OH (2), Boc-Aib-Gpn-OH (3), and Boc-Aib-Gpn-OMe (4) form compact, folded structures, in which a distinct reversal of polypeptide chain direction is observed. In all cases, the Gpn residue adopts a gauche,gauche (g,g) conformation about the C(gamma)[bond]C(beta) (theta(1)) and C(beta)[bond]C(alpha) (theta(2)) bonds. Two distinct Gpn conformational families are observed. In peptides 1 and 3, the average backbone torsion angle values for the Gpn residue are phi=98 degrees, theta(1)=-62 degrees, theta(2)=-73 degrees, and psi=79 degrees, while in peptide 2 and 4 the average values are phi=-103 degrees, theta(1)=-46 degrees, theta(2)=-49 degrees, and psi=-92 degrees. In the case of 1 and 3, an intramolecular nine-membered O[bond]H...O hydrogen bond is formed between the C[double bond]O of the preceding residue and the terminal carboxylic acid OH group. All four alpha-gamma dipeptide sequences yield compact folded backbone conformations; this suggests that the Gpn residue may be employed successfully in the design of novel folded structures.

Interaction of uranium(VI) with various modified and unmodified natural and synthetic humic substances studied by EXAFS and FTIR spectroscopy

The complexation of uranium(VI) by humic acids (HAs) and fulvic acids (FAs) was studied to obtain information on the binding of uranium(VI) onto functional groups of humic substances. For this, various natural and synthetic HAs were chemically modified resulting in HAs with blocked phenolic OH groups. Both from the original and from the modified humic substances, solid uranyl humate complexes were prepared at pH 2. FTIR and extended X-ray absorption fine structure (EXAFS) spectroscopy were applied to study the chemical modification process of humic substances, to study the structure of uranyl humate complexes and to evaluate the effect of individual functional groups of humic substances (carboxylic and phenolic OH groups) on the complexation of uranyl ions. The results confirmed the predominant blocking of phenolic OH groups in the modified HAs. These modified HAs are suitable model substances to study the role of phenolic OH groups of HAs in dependence on pH. By EXAFS spectroscopy, identical structural parameters were determined for all uranyl humates. Axial UO bond distances of 1.78 Å were determined. In the equatorial plane approximately five oxygen atoms were found at a mean distance of 2.39 Å. The blocking of phenolic OH groups of HAs did not change the near-neighbor surrounding of uranium(VI) in uranyl humate complexes. Thus, the results confirmed that predominantly HA carboxylate groups are responsible for binding of uranyl ions and that the influence of phenolic OH groups is insignificant under the applied experimental conditions. The carboxylate groups are monodentate coordinated to uranyl ions.

Role of beta-lactam carboxyl group on binding of penicillins and cephalosporins to class C beta-lactamases.

Molecular models for the Henry Michaelis complexes of Enterobacter cloacae, a class C beta-lactamase, with penicillin G and cephalotin have been constructed by using molecular mechanic calculations, based on the AMBER force field, to examine the molecular differentiation mechanisms between cephalosporins and penicillins in beta-lactamases. Ser318Ala and Thr316Ala mutations in both complexes and Asn346Ala and Thr316Ala/Asn346Ala double mutation in penicillin G complex have also been studied. Results confirm that Thr316, Ser318, and Asn346 play a crucial role in the substrate recognition, via their interactions with one of the oxygens of the antibiotic carboxyl group. Both mutation Ser318Ala and Thr316Ala strongly affect the correct binding of cephalotin to P99, the first mainly by precluding the discriminating salt bridge between carboxyl and serine OH groups, and the second one by the Ser318, Lys315, and Tyr150 spatial rearrangements. On the other hand, Ser318Ala mutation has little effect on penicillin G binding, but the Thr316Ala/Asn346Ala double mutation causes the departure of the antibiotic from the oxyanion hole. Molecular dynamic simulations allow us to interpret the experimental results of some class C and A beta-lactamases.

Carbon-13 NMR spectroscopic studies on chemically modified and unmodified synthetic and natural humic acids

Carbon-13 cross-polarization magic-angle spinning nuclear magnetic resonance spectroscopy ( 13C-CP/MAS-NMR) was applied to study the chemical modification process of humic acids (HA) with diazomethane and the subsequent alkaline hydrolysis of the methylated HA. This modification process results in HA with selectively blocked phenolic OH groups, which can be used for metal ion binding studies with humic substances. Different chemically modified and unmodified natural and synthetic HA with carbon-13 of natural abundance were investigated. In addition, carbon-13 labeled modified synthetic HA, that were synthesized with [ 13C]diazomethane as methylation reagent, were studied to confirm the assumed modification process and to determine the type of functional groups that have the highest affinity for methylation with diazomethane. The results of the NMR studies with carbon-13 labeled modified HA show that predominantly carboxyl and phenolic OH groups are methylated with diazomethane resulting in methyl ester and methyl ether groups, respectively. Due to the alkaline treatment of the methylated HA, the methyl esters of carboxyl groups are hydrolyzed, whereas methyl ethers of phenolic OH groups remain unchanged, which results in modified HA with blocked phenolic OH groups. From the spectra of the modified and unmodified HA with carbon-13 of natural abundance it can be concluded that the applied preparative modification procedure causes only the desired structural changes in HA.

Solubilization of low-rank coal by Trichoderma atroviride: evidence for the involvement of hydrolytic and oxidative enzymes by using 14C-labelled lignite.

The deuteromycete Trichoderma atroviride is able to solubilize lignite in dependence on a given carbon source for growth. When cultivated on media containing glutamate, this mold excreted a set of different enzymes with hydrolytic activity. Addition of lignite to the growth media induced the synthesis of extracellular lignite-specific esterase activity but no evidence has been provided for its direct involvement in the process of lignite solubilization. Hence, the basic capability of T. atroviride enzymes to degrade a variety of ester and ether bonds at the surface or within the bulky lignite structure was tested using coal following its direct labelling with 14C-alkyl iodide. The participation of hydrolytic and oxidative enzymes in lignite degradation was assessed by measuring the release of 14C radioactivity from selectively alkylated carboxylic and phenolic OH groups. T. atroviride cleaved both carboxylic esters using esterases and the phenolic ether bonds by using oxidative enzymes, most likely laccases.