Extent Of Acylation Research Articles

Homogeneous acylation of eucalyptus wood at room temperature in dimethyl sulfoxide/N-methylimidazole

Succinoylation and benzoylation of ball-milled eucalyptus wood using succinic anhydride and benzoyl chloride as acylating reagent, respectively, were investigated at room temperature using dimethyl sulfoxide/N-methylimidazole (DMSO/NMI) as reaction medium without additional catalysts. The effects of the corresponding acylating reagent dosage (1–5:1 for succinoylation and 0.5–5:1 for benzoylation) and reaction time (0.35–5h for succinoylation and 0.5–3h for benzoylation) on the extent of acylation, measured by weight percent gain (WPG), were studied. WPG of succinoylation and benzoylation was in the range of 70.8–144.7% and 17.3–43.1%, respectively. The efficiency of acylation at room temperature significantly increased in DMSO/NMI compared with ionic liquid 1-butyl-3-methylimidazolium chloride because of the role of NMI as solvent, base and catalyst. Fourier transform infrared (FT-IR), and solid-state cross-polarization/magic-angle spinning (CP/MAS) 13C nuclear magnetic resonance (NMR) spectroscopy studies provided evidence for the occurrence of succinoylation and benzoylation reactions and the attachment of functional groups via ester bonds.

Modification of sugarcane bagasse with acetic anhydride and butyric anhydride in ionic liquid 1-butyl-3-methylimidazolium chloride

Bagasse acetate butyrates were prepared homogeneously in 1-butyl-3-methylimidazolium chloride ([C4mim]Cl) ionic liquid from ball-milled sugarcane bagasse by acylation with acetic anhydride and butyric anhydride. The parameters, including reaction temperature, reaction time, feeding method of adding anhydrides, the dosage of total anhydrides to SCB, and the molar ratio of acetic anhydride to butyric anhydride, were considered, and the extent of acylation was measured by weight percent gain (WPG). The results showed the positive effects of reaction duration and total anhydride dosage on WPG and the negative effects of reaction temperature and molar ratio of AA/BA on WPG. The feeding method of acetylation after butyrylation resulted in the increased WPG compared with acetylation before butyrylation. FT-IR and 1H–13C correlation 2D NMR (HSQC) studies provided evidence for acylation. The bagasse acetate butyrates showed increased thermal stability after acylation. This study provides a new way for high value-added utilization of renewable lignocellulosic biomass.

The relationships between physicochemical properties and conformational features of succinylated and acetylated kidney bean (Phaseolus vulgaris L.) protein isolates

The effects of succinylation and acetylation on some physicochemical properties and conformational features of kidney bean protein isolate (KPI) were investigated. The extent of N-acylation progressively increased up to 93–94% with increasing anhydride-to-protein ratio (0–1.0 g g −1 ), while the O-acylation occurred only after extensive blocks of ε-amino groups. Zeta potential and size exclusion chromatography analyses indicated that the acylation and the succinylation in particular led to decreases in isoelectric point and zeta potential at neutral pH, in an extent of acylation dependent manner, while the association/dissociation state of the vicilin was slightly affected by the acylation. The surface hydrophobicity was also remarkably affected by the acylation, but the influence varied with the type and extent of acylation. Differential scanning calorimetry (DSC), intrinsic fluorescence and near-UV and/or far-UV CD spectroscopic analyses showed significant changes in tertiary and/or secondary conformations of the proteins in KPI, with extent of acylation, especially extent of succinylation. These phenomena suggest close relationships between the physicochemical properties and tertiary and/or secondary conformational features of the proteins in acylated KPI samples. It is also suggested that the physicochemical and conformational properties of KPI could be modulated by selecting the type and level of the applied anhydrides.

Tailoring of Stimuli-Responsive Water Soluble Acrylamide and Methacrylamide Polymers

The tailoring of the lower critical solution temperature (LCST) polymers of acrylamide and methacrylamide in water is achieved by chemical modification of freely water soluble precursor polymers poly[N-2-hydroxypropylmethacrylamide], poly[N,N-bis(hydroxyethyl)acrylamide] and poly[N-{tris(hydroxymethyl)methyl}acrylamide]. Two principal reactions, namely acetylation and cinnamoylation, are applied. By varying the acylating agent as well as the extent of acylation, the LCST can be tailored easily. The cloud points observed for the different polymer series do not correlate with the apparent hydrophilicity of the parent polymers according to the content of hydroxyl groups. The results thus exemplify the difficulties to predict the behaviour of modified thermosensitive polymers by simply analysing the balance of hydrophilic to hydrophobic molecular fragments. Chemical modification by cinnamoylation provides photoreactive copolymers e.g susceptible to photocrosslinking. When the polymers are prepared by polymerization using a disulfide-functionalized azo-initiator, efficient grafting of the modified copolymers on gold surfaces is possible to prepare ultrathin hydrogel films, as demonstrated by Surface Plasmon Resonance.

Poly(methyl methacrylate)-supported hydroxamic dithiocarbonic anhydrides: preparation and use as acyl transfer reagents

The use of poly(methyl methacrylate) (PMMA)-supported hydroxamic dithiocarbonic anhydrides in the synthesis of amides is illustrated. The reagent was found to selectively acylate amines and amino acids to give the corresponding amides. The efficiency of the acyl transfer reaction was found to be affected by the nature of the crosslinking agent, the nature of the solvent, the temperature, the reaction period, and the effective concentration of the reagent function. The percentage yield of amides obtained using N,N′-methylene-bis-acrylamide (N,N′-MBA)-crosslinked hydroxamic dithiocarbonic anhydride resins was found to be higher than that of ethylene glycol dimethacrylate (EGDMA)- and divinylbenzene (DVB)-crosslinked ones. It was also observed that the extent of acylation was higher in polar solvents such as acetonitrile. The acyl transfer reaction was favored by high temperature and with a high resin to amine ratio.

Protease-catalyzed synthesis of disaccharide amino acid esters in organic media

An enzymatic synthesis of sugar amino acid esters has been developed in pyridine by using a commercial protease, Optimase M-440, from Bacillus licheniformis. Optimase M-440 showed broad substrate specificity toward amino acid esters as acyl donors and disaccharides as nucleophiles. Analysis of acylation sites indicated sucrose was acylated only at three primary hydroxyls. Trehalose, consisting of two glucose units, was acylated at two primary (6-OH and 6′-OH) and one secondary (3-OH). With sucrose and trehalose, diesters along with monoesters were enzymatically synthesized. In both cases, no triester was formed. Molecular size of nucleophiles, glucose and sucrose, have effects on the extent of acylation with d-amino acids. N-blocked phenylalanine, leucine, and methionine exhibited higher activity toward sucrose than lysine, aspartic acid, and tyrosine. Various leaving groups of tBoc- l-phenylalanine were used and cyanomethyl ester gave the highest rate of reaction. Optimization of initial water activity of Optimase M-440 ( A w = 0.2–0.3) was necessary to maintain catalytic activity and to prevent undesirable hydrolysis of activated esters.

Synthesis and antioxidant activity of water‐soluble chitosan derivatives

AbstractWe prepared three different water‐soluble chitosan derivatives by the acylation of chitosan with corresponding acid anhydrides. The extent of acylation was determined by using the electric conductivity method. All of the chitosan derivatives were soluble enough to examine the respective mltioxidant activity in aqueous media. The chitosan derivatives inhibited thiobarbituric acid reactive substrate (TBARS) formation in t‐butyl hydroperoxide and benzoyl peroxide induced lipid peroxidations. However, they did not show any scavenging activity toward superoxide.

Site-specific acylation of GABA-gated Cl − channels: effects on 36Cl − uptake

Radioligand binding studies indicate that p-isothiocyanato-t-butylbicycloorthobenzoate (p-NCS-TBOB) specifically acylates GABA-gated chloride channels. Preincubation of synaptoneurosomes with p-NCS-TBOB followed by washing resulted in a concentration dependent (63–500 nM) inhibition of both muscimol-stimulated chloride uptake and [355]t-butylbicyclophosphorothionate (TBPS) binding. The extent of acylation (assessed by inhibition of [ 35S]TBPS binding) was highly correlated (r = 0.89; p < 0.0001) with the inhibition of muscimol-stimulated Cl − uptake. Neither basal Cl − uptake nor [ 3H]muscimol binding to GABA A receptors were affected by p-NCS-TBOB. Preincubation with the nonacylating ‘cage’ convulsant t-butylbicycloorthobenzoate (500 nM) followed by washing had no effect on either muscimol-stimulated Cl − uptake or [ 35S]TBPS binding. These findings indicate that p-NCS-TBOB interferes with the efficacy of muscimol promoted channel openings, but does not affect the recognition qualities of GABA A receptors. p-NCS-TBOB should prove useful in electrophysiological and biochemical studies examining the properties of GABA-gated Cl − channels.

Effect of Growth Conditions on the Production, Composition and Viscosity of Xanthomonas campestris Exopolysaccharide

Summary: A streptomycin-resistant variant of Xanthomonas campestris was grown in defined nutrient-deficient media in both batch and continuous culture. The production, composition and viscosity of the extracellular polysaccharide (xanthan) synthesized by this strain were influenced by the fermentation time and nutrient exhaustion in batch culture and by the dilution rate in continuous culture. The specific rate of exopolysaccharide synthesis was maximal during exponential growth in all of the nutrient-deficient media studied although some xanthan was formed during stationary phase. The concentration of exopolysaccharide decreased at later stages of stationary phase in some cultures. Both the extent of acylation and the consistency index of xanthan isolates were low or minimal during exponential growth, maximal in polysaccharide isolated as the growth rate fell and usually lower after this time. Between dilution rates of 0·03 and 0·06 h−1 in chemostat culture, the cell and exopolysaccharide dry weights were independent of dilution rate, the specific rate of xanthan synthesis decreasing at lower growth rates. Although the variation in the acyl content and consistency index was less than that observed in batch culture, exopolysaccharide isolated at higher dilution rates tended to have a higher acetyl content, lower pyruvyl content and lower consistency index.

Selective acylation of membrane proteins in Acholeplasma laidlawii

In membranes of the cell-wall-less prokaryote Acholeplasma laidlawii most proteins are of the integral type. A substantial fraction of these proteins are enriched in hydrophilic amino acid residues. Approximately 20 different major as well as minor proteins were found to be covalently modified with acyl chains. The same set of proteins are acylated when cells are grown in different fatty-acid-supplemented media. In individual proteins the ratio of palmitoyl/oleoyl acyl chains was 12-14 times larger than the acyl chain ratio in polar membrane lipids. The transmembrane protein D12 has close to two acyl chains per molecule. Proteins T2 and T4a, localized in the outer and inner leaflet of the membrane, respectively, occur each as pairs with a difference in relative molecular mass within each pair of approximately 2000. Each of these proteins as well as the other acyl proteins, except the light form of T4a, has close to one acyl chain per molecule. The extent of acylation was increased for certain proteins and decreased for others by treatment with globomycin or phenethylalcohol. The relative amounts of the T2 and T4a pairs were affected by these drugs. It is concluded that the mechanism of acylation is different from that in Escherichia coli lipoprotein and Bacillus penicillinase. The mean hydrophobicity [Kyte & Doolittle (1982) J. Mol. Biol. 157, 105-132] of the A. laidlawii acyl proteins are similar to those of other bacterial acyl proteins but significantly lower than for non-acylated integral membrane proteins, supporting an anchoring function of the acyl chains. The number of membrane acyl proteins in A. laidlawii and two other mycoplasmas are at least twice that in other bacteria.

Effect of Acetylation and Succinylation on the Functional Properties of Winged Bean (Psophocarpus tetragonolobus) Flour

ABSTRACTWinged bean flour was acylated to various degrees and changes in functional properties were monitored. Nitrogen solubility in water increased up to ?80% acylation and then decreased. Water absorption capacity increased continuously with the extent of acylation. The fat absorption capacity of the acetylated flour increased up to ?30% acetylation and then decreased while that of the succinylated flour remained unchanged. Emulsion capacity showed the same trend as nitrogen solubility in water. Foam capacity increased due to acylation but foam stability decreased. Bulk density of the acetylated flour increased up to ?80% acetylation and then decreased. Succinylated flour had higher bulk density at all levels of succinylation.

On the function of half-site reactivity: Intersubunit NAD +-dependent activation of acyl-glyceraldehyde 3-phosphate dehydrogenase reduction by NADH

Glyceraldehyde 3-phosphate dehydrogenase extracted from sturgeon muscle, exhibits half-site reactivity in its reaction with the pseudo-substrate β -(2-furyl) acryloyl phosphate ( Malhotra & Bernhard, 1968 ). The product is the difurylacryloyl thiol ester enzyme tetramer formed from the active site cysteinyl residue. The electronic spectrum of the furylacryloyl thiol ester linkage is perturbed upon binding of oxidized coenzyme (NAD + ) at the acyl site ( Malhotra & Bernhard, 1973 ). Likewise, we now report the perturbation in electronic spectrum of this furylacryloyl thiol ester upon interaction with NADH at the acyl site. The spectral perturbation accompanying NADH binding is large and in the opposite direction from that due to the binding of NAD + . The “color” of the chromophoretic acyl-enzyme linkage is a reflection of its chemical properties: furylacryloyl-enzyme-NAD complex is “red-shifter” (λ max =360 nm), and reacts uniquely with inorganic phosphate to yield the corresponding acyl phosphate. Furylacryloyl-enzyme-NADH complex is “blue-shifter” (λ max =330 nm) and is uniquely capable of reduction by NADH. Despite the presence of a ubiquitous NADH-induced color change, NADH reduction occurs only when NAD is bound to the remaining two non-acylated subunits. Hence, these results provide a direct example of allosteric regulation of enzyme function by ligand interaction at an adjacent subunit site. This site is known to be at least 17 A removed from the site of reduction ( Buehner et al. , 1974 ). Carboxymethylation of the two remaining (active) thiol groups of the diacyl-enzyme tetramer, a process which modifies the affinity for NAD at this site to only a small extent, does not affect the NAD-dependent activation of the acyl-NADH site for reduction. Qualitative transient kinetic evidence suggests that NAD binding to the non-acylated sites favors a slow conformational change leading to reducibility at acyl sites. The affinity of the various sites, acylated and non-acylated, for coenzyme ligands varies with the extent of acylation and the extent and nature of the ligand occupancies at adjacent subunits. Since the molar concentration of glyceraldehyde 3-phosphate dehydrogenase sites in muscle sarcoplasmic fluid is high (of the order of 1 pm), these results suggest that the “enzyme” functions as a regulatory warehouse for energy via acyl-enzyme-ATP interconversion and NADH/NAD redox potential regulation.

Nature of the Interaction of Dextran Sulfate with Low Density Lipoproteins of Plasma

Abstract The nature of the interaction between dextran sulfate and low density human plasma lipoproteins of the Sf 0 to 10 class (LDL) was investigated by modifying LDL with succinic anhydride or acetic anhydride. In the absence of divalent metal ions, the formation of insoluble and soluble dextran sulfate-LDL complexes was prevented by the acylation of approximately 16% and 25%, respectively, of the LDL free amino groups. The decrease in the positive charges of the protein moiety was primarily responsible for the interference with the complex formation. An increase in the negative charges introduced by succinylation did not interfere as extensively as the reduction in positive charges. The presence of magnesium sulfate facilitated the conversion of succinylated and acetylated lipoproteins to insoluble complex. The amount of Mg++ required for the conversion progressively increased with increases in the extent of acylation. Although the succinylated LDL required more Mg++ than the acetylated LDL, the difference was relatively small until approximately 75% of the lipoprotein amino groups was modified. A steep increase in the magnesium requirement observed upon further increase in the extent of succinylation reflected an extensive increase in negative charges due to succinylation of LDL free hydroxyl groups. The evaluation of the requirement for Mg++ by acetylated and succinylated LDL revealed the involvement of both LDL positive and negative charges in the formation of insoluble complex in the presence of divalent metal ions. Although the treatment of LDL with phospholipase C hydrolyzed as much as 56% of the phospholipids, the LDL was almost completely converted to insoluble complex in both the presence and absence of Mg++. Since minor modification by succinylation or acetylation drastically reduced the reactivity of LDL with dextran sulfate, the protein-charged groups rather than the phospholipid-charged groups seem to play the major role in the dextran sulfate-LDL interaction.

Thiolation for labeling of proteins, using catalysis by silver and imidazole combined

1. 1.|Compounds containing primary amino groups could be thiolated rapidly at pH 7.5–8 by N-acetylhomocysteine thiolactone with catalysis by Ag + and imidazole present together. 2. 2.|Using equimolar thiolactone and Ag + the characteristics of these reagent mixtures at varying ratios of imidazole were investigated. Suggestions are made as to the species present in the course of these reactions. 3. 3.|Hydrolysis of the thiolactone competed with aminolysis and limited acylation of amino compounds. The extent of acylation was measured on glycine and was improved by increasing the concentration of Ag + and thiolactone (equimolar) but not by increasing the relative imidazole content, which somewhat increased the rates. 4. 4.|Rabbit γ-globulin was extensively thiolated using 0.3 M Ag + and thiolactone, with 2 and with 8 equiv of imidazole. For comparison, globulin thiolated with catalysis by imidazole alone was also extensively (although slowly) acylated but lost many of the thiol groups in the absence of the protecting Ag +. 5. 5.|A procedure for protein thiolations is given, and factors governing the choice of the reagent mixtures are discussed.