Resolution of non-protein amino acids via the microbial protease-catalyzed enantioselective hydrolysis of their N-unprotected esters

In the Aspergillus oryzae protease-catalyzed ester hydrolysis, substitution of N-unprotected amino acid esters for the corresponding N-protected amino acid esters resulted in a large enhancement of the hydrolysis rate, while the enantioselectivity was deteriorated strikingly when the substrates employed were the conventional methyl esters. This difficulty was overcome by employing esters bearing a longer alkyl chain such as the isobutyl ester. Utilizing this ester, amino acids carrying an aromatic side chain were resolved with excellent enantioselectivities (E=50 to >200). With amino acids bearing an aliphatic side chain also, good results in terms of the hydrolysis rate and enantioselectivity were obtained by employing such an ester as the isobutyl ester. Moreover, the enantioselectivity proved to be enhanced further by conducting the reaction at low temperature. This procedure was applicable to the case where the enantioselectivity was not high enough even by the use of the isobutyl ester.     

Equilibrium and hydrolysis ofα-amino acid esters in ternary complexes of copper(II) involving glycyl-L-tyrosine

Summary Binary and ternary complexes of copper(II) with glycyl-L-tyrosine and an amino acid ester were investigated by potentiometric and spectrophotometric techniques. The kinetics of base hydrolysis of the above esters in the presence of copper(II)-glycyl-L-tyrosine complex was studied at 30°C.     

Ternary complexes involving copper(II) and amino acids,peptides and DNA constituents. The kinetics of hydrolysis of α-amino acid esters

Binary and ternary complexes of copper(II) involving picolylamine (Pic) and amino acids, peptides (HL) or DNA constituents have been investigated. Ternary complexes of amino acids or peptides are formed by simultaneous reactions. Amino acids form the Cu(Pic)L complex, whereas peptides form Cu(Pic)L and Cu(Pic)(LH_–1). The ternary complexes of copper(II) with picolylamine and DNA are formed in a stepwise process, whereby binding of copper(II) to picolylamine is followed by ligation of the DNA components. The stability of the ternary complexes is compared with the stabilities of the corresponding binary complexes. The hydrolysis of glycine methyl ester (MeGly) is catalysed by the Cu(pic)²⁺ complex. The kinetic data is fitted assuming that the hydrolysis proceeds in two steps. The first step, involving coordination of the amino acid ester by the amino and carbonyl groups, is followed by rate-determining attack by the OH^– ion. The second step involves equilibrium formation of the hydroxo-complex, Cu(pic)(MeGly)(OH), followed by intramolecular attack.     

Equilibrium and hydrolysis of α-amino acid esters in mixed-ligand complexes withN-(acetamido)-iminodiacetatecopper(II)

Summary The formation equilibria of the binary and ternary complexes of Cu^II withN-(acetamido)-iminodiacetic acid (ADA) and amino acids or their esters were investigated potentiometrically. The chelation mode was ascertained by conductivity measurements. The kinetics of the base hydrolysis of amino acid esters in the presence of the copper(II)-ADA complex were studied. The rate and catalysis constants were estimated.     

Organosilicon synthesis of isocyanates: IV. Synthesis of isocyanates from aliphatic and alkylaromatic amino acid esters

Treatment of an alcoholic suspension of amino acids with trimethylchlorosilane yielded phenylglycine, valine, β-phenylalanine, and homovaline ester hydrochlorides. Their saccharin-catalyzed silylation with hexamethyldisilazane proceeds quantitatively and involves only one proton of the amino group. The best conversion of the amino acid esters to the corresponding isocyanates was achieved by phosgene treatment of their monosilyl urethanes, rather than of the silylated amino esters. Monosilyl urethanes are formed quantitatively by treatment of the amino acid ester hydrochlorides with the hexamethyldisilazane-CO₂ system. The ¹H NMR spectra show that monosilyl urethanes derived from α-and β-amino acid esters are characterized by intramolecular interaction of the silicon atom and the oxygen atom of the carboxy group.     

Synthesis and specific biodegradation of novel polyesteramides containing amino acid residues

Novel polyesteramides were synthesized from p‐nitrophenyl esters of sebacic or adipic acids and diamines containing α‐amino acid ester groups. The optimal polymerization condition was 60 °C in N,N‐dimethylformamide. The structures of these polymers were confirmed by IR and NMR. The number‐average molecular weights of these polyesteramides ranged from 2280 to 23,600 (except for the polymers containing glycine residues), depending on the nature of the amino acid used. The biodegradability of the polyesteramides was investigated by in vitro hydrolysis with proteases and a lipase as catalysts in borate buffer solutions. The results indicated that the polymers containing L ‐phenylalanine were hydrolyzed most effectively by α‐chymotrypsin, subtilisin Carlsberg, and subtilisin BPN′. The polyesteramides containing other amino acid residues also underwent hydrolysis to different extents, reflecting the substrate specificity of the proteases. Lipase had almost no effect on the hydrolytic degradation of these polyesteramides. The polymers containing glycine residues were hardly decomposed by any of the enzymes used. © 2001 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 39: 1318–1328, 2001     

Environmentally benign process for the synthesis of N-formyl amino acid esters

Several amino acid ester hydrochlorides were reacted with ammonium formate to give N-formyl amino acid esters in good yields.     

Calix[4]resorcinols functionalized with amino acid residues

By the Mannich reaction in ternary systems calix[4]resorcinol-amino acid (amino acid ester, amino acid ester hydrochloride)-formaldehyde calix[4]resorcinols were obtained functionalized on the upper molecule rim by amino acid residues.     

Assignment of the configurations of the amino acids in peptidic siderophores

Pyoverdins and azotobactins contain beta-hydroxyaspartic acid, N delta-hydroxyornithine, citrulline and homoserine, in addition to the common protein amino acids. Configuration assignment of all of these was achieved by acid hydrolysis of the peptide, derivatization of the constituent amino acids to the N-pentafluoropropionyl amino acid esters and gas chromatographic separation of the stereoisomers on capillaries coated with Chirasil-Val. This approach is straightforward for the protein amino acids, but the less common amino acids are either partially degraded during acid hydrolysis or their derivatives exhibit unfavourable gas chromatographic properties. By judicious combination of partial and total hydrolysis and dual derivatization, these problems may be overcome.     

Synthesis of Optically Active,Ring-Substituted N-Benzyloxycarbonylphenylalanines via 2-Benzyloxycarbonylamino-2-arylalkylmalonates

Diethyl or dimethyl benzyloxycarbonylaminomalonate was reacted with ring substituted benzyl halides and the resulting arylalkyl derivatives ( 3 to 6 ) half-saponified to the DL -monoacid-monoesters ( 7 to 10 ). Decarboxylation by refluxing in dioxane afforded the N-benzyl oxycarbonyl-DL -amino acid esters ( 11 to 14 ), which were resolved into their optical antipodes by enzymic hydrolysis of the ester group with Subtilisin, type Carlsberg. Enzymic hydrolysis led to the N-benzyloxycarbonyl-L -amino acids ( 15 to 18 ) and to the corresponding D -amino acid esters. The latter were converted to the N-benzyloxycarbonyl-D -amino acids ( 19 and 20 ) by alkaline hydrolysis of the ester groups. These derivatives could be used directly for further peptide synthesis. The following compounds were prepared: N-benzyloxycarbonyl derivatives of p-methyl-L -phenylalanine ( 15 ), p-methyl-D -phenylalanine ( 19 ), p-fluoro-L -phenylalanine ( 16 ), m-fluoro-L -phenylalanine ( 17 ), m-fluoro-D -phenylalanine ( 20 ) and penta-fluoro-L -phenylalanine ( 18 ). The free amino acids were obtained by removal of the benzyloxycarbonyl group with HBr in acetic acid.     

Synthesis of symmetric bis(imidazole-4,5-dicarboxamides) substituted with amino acids

A series of symmetric bis(imidazole-4,5-dicarboxamides) (bis-I45DCs) were prepared with amino acid esters and a variety of linker groups. The critical pyrazine intermediates, substituted with amino acid esters, were synthesized by stoichiometric control of the amino acid ester, even though primary alkanamines, in comparison, generally offer less selectivity for this reaction. Diamines are added to subsequently react with and open the remaining acyl imidazole bonds in the pyrazine intermediates and thereby yield the bis-I45DCs.     

Synthesis of C-11 linked active ester derivatives of vitamin D3 and their conjugations to 42-residue helix-loop-helix peptides

Derivatives of vitamin D₃ carrying an 8-carbon linker at C-11 terminating in an active ester were synthesized from commercial vitamin D₃ using a disassembly-reassembly strategy. Vitamin D₃ was cleaved at the C6-C7 double bond and the ‘upper’ fragment was converted, via a series of reactions, to derivatives substituted at C-11 with an 8-carbon linker terminating in an ethyl ester. Reassembly with modified ‘lower’ fragments using Horner-Wittig olefination followed by linker ester hydrolysis and re-esterification with p-nitrophenol gave C-11 substituted p-nitrophenyl esters. These vitamin D derivatives were conjugated to 42-amino acid helix-loop-helix peptides by reaction of their p-nitrophenyl esters with lysyl side-chain amino groups on the peptides. The vitamin D—peptide conjugates, being potential specific binder candidates for vitamin D-binding protein, were characterized by mass spectroscopy and CD measurements.     

Complex-formation reactions of dicholoro(S-methyl-L-cysteine)palladium(II) with bio-relevant ligands. Labilization induced by S-donor chelates

The complex-formation equilibria of [Pd(SMC)(H2O)2]+, where SMC = S-methyl-l-cysteinate, with bio-relevant ligands such as amino acids, peptides, dicarboxylic acids and DNA constituents were studied and their formation constants were determined. The binding mode of the ligands containing various functional groups was studied and the speciation diagrams were evaluated. The kinetics of base hydrolysis of amino acid esters bound to [Pd(SMC)(H2O)2]+ was studied in aqueous solution at 25 degrees C and 0.1 M ionic strength. The effect of solvent polarity and temperature on the hydrolysis of coordinated glycine methyl ester was investigated. The activation parameters are evaluated and discussed.     

N-(5-氯-2-羟苯基)氨基酸酯的合成、表征及抑菌活性

以5-Cl水杨醛和L-苯丙氨酸、L-亮氨酸及L-甘氨酸的脂肪酸酯为原料,通过碱催化的席夫碱缩合反应,合成了6种N-（5-氯-2-羟苄基）席夫碱氨基酸酯及其还原产物N-（5-氯-2-羟苄基）氨基酸酯。 化合物的结构及组成经过IR、¹H NMR和元素分析测试技术进行了表征。 合成的席夫碱及其还原产物对革兰氏阴性菌、革兰氏阳性菌及真菌均有不同程度的抑制作用。 质量分数为0.01%的N-（5-氯-2-羟苄基）席夫碱氨基酸酯对大肠杆菌的抑菌率达90%以上,而N-（5-氯-2-羟苄基）氨基酸酯对金黄色葡萄球菌的抑菌率也在90%以上,均为强抑菌活性,其中N-（5-氯-2-羟苄基）苯丙氨酸酯的抑菌率达98%以上。     

Amphiphilic Anthracene-Amino Acid Conjugates as Simple Carbohydrate Receptors in Water

N -Alkylation of amino acid methyl esters with 9,10-bis-chloromethylanthracene, followed by methyl ester hydrolysis, yielded water-soluble amphiphilic amino acid-anthracene conjugates. In NMR-titration experiments, a l -arginine-anthracene conjugate was demonstrated to recognize p -nitrophenyl glycosides and sialylated oligosaccharides in D 2 O in a manner sensitive to the carbohydrate structure.     

In situ formation of N-trifluoroacetoxy succinimide (TFA-NHS): one-pot formation of succinimidyl esters, N-trifluoroacetyl amino acid succinimidyl esters, and N-maleoyl amino acid succinimidyl esters

A method for the in situ formation of N-trifluoroacetoxy succinimide (TFA-NHS) and its application in the formation of succinimidyl esters is presented. The developed method provides N-trifluoroacetyl and N-maleoyl amino acid succinimidyl esters from a variety of amino acids using a one-pot, high-yielding protocol. Investigations into the formation of an N-maleoyl amino acid succinimidyl ester supported the proposal of a revised reaction mechanism, and contributed to the optimization of the reaction conditions.     

Amino acids and peptides. XXIII. Synthesis of N alpha-protected amino acid 6-chloro-2-pyridyl esters and their evaluation for peptide synthesis

6-Chloro-2-pyridyl esters (OPyCl) of N alpha-benzyloxycarbonyl and tert-butyloxycarbonylamino acids were synthesized by the N,N'-dicyclohexylcarbodiimide (DCC) method from the acids and 6-chloro-2-hydroxypyridine in dimethylformamide (DMF). The reactivity of the 6-chloro-2-pyridylester with amino group is much higher than that of the corresponding 2-pyridyl ester (OPy) and p-nitrophenyl esters (ONp) in dioxane and DMF, and a peptide bond is formed without acylation at the side chain hydroxyl group of amino acids. Z-Asp(OBzl)-OPyCl reacted with amino acid methyl esters in dioxane to give the corresponding dipeptide without any detectable aspartimide formation.     

Caged O-phosphorothioyl amino acids as building blocks for Fmoc-based solid phase peptide synthesis

The synthesis of 1-(2-nitrophenylethyl) caged O-phosphorothioylserine, -threonine, and -tyrosine derivatives is reported. These amino acid building blocks can be directly incorporated into peptides by Fmoc-based solid phase synthesis as their pentafluorophenyl esters or as symmetric anhydrides. Upon irradiation with UV light, the thiophosphate group, representing a hydrolysis resistant phosphate analog, is revealed.     

Gas-liquid chromatographic analysis of protein amino acids as N-isobutyloxycarbonylamino acid methyl esters.

A practical method for the quantitative determination of protein amino acids by gas-liquid chromatography (GLC) is described. All of the common protein amino acids except arginine can be readily converted into their N-isobutyloxycarbonyl (N-isoBOC) methyl ester derivatives by a simple procedure involving isobutyloxycarbonylation with isobutyl chloroformate in aqueous medium, followed by methylation with diazomethane. Arginine was converted into N-isoBOC ornithine methyl ester by treatment with arginase, followed by the above derivatization procedure. The resulting N-isoBOC methyl esters of the amino acids have good GLC properties. Complete resolution of the derivatives of 20 protein amino acids was achieved by using a dual-column system consisting of a 0.65% Poly-A-101A column and a 0.70% FFAP-Poly-A-101A (1:1, w/w) column. The reproducibility of response was found to be good for derivatives carried through the entire chemical and chromatographic procedure. The calibration graphs were linear and showed no statistical bias. The results of recovery experiments with synthetic mixtures containing known amounts of the amino acids were satisfactory, the recoveries ranging from 94.3 to 106.2%.     

Enantiomer separation of amino acids as their N-alkyloxycarbonyl alkylamide derivatives by chiral phase capillary GC

The enantiomers of amino acids were first converted into N-alkyloxycarbonyl 2,2,2-trifluoroethyl esters, and then into N-alkyloxycarbonyl alkylamides by nucleophilic substitution of the ester group with amines. The first reaction proceeds instantaneously, while the second substitution occurs smoothly with n-propylamine and isobutylamine. The final derivatives were produced for separation on a capillary column coated with Chirasil-Val by GC. Pro, which is difficult to separate completely as its N-perfluoroacyl alkyl ester derivative, showed complete separation of the enantiomeric pair. All amino acids examined in this study showed an increased separation factor.