Synthesis of Fmoc-amino acid chlorides assisted by ultrasonication,a rapid approach

Summary The chloride derivatives of all common Fmoc-amino acids lacking polar side chains as well as a number of benzyl based side chain protection have been prepared using the corresponding amino acid and thionyl chloride assisted by ultrasound has been described. The protocol is simple, efficient and rapid. All the acid chlorides prepared have been obtained in good yield and purity.     

Rapid and Efficient Synthesis of Peptides Containing α,α-dialkylamino acids employing KOBt

Summary Several Fmoc-α,α-dialkylamino acids and their acid chlorides have been prepared, isolated and characterised. The synthesis of peptides containing sterically hindered dialkylamino acids has been accomplished using acid chloride/KOBt in dichloromethane. The yields as well as the purity of the peptides were satistactory.     

Synthetic Studies on the Acyclic Nucleosides of 5-Substituted-6-Azauracils

Abstract

A number of acyclic nucleosides have been prepared. 5-substituted-6-azauracils were persilylated with HMDS and then alkylated with aliphatic side chains e.g., (2-acetoxyethoxy)methyl bromide, 1,3-dibenzuloxy-2-chloromethoxypropane, (1-benzyloxy-3-phthalimido-2-propoxy)methyl chloride, and 1-benzyloxy-2-chloro-methoxybutane to yield protected acyclic nucleosides which were deprotected by Lewis acid or palladium to give various 6-azauracil acyclonucleosides.     

Selective acylation of cholic acid derivatives with multiple methacrylate groups

Bile acids in the family of steroid compounds can be chemically modified for biochemical and other applications. Derivatives of cholic acid with multiple methacrylate groups can be prepared by the use of methacrylic acid, methacryloyl chloride and methacryloyl anhydride as the acylating agents. The hydroxyl groups of cholic acid methyl ester and cholic acid ethylene glycol ester have been selectively acylated by changing the acylating agents and the number of substitutions have been varied by changing the amount of the acylating agents used. In the acylation reactions with methacryloyl chloride, the reactivity of secondary hydroxyl groups on the steroid skeleton of cholic acid derivatives follows the order of C3>C12>C7.     

The preparation of amino bile acid derivatives

Bile acid derivatives have been prepared by reaction of their mixed anhydrides with diaminoethane. The new compounds have side chains modified by amide bond formation which extends the side chain by two carbon atoms and terminates in a primary amino group. Important properties of the new compounds are their solubility in acid, and their ability to act as nucleophiles in the carbodiimide reaction or mixed anhydride reaction. The derivatives have been used to prepare high molar ratio immunogens with bovine serum albumin and to prepare I labelled ligands for radioimmunoassay     

Depigmenting activity of new kojic acid derivative obtained as a side product in the synthesis of cinnamate of kojic acid

We synthesized cinnamate derivatives of kojic acid for use as depigmenting agents by various esterification methods. The cinnamate of 5-position of kojic acid (6) was obtained by EDC coupling, DCC coupling, acid chloride, and mixed anhydride methods. To obtain the cinnamate of the 2-position of kojic acid (7), we carried out the nucleophilic addition of the potassium salt of cinnamic acid to kojyl chloride. In this reaction, we discovered the occurrence of a side reaction and identified the structure of the side product thus formed. We evaluated the depigmenting activities of both the side product and the cinnamate derivatives of kojic acid. Interestingly, the side product (11) showed more potent depigmenting activity (IC(50)=23.51μM) than compound 7 (IC(50)>100μM) which is the mother compound of the side product. However, it has no tyrosinase inhibitory activity. Compound 6, the cinnamate of 5-position of kojic acid, also showed moderate depigmenting activity (IC(50)=46.64μM) without tyrosinase inhibitory activity. Production of this side product (11) may have originated from the proton exchange between the potassium salt of cinnamic acid and kojyl chloride. We then efficiently reduced the yield of the side product by controlling the equilibrium of the potassium salt of cinnamic acid. The addition of cinnamic acid greatly reduced the amount of the side product produced.     

Synthesis of 4beta-amido and 4beta-sulphonamido analogues of podophyllotoxin as potential antitumour agents

The new 4beta-amido analogues of podophyllotoxin or 4'-O-demethylepipodophyllotoxin have been prepared either by the coupling of 4beta-amino podophyllotoxin or 4beta-amino-4'-O-demethyl epipodophyllotoxin with the corresponding acids in presence of DCC in dichloromethane or by treating the appropriate acid chloride or sulphonyl chloride in presence of Et(3)N. These 4beta-amido and 4beta-sulphonamido derivatives of podophyllotoxin have been evaluated for their cytotoxicity against six human cancer cell lines. Some of these analogues have shown promising anticancer activity.     

Alpha-azido acids for direct use in solid-phase peptide synthesis.

Several new alpha-azido acids have been synthesized and their use in solid-phase peptide synthesis has been demonstrated. The azido group allows for high activation of the carboxyl group as an acid chloride without formation of byproducts and with no detectable racemization. An analog of Leu-enkephalin has been prepared and tested in the mouse vas deferens and guinea pig ileum bioassays: it displays moderate activity at the delta-opiod receptor.     

Protein immobilization on poly-(N-substituted) acrylamides

A series of water-soluble polymers containing side chains derived from N-acryloyl-β-alanine, N-ethylacrylamide and N-[3-(N′,N′,N′-trimethylammonio)propyl] acylamide chloride has been prepared and characterized. A related series of insoluble gels was also prepared. Protein may be attached to these materials by means of amide bond formation between carboxyl groups on the polymers and amino groups of the protein; the preparation and characterization of conjugates formed with α-chymotrypsin are described. Polymers bearing negatively or positively charged side chains are attached to this enzyme at only a single amino acid and the integrity of the active site is largely preserved in these systems. The corresponding gels are not able to bind as much enzyme as are the soluble polymers and bound enzyme is less active in these cases.     

Preparation of N-9-fluorenylmethoxycarbonylamino acid chlorides from mixed anhydrides by the action of hydrogen chloride.

N-9-Fluorenylmethoxycarbonyl-(Fmoc) amino-acid chlorides have been prepared by reaction of hydrogen chloride on purified mixed Fmoc-amino acid-monoalkyl carbonic acid anhydrides in dichloromethane. The products partially undergo subsequent conversion to the corresponding esters due to the presence of the liberated alcohol, the extent depending on the nature of the alkyl group. Esterification occurred to 5-20% when the alkyl group was isopropyl. Anhydrides of monoisopropenyl carbonic acid which liberate acetone instead of an alcohol gave products uncontaminated with ester. The three components in a reaction mixture could be determined as the reaction progressed by normal phase high-performance liquid chromatography of aliquots, which had been freed of excess hydrogen chloride, on a mu Porasil (underivatized silica) column using tert.-butanol-hexane (1.5:98.5) as solvent.     

Rapid and Efficient Synthesis of Peptides Containing α,α-dialkylamino acids employing KOBt

Several Fmoc-,-dialkylamino acids and their acid chlorides have been prepared, isolated and characterised. The synthesis of peptides containing sterically hindered dialkylamino acids has been accomplished using acid chloride/KOBt in dichloromethane. The yields as well as the purity of the peptides were satisfactory.     

Synthesis of Some New Halogenated N-Thiazolyl Substituted 2-Mercapto Benzoic Acid Amides and Their Use as Possible Fungicides

One hundred and four acid amido compounds have been prepared by condensing 2-mercapto benzoic acid and its 5-bromo, 3, 5-dibromo and 3, 5-dichloro derivatives with 2-amino-4-substituted thiazoles and their 5-halogenated derivatives via the acid chloride. All these compounds have been tested against Helminthosporium. Four of the compounds were tested for their antibacterial, antihelminthic activities and also for their hypoglycemic activity.     

Gastric acid secretion in the lizard. Ionic requirements and effects of inhibitors.

1. The effects of ion substitution and various inhibitors on the transmucosal potential, short circuit current, mucosal resistance and acid secretion of the lizard gastric mucosa, incubated in an Ussing chamber, have been determined. 2. Ion substitution experiments indicate that the serosal potential step consists of a combined C1- and K+ diffusion potential, and that the mucosal potential step is Na+ dependent and behaves primarily as a Na+ diffusion potential. 3. Experiments with ouabain indicate that the major (Na+, K+)-ATPase activity responsible for maintenance of cation gradients is located on the serosal side of the mucosal cells, and that this pump activity is non-electrogenic. 4. Experiments with amiloride indicate that a passive sodium influx on the mucosal side is essential for the maintenance of the transmucosal potential and short circuit current. 5. Acid secretion requires the presence of sodium and chloride on the serosal side and the maintenance of a high intracellular potassium level through the (Na+, K+)-ATPase system. 6. The effects of acetazolamide and thiocyanate are compatible with an involvement of carbonic anhydrase and anion-dependent ATPase in acid secretion. 7. Upon initiation of acid secretion the serosal membrane permeability for chloride increases and that for potassium decreases.     

2-alpha-(N-dansyl-4-aminophenylthio)-N-acetyl-9-O-acetylneuraminic acid. A new specific and highly sensitive substrate in sialate-O-acetylesterase assay.

2-alpha-(N-Dansyl-4-aminophenylthio)-N-acetyl-9-O-acetylneuraminic acid (10) was prepared as a new specific and highly sensitively detectable sialate-9-O-acetyl-esterase substrate. It is built up from a sialidase-stable aminophenyl-alpha-thioketoside of N-acetylneuraminic acid. By labeling this thioketoside with dansyl chloride a fluorescent neuraminic acid derivative was prepared which allows determinations down to the picomol range. Regioselective acetylation with trimethylorthoacetate results in the corresponding 9-O-acetyl derivative. After incubation with esterase from bovine brain the hydrolysis products were separated on a HPLC column and fluorimetrically detected at 334 nm excitation and 564 nm emission. The Km value of 2.5 mM was in the range between the values of the completely unspecific methylumbelliferyl acetate and the less sensitively detectable N-acetyl-9-O-acetylneuraminic acid which have been used up to now as standard substrates.     

Side-dependent inhibition of a prokaryotic ClC by DIDS

The x-ray structure of the Escherichia coli chloride/proton antiporter ClC-ec1 provides a structural paradigm for the widespread and diverse ClC family of chloride channels and transporters. To maximize the usefulness of this paradigm, it is important to directly relate structure to function via studies of ClC-ec1 itself; however, few functional studies of this protein have been performed. In an endeavor to develop new tools for functional analysis of ClC-ec1, we have discovered that this transporter is inhibited by the stilbenedisulfonate 4,4-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS). In planar lipid bilayers, DIDS inhibits ClC-ec1 activity reversibly, with an apparent affinity in the micromolar range. Since ClC-ec1 is randomly oriented in the bilayers, ascertaining whether DIDS inhibits from the intracellular or extracellular side required an indirect approach. Using the ClC-ec1 structure as a guide, we designed a strategy in which modification of Y445C was monitored in conjunction with inhibition by DIDS. We found that DIDS inhibits transporters specifically from the intracellular side. Transporters with their extracellular side exposed to DIDS function normally, maintaining stoichiometric proton/chloride antiport over a wide range of proton and chloride concentrations. The side-dependent nature of DIDS inhibition will be useful for generating "functionally oriented" preparations of ClC-ec1, in which DIDS is used to silence transporters in one orientation but not the other.     

The synthesis and P388 cytotoxicity of mycalazol 11 and related 5-acyl-2-hydroxymethylpyrroles

We report a general method for the synthesis of mycalazol 11 and some related 5-acyl-2-hydroxymethylpyrroles using a Stille coupling of 5-(tri-n-butylstannyl)pyrrole-2-carboxaldehyde with an acid chloride as the key step. The newly prepared 5-acyl-2-hydroxymethylpyrroles 5-7, together with the 5-carboxamido-2-hydroxymethylpyrrole 10, have been assayed for in vitro cytotoxicity against the P388 murine leukemia cell line.     

Azobenzene-modified poly(l-glutamic acid) (AZOPLGA): its conformational and photodynamic properties

Azobenzene-modified poly(l-glutamic acid) (AZOPLGA) polymers with 22 and 35 mol % of azo chromophores in the side chains have been synthesized by condensing 4-methoxy-4'-aminoazobenzene and poly(l-glutamic acid). These polymers have been characterized by NMR, FT-IR, and UV-visible spectroscopic techniques. The conformational features of the polymer backbone chains in the films that were cast from the polymer solutions prepared in different solvents have been investigated by circular dichroism spectroscopy. Experimental data suggested that the thermal cis-trans relaxation and photoinduced birefringence, which are related to the azo chromophores in the side chains of polymer, are not affected by the conformations of polymer backbones. However, the modulations of the surface relief gratings, the result of photoinduced mass transport process, recorded on these polymers are sensitive to polymer main chain conformation, as well as the degree of functionalization.     

Synthesis of steroidal 17 β-carboxamide derivatives

Several 17 β-carboxamide derivatives of natural and fluorinated glucocorticoids have been synthesized. The 17 β-carboxylic derivatives were obtained by periodic acid oxidation of their side chains. They were then activated by N-hydroxybenzotriazole (HOBT) and coupled to several primary amines. Using this method eleven 17 β-carboxamide derivatives have been prepared in good yields.     

Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease

Intact ribonucleic acid (RNA) has been prepared from tissues rich in ribonuclease such as the rat pancreas by efficient homogenization in a 4 M solution of the potent protein denaturant guanidinium thiocyanate plus 0.1 M 2-mercaptoethanol to break protein disulfide bonds. The RNA was isolated free of protein by ethanol precipitation or by sedimentation through cesium chloride. Rat pancreas RNA obtained by these means has been used as a source for the purification of alpha-amylase messenger ribonucleic acid.     

Methanolysis of sphingomyelin. Toward an epimerization-free methodology for the preparation of D-erythro-sphingosylphosphocholine

It is well known that acid hydrolysis of natural sphingomyelin in aqueous methanol or 1-butanol at refluxing temperature is accompanied by epimerization at the C-3 position of the long-chain base. An improved procedure for the hydrolysis of commercially available, naturally occurring sphingomyelin is described. Prolonged exposure (3;-4 days) of sphingomyelin to freshly prepared 0.5 M anhydrous methanolic hydrogen chloride (generated by trapping the gas evolved from the reaction of concentrated sulfuric acid with solid sodium chloride in anhydrous methanol) at 50 degrees C resulted in cleavage of the amide side chain. The extent of epimerization of the allylic alcohol stereocenter was quantified by integration of the C-5 signal of the (13)C nuclear magnetic resonance spectrum of lysosphingomyelin.The method described here is superior to the traditional acid hydrolysis methods because it provides the product as a approximately 10:1 ratio of d-erythro/l-threo epimers; in contrast, a ratio of approximately 1. 3:1 was obtained by the previous methods. We also report that use of dichloromethane as a cosolvent with N,N-dimethylformamide in the reaction of lysosphingomyelin with an activated fatty acid reduced the time required for completion of the N-acylation reaction.