Major Side Product Research Articles

Solid phase peptide synthesis of human endothelin precursor peptide using two‐step hard acid deprotection/cleavage methods

Syntheses are described for the putative human and porcine biosynthetic precursors (hET-38 and pET-39) of endothelin, with the sequence previously deduced from human- and porcine-cDNA coding for preproendothelin. The Boc based solid phase synthetic method was applied, followed by weak hard acid, trimethylsilyl bromide, cleavage. The peptide removal from the resin was optimally accomplished with hydrogen fluoride. Disulfide bridges were formed by air-oxidation, and the linkage modes determined by enzymic (Endoproteinase Asp-N) digestion and HPLC. Five additional C-terminally elongated endothelin homologs were also synthesized. For alternative synthesis of pET-39, the use of trimethylsilyl trifluoromethanesulfonate for the removal of peptide from the resin generated a major side product, which was characterized. hET-38 was found to be less effective in vitro, when compared to endothelin. The vasoconstrictor activity in vitro of other related peptides was comparable to that of hET-38.

Expression of native rabbit light meromyosin in Escherichia coli: Observation of a powerful internal translation initiation site

The cDNA-sequence coding for rabbit skeletal muscle light meromyosin (LMM) was placed under the control of the lambda promoter (P L) of an Escherichia coli expression vector. The resulting plasmid pEXLMM74 expressed non-fused rabbit skeletal muscle LMM with yields ranging from 1 to 5% of the total proteins of E. coli. This LMM was specifically recognized by polyclonal antibodies raised against chicken pectoralis muscle myosin. It could be highly enriched from E. coli extracts by using two cycles of high and low ionic strength buffer. The partially purified protein contained a major side-product, with a calculated molecular mass of 59 kilodaltons, that is produced by translation initiation from a site in the coding region of LMM. After deletion of the translation initiation site derived from the expression plasmid, only the 59 kilodalton protein is expressed in E. coli from the resulting plasmid pEXLMM59. Both the 74 and 59 kilodalton proteins were shown to form paracrystals. They were studied by electron microscopy using negative staining and were found to show characteristic striations with an axial periodicity of about 43 nm. By circular dichroism measurement we showed that the purified 59 kilodalton protein is folded mostly as an alpha-helix.

Mechanism of ruthenium-catalyzed linear dimerization of acrylonitrile: a kinetic study

A detailed kinetic study of the RuCl 2(AN) 3-catalyzed linear dimerization of acrylonitrile reveals that the reaction is first order in Ru and H 2, and zero order in acrylonitrile. A reaction model is proposed which is consistent with the observed kinetic behavior and product distribution data, including our previous deuterium-labelling study [4]. It is visualized that two equivalents of base are first utilized to facilitate the generation of the active catalyst, a zero-valent Ru complex. The Ru(0) complex interacts reversibly with H 2 to give a ruthenium dihydride, which then enters into the dimer-forming catalytic cycle. Two subsequent acrylonitrile insertions, the first into one of the Ru—H bonds followed by another into the Ru—C bond formed in the first insertion, and a β-hydride elimination produce the observed linear dimer. The major side product, propionitrile, is formed by a competing reductive elimination from the hydrido(α- or β-cyanoethyl)ruthenium intermediate. Detailed product distribution analysis reveals the presence of one base ligand in the coordination sphere of the Ru. The coordination of base favors the production of saturated dimer vs. unsaturated dimer, but has no effect on the propionitrile: linear dimer ratio.

Preparation of a glycosyl donor suitable for synthesis of glycoprotein “core” oligosaccharides

2- O-Acetyl-3- O-allyl-4- O-benzyl-6- O-tert-butyldiphenylsilyl- d-glucopyranosyl chloride ( 14), a glycosyl donor suitable for synthesis of oligosaccharides corresponding to the N-glycoprotein saccharide “core”, was synthesized by an efficient, six-stage route from 3,4,6-tri- O-acetyl-1,2- O-[1-( exo-ethoxy)ethylidene]-α- d-glucopyranose. Silver triflate-promoted coupling of 14 with benzyl 2-acetamido-3,6-di- O-benzyl-2-deoxy-α- d-glucopyranoside gave a protected β- d-(1→4)-linked disaccharide in 38% yield, but a major side-reaction also occurred. When the tert-butyldiphenylsilyl group was quantitatively removed from 14 prior to the coupling reaction, and replaced afterwards, the yield in the glycosidation was increased to 55%, and major side-products were avoided.

4-Chloromercurioandrosta-4,6-diene-3,17-dione: preparation, X-ray structure determination, and potential utility

The novel steroid 4-chloromercurioandrosta-4,6-diene-3,17-dione (2) was obtained as a major side-product during conversions of 6βbromoandrost-4-ene-3,17-dione (1b) into its 6β-fluoro derivative (1a) using HF(pyridine)n and mercuric oxide. Compound (2) was successfully converted into various 4-substituted steroids (using standard vinyl-mercury methodology), including the disteroidal mercury compound (3c). Crystals of the acetone solvate of (2). 2(HgClC19H40O2)· acetone, were monoclinic, space group P21,Z= 2 with dimensions a= 7.295(8),b= 24.12(1),c= 12.215(12)A,β= 108.0(1)°. The two independent molecules have equivalent structures with the mercury atoms strongly bound to a chlorine atom and to carbon atom C(4) of the steroid. The acetone molecule forms a weak bridge between the two mercury atoms.

Addition of aromatic compounds to oleic acid catalyzed by heterogeneous acid catalysts

AbstractThe addition of aromatic compounds to the double bond of oleic acid was studied using solid acid catalysts. For example, in the presence of an acid clay catalyst (bentonite), phenol reacted with oleic acid to yield 96% of an alkylphenol addition product. When toluene was used as the aromatic reactant, however, the yield of alkylbenzene addition product was less than 2%. In this instance, the major reactions observed were elaidinization and migration of the double bond of oleic acid. The addition of phenol to oleic acid in greater than 95% yield also was accomplished with the use of a sulfonic acid ion‐exchange resin catalyst. This same catalyst also catalyzed the addition of toluene and benzene to oleic acid to yield 82% and 22%, respectively, of alkylbenzene‐type addition products. In the latter instances, the major side product formed was γ‐stearolactone. Capillary gas chromatographic (GC) analyses of the alkylbenzene addition products obtained showed them to be mixtures of positional isomers. The isomer distributions were subsequently determined by GC‐mass spectrometry (MS).

Use of alcohols as sources of hydrogen in the transformation of co-olefin mixtures

A study has been made of the catalytic activities of various ruthenium complexes in the synthesis of ketones from olefin-carbon monoxide mixtures and an alcohol as source of hydrogen: major side products were shown to be the alkyl propionates and ethane. The effects of base, concentration of catalyst (for RuCl3- and Ru3(CO)12- based systems), pressure, temperature, CO/olefin ratio and the water content of the alcohol have been examined.

Reaction of tetramethyl‐1,2‐dioxetane with triphenylphosphine: Activation parameters for the formation of 2,2‐dihydro‐4,4,5,5‐tetramethyl‐2,2,2‐triphenyl‐1,3,2‐dioxaphospholane

AbstractThe reaction of tetramethyl‐1,2‐dioxetane (1) and triphenylphosphine (2) in benzene‐d6 produced 2,2‐dihydro‐4,4,5,5‐tetramethyl‐2,2,2‐triphenyl‐1,3,2‐dioxaphospholane (3) in ˜90% yield over the temperature range of 6–60°. Pinacolone and triphenylphosphine oxide (4) were the major side products [additionally acetone (from thermolysis of 1) and tetramethyloxirane (5) were noted at the higher temperatures]. Thermal decomposition of 3 produced only 4 and 5. Kinetic studies were carried out by the chemiluminescence method. The rate of phosphorane was found to be first order with respect to each reagent. The activation parameters for the reaction of 1 and 2 were: Ea ≅ 9.8 ± 0.6 kcal/mole; ΔS≠ = −28 eu; k30° = 1.8 m−1sec−1 (range = 10–60°). Preliminary results for the reaction of 1 and tris (p‐chlorophenyl)phosphine were: Ea ˜ 11 kcal/mole, ΔS≠ = −24 eu, k30° = 1.3 M−1sec−1 while those for the reaction of 1 and tris(p‐anisyl)phosphine were: Ea ˜ 8.6 kcal/mole, ΔS≠ = −29 eu, k30° = 4.9 M−1 sec−1.

Cyanamide mediated syntheses of peptides containing histidine and hydrophobic amino acids.

Using the model of a primitive earth evaporation pond, the synthesis of three histidyl peptides in yields up to 11% was demonstrated when aqueous solutions of histidine, leucine, ATP, cyanamide, and MgCl2 were evaporated and heated for 24 h at 80 degrees C. In addition, peptides were formed in yields of up to 56%, 35%, and 21%, respectively for phenylalanine, leucine, and alanine when aqueous solutions of the appropriate amino acid were evaporated and heated with cyanamide and one or more of the following components: ATP, AMP, 4-amino-5-imidazole carboxamide, or MgCl2. The greatest peptide yield occurred at pH 3. But peptide formation was demonstrated for a system of Leu, cyanamide, and MgCl2 adjusted to pH 7 with NH4OH. Peptide synthesis was also studied in the presence of CaCl2, ZnCl2, different adenosine nucleotides, and UTP to compare their effects on peptide synthesis. The optimum conditions for cyanamide mediated peptide synthesis were also studied in terms of pH, reaction time, reaction temperature, and cyanamide concentration. The major side product in nearly all reactions studied appears to be an amino acid-cyanamide adduct. Peptides were analyzed and identified by thin layer chromatography, acid hydrolysis, and enzymatic degradation.

Structures and mechanisms of formation of the major side products from the preparation of 4,4-dimethylcyclohex-2-enone by the potassium hydroxide promoted annelation of methyl vinyl ketone and isobutyraldehyde

Structures and mechanisms of formation of the major side products from the preparation of 4,4-dimethylcyclohex-2-enone by the potassium hydroxide promoted annelation of methyl vinyl ketone and isobutyraldehyde

ChemInform Abstract: SYNTHESIS OF ISOCYANIC ACID FROM NITRIC OXIDE OVER PALLADIUM AND IRIDIUM CATALYSTS

AbstractIn einem Quarzreaktor wird die Umsetzung eines Gemisches von 0,3% Stickstoffmonoxid, 5% Kohlenmonoxid und 0,5% Wasserstoff in einem Heliumstrom an einem Fixbettkatalysator im Temperaturbereich 280 ‐ 450° untersucht.

Synthesis of Isocyanic Acid from Nitric Oxide over Palladium and Iridium Catalysts

A novel and convenient synthesis of isocyanic acid (HNCO) and its trimer isocyanuric acid has been obtained by the conversion of nitric oxide (NO). Nitric oxide, carbon monoxide, and hydrogen mixed in helium at the level of 0.3 to 5 percent reacted over iridium and palladium catalysts at 280 degrees to 450 degrees C to form HNCO in 60 to 75 percent yield. Major side products were ammonium cyanate at low temperature and ammonia or nitrogen at high temperature. Total conversion of nitrogen from NO to NCO reached 73 percent over palladium and 86 percent over iridium.

The oxidation of isobutene to p‐xylene over bismuth oxide‐based catalysts

AbstractStudies have been carried out of the optimisation of a bismuth oxide based catalyst for the oxidation of isobutene to p‐xylene. Using a Bi2O3‐P2O5 (Bi:P = 2) catalyst located sequentially before a 3% Pt/Al2O3 catalyst, up to 60% selectivity can be observed at 50% conversion. The kinetics of the oxidation of isobutene over the Bi2O3‐P2O5 catalyst have been measured for all major products. The results support a mechanism involving allylic oxidation of isobutene. Dimethylhexadiene and p‐xylene are suggested to originate from reaction between two adsorbed isobutene molecules and monatomic oxygen species adsorbed at different sites. A major side product, methacrolein, appears to originate from reaction between two adsorbed isobutene molecules and a diatomic oxygen species.

The formation of grignard compounds—V

Abstract Analysis of product formation reveals that in the reaction of iodomethane and magnesium in anyl phenyl ether ethane and 1-butane are the major side products. The latter product is formed mainly by addition of methyl radicals to the terminal CH2 group of allyl phenyl ehter, followed by β-scission under extrusion of a phenoxy radical. This mechanism f is confirmed by an analysis of the CIDNP spectra during the reaction; CH3 radicals escaping from CH 3 CH 3 pairs and adding to allyl phenyl ether give rise to a very clear example of a second order CIDNP spectrum in the product 1-butane, while polarization in CH 3 C 6 H 5 O-CH-CHCH 2 pairs, formed after α-H abstraction from allyl phenyl ether, leads to net effects in the products ethane (E) and 1-methylallyl phenyl ether (A).

The carboxylic acid groups of bovine luteinizing hormone. The effects of their modification on receptor site binding and subunit-subunit interaction.

The modification of the carboxyl groups of the subunits of bovine luteinizing hormone to neutral derivatives by carbodiimide-mediated coupling with glycine methyl ester has been studied. The modified alpha subunit, which has 8 residues of glycine methyl ester incorporated, will no longer recombine with native beta (hormone-specific) subunit, but the modified beta subunit, with 6 to 7 glycine methyl esters incorporated, will recombine with native alpha to yield a partially active hormone. Derivatization of the intact hormone results in dissociation to subunits together with formation of a major side product which is covalently cross-linked. Significant cross-linked product was not obtained during modification of individual subunits, thus indicating an orientation between an activated carboxyl group(s) and a nucleophile(s) in the intact hormone which favors coupling. Separation of subunits from the derivatized, noncross-linked fraction by countercurrent distribution reveals a heterogeneous preparation of the modified alpha subunit which also will not recombine with either a native or modified beta subunit. The beta subunit from the modified intact hormone was indistinguishable from the modified isolated beta subunit in amino acid composition and in ability to recombine with native alpha subunit. The results are consonant with data from this and other laboratories in which various modifications of the alpha chain, the subunit common to the glycoproteins, more seriously affect recombination than similar modifications of the beta subunits. The number of carboxyl groups modified in each subunit is compatible with but not in total agreement with assignments of amides reported from sequence studies.

The nitric oxide catalyzed positional isomerization of 3‐methylene‐1,5,5‐trimethylcyclohexene and the stabilization energy in the 3‐methylenecyclohexenyl radical

AbstractThe gas phase, nitric oxide catalyzed positional isomerization of 3‐methylene‐1,5,5‐trimethylcyclohexene (MTC) into 1,3,5,5‐tetramethyl‐1,3‐cyclohexadiene (TECD) has been studied for temperatures ranging between 296° and 425°C. The major reaction was first order with respect to nitric oxide and to MTC.The major side product, mesitylene, usually amounted to less than 10% of the TECD isomer formed. Only at high temperatures and large conversions has up to 20% been observed.Conditioned pyrex or quartz vessels coated with KCl have been used. The nitric oxide catalyzed isomerization is apparently a homogeneous process, as demonstrated by the insensitivity of the observed rate constants towards a 15‐fold increase in the surface to volume ratio of the reaction vessels. However, a residual, presumably heterogeneous, thermal isomerization of the starting material could not be eliminated. Good mass balances were obtained for both NO and hydrocarbons.After correcting for the thermally induced conversion the observed rate constants for the nitric oxide catalyzed isomerization yield log k1 (1 mole−1 sec−1) = (10.7 ± 0.2) – (37.3 ± 0.9)/θ where θ is 2.303 × 10−3 RT (kcal mole−1). Plotting log k1 versus the ratio of the starting materials (MTC/NO)0 it was found that for temperatures ≥ 365°C the rate constants were systematically too high.Using extrapolated values for the higher temperature range yields the more reliable corrected Arrhenius equation log k = 8.6 – 31.7/θ. The reaction mechanism is outlined and the implications with respect to the stabilization energy generated in the MTCċ radical intermediate and the activation energy of the backreaction MTCċ + HNO are discussed.Using for the activation energy E−1 of the backreaction (Rċ + HNO) a literature value of 9.2 ± 0.9 kcal mole−1 reported for the cyclohexadiene1,3system, this yields 23.4 ± 2 kcal mole−1 for the stabilization energy in the methylenecyclohexenyl radical, which is to be compared with the corresponding values for the allyl (10.2 ± 1.4), methallyl (12.6 ± 1) pentadienyl (15.4 ± 1) and cyclohexadienyl (24.6 ± 0.7) radicals.The pre‐exponential factor agrees well with the value of (8.4 ± 0.2) reported by Shaw and co‐workers for the similar reaction of NO with 1,3‐cyclohexadiene. It is noteworthy that HNO, acting as sole hydrogen donor in the system, is surprisingly stable under the reaction conditions used. Nitrous oxide, HCN, H2O and N2 are observed in the product mixture of experiments carried out to high conversions at higher temperatures.