Autoxidation of methyl linoleate. Separation and analysis of isomeric mixtures of methyl linoleate hydroperoxides and methyl hydroxylinoleates

The mixture of conjugated diene hydroperoxide isomers obtained from autoxidation of methyl linoleate was separated by high performance liquid chromatography (HPLC). Four major isomers were obtained from adsorption chromatography and identified as the 9 and 13 positional isomers having thetrans-trans andcis-trans configurations. The latter geometrical isomers have thetrans double bond adjacent to the hydroperoxide group. The hydroxy compounds (methyl hydroxylinoleates) obtained from the hydroperoxides by NaBH₄ reduction were similarly separated but with improved resolution. This is the first instance of the complete separation of these compounds and provides a rapid method for their analysis. Unlike adsorption chromatography, reversed-phase chromatography separates the mixtures only according to the geometrical isomerism of the double bonds and not according to the position of the hydroxy or hydroperoxide function.     

Synthesis of γ‐Fluoro‐α, β‐unsaturated Carboxylic Esters from Saturated α‐Fluoro Aldehydes

γ‐Fluoro‐α, β‐unsaturated carboxylic esters 7a, 7b and 7d and 4‐fluoro‐4‐phenylbut‐3‐enoic ester ( 8 ) are obtained by two alternative pathways from 2‐fluoro aldehydes 5a—d , either by Horner—Wadsworth—Emmons reaction or by Wittig reaction. The aldehydes 5a—d are prepared by Swern oxidation of the corresponding fluorohydrins 4a—d . These are available from α‐olefins by bromofluorination, bromineby‐acetate replacement and subsequent hydrolysis.     

Cyclisierung von β‐[(1, 4‐Chinon‐2‐yl)methyl]‐enaminon‐Derivaten

Starting from 2,5‐bisbenzyloxy‐4‐methyl‐benzaldehyd 2‐(3‐amino‐2‐acetyl‐but‐2‐enyl)‐5‐methyl[1,4]benzoquinones 9a—e are synthesized as model compounds in order to study the ambident reactivity of enaminones and quinones. Spontaneus cyclization of 9b—e in ethanol or acetic acid to 1‐aza‐spiro[4,5]deca‐2,7‐dien‐6,9‐dion 20b—e in good yield is observed. 6‐Hydroxy‐3‐acetyl‐quinoline 21 is obtained from 9a . In one case ( 9c ) 2‐acetyl‐3‐benzylamino‐7‐hydroxy‐naphthalene ( 17 ) was formed as by‐product. 9d in alkoholic perchloric acid leads to quinolinium salt 18 in low yield.     

β‐Aminopeptidase‐Catalyzed Biotransformations of β2‐Dipeptides: Kinetic Resolution and Enzymatic Coupling

We have previously shown that the β‐aminopeptidases BapA from Sphingosinicella xenopeptidilytica and DmpA from Ochrobactrum anthropi can catalyze reactions with non‐natural β³‐peptides and β³‐amino acid amides. Here we report that these exceptional enzymes are also able to utilize synthetic dipeptides with N‐terminal β²‐amino acid residues as substrates under aqueous conditions. The suitability of a β²‐peptide as a substrate for BapA or DmpA was strongly dependent on the size of the C_α substituent of the N‐terminal β²‐amino acid. BapA was shown to convert a diastereomeric mixture of the β²‐peptide H‐β²hPhe‐β²hAla‐OH, but did not act on diastereomerically pure β²,β³‐dipeptides containing an N‐terminal β²‐homoalanine. In contrast, DmpA was only active with the latter dipeptides as substrates. BapA‐catalyzed transformation of the diastereomeric mixture of H‐β²hPhe‐β²hAla‐OH proceeded along two highly S‐enantioselective reaction routes, one leading to substrate hydrolysis and the other to the synthesis of coupling products. The synthetic route predominated even at neutral pH. A rise in pH of three log units shifted the synthesis‐to‐hydrolysis ratio (v_S/v_H) further towards peptide formation. Because the equilibrium of the reaction lies on the side of hydrolysis, prolonged incubation resulted in the cleavage of all peptides that carried an N‐terminal β‐amino acid of S configuration. After completion of the enzymatic reaction, only the S enantiomer of β²‐homophenylalanine was detected (ee>99 % for H‐(S)‐β²‐hPhe‐OH, E>500); this confirmed the high enantioselectivity of the reaction. Our findings suggest interesting new applications of the enzymes BapA and DmpA for the production of enantiopure β²‐amino acids and the enantioselective coupling of N‐terminal β²‐amino acids to peptides.     

Cationic copolymers of α,β‐poly‐(N‐2‐hydroxyethyl)‐DL‐aspartamide (PHEA) and α,β‐polyasparthylhydrazide (PAHy): synthesis and characterization

In the present study the derivatization of two water‐soluble synthetic polymers, α,β‐poly(N‐2‐hydroxyethyl)‐DL ‐aspartamide (PHEA) and α,β‐polyasparthylhydrazide (PAHy), with glycidyltrimethylammonium chloride (GTA) is described. This reaction permits the introduction of positive charges in the macromolecular chains of PHEA and PAHy in order to make easier the electrostatic interaction with DNA. Different parameters affect the reaction of derivatization, such as GTA concentration and reaction time. PHEA reacts partially and slowly with GTA; on the contrary the reaction of PAHy with GTA is more rapid and extensive. The derivatization of PHEA and PAHy with GTA is a convenient method to introduce positive groups in their chains and it permits the preparation of interpolyelectrolyte complexes with DNA. © 2000 Society of Chemical Industry     

Kinetic Resolution of Aliphatic β‐Amino Acid Amides by β‐Aminopeptidases

Access to enantiopure β‐amino acids : β‐Aminopeptidases are hydrolases that possess the unique ability to cleave N‐terminal β‐amino acids from peptides and amides. Hydrolysis of racemic β‐amino acid amides catalyzed by these enzymes displays enantioselectivity with strong preference for substrates with the L ‐configuration, and gives access to various aliphatic β‐amino acids of high enantiopurity.

     

An Efficient and Simple Procedure for the Preparation of α‐Keto‐β‐lactams

The methods for the preparation of α‐keto‐β‐lactams described in the literature are generally specific for a particular target molecule and lack generality. A short route to several of these compounds has been developed and is described in this communication. The protocol based on an efficient cyclization procedure followed by hydrolysis and oxidation allows preparation of α‐keto‐β‐lactams 5a—g with sensitive substituents.     

Effect of α-tocopherol and Trolox on the decomposition of methyl linoleate hydroperoxides

To clarify the mechanisms of antioxidant action, the effect of α-tocopherol and its water-soluble carboxylic acid derivative, Trolox, was studied on the decomposition of methyl linoleate hydroperoxides (MeLoOOH). Decomposition rate and the distribution of autoxidation products formed from MeLoOOH were followed by analyzing the volatile and nonvolatile products by static headspace gas chromatography and normal-phase high-performance liquid chromatography, respectively. Both α-tocopherol and Trolox markedly inhibited the decomposition of MeLoOOH in a concentration-dependent way. In the absence of antioxidants, MeLoOOH was completely decomposed after incubation for 48 h at 60°C, and in the presence of equal molar concentration of antioxidants only 6–7% of initial MeLoOOH was decomposed even after 280 h of incubation. MeLoOOH produced 1.2% methyl linoleate hydroxy compounds (MeLoOH) in the presence of α-tocopherol and 3.8% in the presence of Trolox. Both antioxidants inhibited the formation of volatile decomposition products and the formation of ketodiene compounds. The hydroxy compounds may be formed by the reaction of alkoxy radical and hydrogen donating antioxidants. Conversion of MeLoOOH into stable MeLoOH demonstrated that the antioxidants α-tocopherol and Trolox trap alkoxyl radicals by H-donation.     

Gold(I)‐Catalyzed Tandem Alkoxylation/Lactonization of γ‐Hydroxy‐α,β‐Acetylenic Esters

The formation of 4‐alkoxy‐2(5H)‐furanones was achieved via tandem alkoxylation/lactonization of γ‐hydroxy‐α,β‐acetylenic esters catalyzed by 2 mol% of [2,6‐bis(diisopropylphenyl)imidazol‐2‐ylidine]gold bis(trifluoromethanesulfonyl)imidate [Au(IPr)(NTf₂)]. The economic and simple procedure was applied to a series of various secondary propargylic alcohols allowing for yields of desired product of up to 95%. In addition, tertiary propargylic alcohols bearing mostly cyclic substituents were converted into the corresponding spiro derivatives. Both primary and secondary alcohols reacted with propargylic alcohols at moderate temperatures (65–80 °C) in either neat reactions or using 1,2‐dichloroethane as a reaction medium allowing for yields of 23–95%. In contrast to [Au(IPr)(NTf₂)], reactions with cationic complexes such as [2,6‐bis(diisopropylphenyl)imidazol‐2‐ylidine](acetonitrile)gold tetrafluoroborate [Au(IPr)(CH₃CN)][BF₄] or (μ‐hydroxy)bis{[2,6‐bis(diisopropylphenyl)imidazol‐2‐ylidine]gold} tetrafluoroborate or bis(trifluoromethanesulfonyl)imidate – [{Au(IPr)}₂(μ‐OH)][X] (X=BF₄, NTf₂) – mostly stop after the alkoxylation. Analysis of the intermediate proved the exclusive formation of the E‐isomer which allows for the subsequent lactonization.     

Structural Effects on ss‐ and dsDNA Recognition by a β‐Hairpin Peptide

Form defines function : The effects of β‐hairpin structure on the binding affinity and selectivity for ssDNA versus dsDNA were investigated; this provided insights into the factors that contribute to the selective recognition of both ss‐ and dsDNA and suggested new approaches for designing biomimetic receptors. Binding studies showed that 1) folding is crucial for binding to both ss‐ and dsDNA, and 2) chirality affects binding for duplex but not for ssDNA.

     

Effects of Temperature and Heating Time on the Formation of Four Toxic α,β‐Unsaturated‐4‐Hydroxyaldehydes in Vegetable Oils

The formation of four α,β‐unsaturated hydroxyaldehydes, 4‐hydroxy‐2‐trans‐hexenal (HHE), 4‐hydroxy‐2‐trans‐octenal (HOE), 4‐hydroxy‐2‐trans‐nonenal (HNE), and 4‐hydroxy‐2‐trans‐decenal (HDE), was detected in commercial corn, soybean, peanut, and canola oils heated for 1, 3, and 5 hours at 145, 165, and 185 °C. These four toxic aldehydes were investigated using high‐performance liquid chromatography (HPLC). These oils were selected based upon different degrees of unsaturations, especially their linoleic and linolenic acid concentrations. To select the appropriate conditions of temperatures and heating times, preliminary experiments were conducted using the thiobarbituric acid assay, which detects the formation of secondary‐oxidation products such as aldehydes and related carbonyl compounds. After various heat treatments, the formation of HHE, HOE, HNE, and HDE was detected as 2,4‐dinitrophenyl hydrazine derivatives using HPLC. In general, HHE, HOE, HNE, and HDE formation increased in all four oils with higher temperatures, longer heating times, and higher concentrations of linoleic and linolenic acids in the oils. The formation of HNE in the oils was mostly much higher than the other three 4‐hydroxyaldehyde isomers under the same conditions.     

RuO4 staining and lamellar structure of α‐ and β‐PP

Monoclinic (α) and hexagonal (β) polypropylene (α‐ and β‐PP) were stained in the vapor of a ruthenium tetroxide solution prepared in situ. The effect of staining on the fusion behavior was investigated using a DSC. A staining duration between 10 and 24 h was found suitable for obtaining a good electron contrast between the crystalline and amorphous regions for TEM examination without causing severe damage to the crystals. The spherulites of the water‐quenched α‐PP were found to be composed of very fine cross‐hatched lamellae whose long period was about 10 nm. In comparison, the β‐PP spherulites crystallized isothermally at 130°C had a category 2 morphology and the lamellae have a long period of 20 nm. The morphology of the spherulite boundary varied depending on the contact angle between the lamellae of the neighboring spherulites. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 72: 1529–1538, 1999     

α‐ and β‐crystalline forms of isotactic polypropylene investigated by nanoindentation

Under special crystallization conditions from the melt, both α‐ and β‐forms of isotactic polypropylene were produced simultaneously. The α‐ and β‐spherulites of polypropylene were differentiated under optical microscope, allowing the nanoindentation of individual spherulites of each crystallographic form. Elastic modulus and hardness of β‐spherulites were found to be 10 and 15% respectively lower than in α‐spherulites. The higher stiffness of α may be related to the particular interlocked structure with cross‐hatched lamellae, and to a lower molecular mobility in the crystallites. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 74: 195–200, 1999     

Concurrent oxidation of accumulated hydroperoxides in the autoxidation of methyl linoleate

Summary 1. Kinetic studies showed that concurrent oxidation of preformed hydroperoxides may be expected to take place at all stages of the autoxidation of methyl linoleate. The rate of oxidation relative to the rate of autoxidation of unoxidized ester is determined chiefly by the extent of the accumulation of hydroperoxides. 2. Infrared spectral analysis of hydroperoxides oxidized to various degrees indicated thattrans, trans diene conjugation and isolatedtrans double bonds produced in the autoxidation of methyl linoleate are related to the concurrent oxidation of the accumulated hydroperoxides. 3. The low absorptivity observed for diene conjugation, compared to that which may be expected for the exclusive production ofcis, trans diene conjugated hydroperoxide isomers during the autoxidation of methyl linoleate is attributed to the concurrent oxidation of accumulated hydroperoxides. 4. The effect of antioxidants in giving a well-defined induction period in the oxidation of hydroperoxides isolated from autoxidized methyl linoleate indicated that the oxidation proceeds by a chain reaction. 5. The primary reaction products of the oxidation of hydroperoxides isolated from autoxidized methyl linoleate were found to be polymers formed in a sequence of reaction involving the diene conjugation. 6. Studies on the autoxidation of methylcis-9,trans-11-linoleate showed thatcis, trans isomerization of the conjugated diene took place with the concurrent production of isolatedtrans double bonds and loss of diene conjugation. Hormel Institute publication no. 138. Presented before the American Oil Chemists’ Society, Philadelphia, Pa., Oct. 10–12, 1955. This work was supported by a grant from the Hormel Foundation.     

Counter‐ion‐specific helix formation of poly(γ‐methyl L‐glutamate) derivatives containing sulfonate group in aqueous alcohols

Charged polypeptides containing sulfonate groups were prepared by transesterification of poly(γ‐methyl L ‐glutamate) with isethionic acid. The coil–helix transition of the sulfonated polypeptides was investigated in aqueous alcohols. Marked counter‐ion specificity was observed for helix formation: Li⁺ < Na⁺ < Cs⁺ ≦ Rb⁺ ≦ K⁺; this was different to that for poly(L ‐glutamate) (PLG): Cs⁺ ? K⁺ < Li⁺ < Na⁺. Specific helix stabilization by counter‐ion mixing, which has been found for the PLG system, was not observed for the sulfonated polypeptides. The counter‐ion‐ and solvent‐specific helix formation is discussed and compared with that in PLG. © 2001 Society of Chemical Industry     

Gas chromatographic study on high-temperature thermal degradation products of methyl linoleate hydroperoxides

Chromatographic techniques were used to separate secondary products generated by thermal degradation of methyl linoleate hydroperoxides (MLHP). The MLHP were obtained by oxidation, selected, and concentrated by solid-phase extraction (SPE) and thin-layer chromatography (TLC). The purified MLHP were then thermo-degraded in the gas-chromatographic glass liner and analyzed on-line by gas chromatography-mass spectrometry (GC-MS). The MLHP were also thermodegraded and collected in a short silicic acid-packed column, eluted, separated by TLC, and then analyzed by GC. By considering the elution in TLC, the GC retention times and the GC-MS analyses, it was possible to characterize the mono- and the dioxygenated secondary products, particularly those having a boiling point higher than methyl linoleate. The peaks that corresponded to the mono-oxygenated products (epoxy, hydroxy, and keto) were identified, and, on the basis of their MS spectra, molecular structures were proposed. A specific elution order was suggested for keto derivatives: 9-keto,Δ^10,12- and 13-keto,Δ^9,11-octadecadienoate. The hydroxy derivatives, which show the typical fragmentations of 9-hydroxy,Δ^10,12- and 13-hydroxy,Δ^9,11-octadecadienoate, were also identified. On the other hand, identification of the di-oxygenated compounds was more difficult, and, therefore, it was not possible to indicate each positional isomer; however, their elution order could be epoxy-hydroxy and epoxy-keto derivatives.     

Acetylcholine Promotes Binding of α‐Conotoxin MII at α3β2 Nicotinic Acetylcholine Receptors

α‐Conotoxin MII (α‐CTxMII) is a 16‐residue peptide with the sequence GCCSNPVCHLEHSNLC, containing Cys2–Cys8 and Cys3–Cys16 disulfide bonds. This peptide, isolated from the venom of the marine cone snail Conus magus, is a potent and selective antagonist of neuronal nicotinic acetylcholine receptors (nAChRs). To evaluate the impact of channel–ligand interactions on ligand‐binding affinity, homology models of the heteropentameric α₃β₂‐nAChR were constructed. The models were created in MODELLER with the aid of experimentally characterized structures of the Torpedo marmorata‐nAChR (Tm‐nAChR, PDB ID: 2BG9) and the Aplysia californica‐acetylcholine binding protein (Ac‐AChBP, PDB ID: 2BR8) as templates for the α₃‐ and β₂‐subunit isoforms derived from rat neuronal nAChR primary amino acid sequences. Molecular docking calculations were performed with AutoDock to evaluate interactions of the heteropentameric nAChR homology models with the ligands acetylcholine (ACh) and α‐CTxMII. The nAChR homology models described here bind ACh with binding energies commensurate with those of previously reported systems, and identify critical interactions that facilitate both ACh and α‐CTxMII ligand binding. The docking calculations revealed an increased binding affinity of the α₃β₂‐nAChR for α‐CTxMII with ACh bound to the receptor, and this was confirmed through two‐electrode voltage clamp experiments on oocytes from Xenopus laevis. These findings provide insights into the inhibition and mechanism of electrostatically driven antagonist properties of the α‐CTxMIIs on nAChRs.     

Preparation and swelling behavior of biodegradable hydrogels based on α,β‐poly(N‐2‐hydroxyethyl‐DL‐aspartamide)

Biodegradable polymers and the hydrogels have been increasingly applied in a variety of biomedical fields and pharmaceutics. α,β‐Poly(N‐2‐hydroxyethyl‐DL ‐aspartamide), PHEA, one of poly(amino acid)s with hydroxyethyl pendants, are known to be biodegradable and biocompatible, and has been studied as an useful biomaterial, especially for drug delivery, via appropriate structural modification. In this work, hydrogels based on PHEA were prepared by two‐step reaction, that is, the crosslinking of polysuccinimide, the precursor polymer, with oligomeric PEG or PEI‐diamines and the following nucleophilic ring‐opening reaction by ethanolamine. Soft hydrogels possessing varying degrees of gel strength could be prepared easily, depending on the amount of different crosslinking reagents. The swelling degrees, which were in the range of 10–40 g–water/dry gel, increased somewhat at higher temperature, and also at alkaline pH of aqueous solution. A typical hydrogel remained almost unchanged for 1 week, at 37°C in phosphate buffer of pH 7.4, and then seemed to degrade slowly as time. A porous scaffold could be fabricated by the freeze drying of water‐swollen gel. The PHEA‐based hydrogels have potential for useful biomaterial applications including current drug delivery system. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 90: 3741–3746, 2003