Bis-2-oxo amide triacylglycerol analogues: a novel class of potent human gastric lipase inhibitors.

A novel class of potent human gastric lipase inhibitors, bis-2-oxo amide triacylglycerol analogues, was developed. These analogues of the natural substrate of lipases were prepared starting from 1,3-diaminopropan-2-ol. They were designed to contain the 2-oxo amide functionality in place of the scissile ester bond at the sn-1 and sn-3 position, while the ester bond at the sn-2 position was either maintained or replaced by an ether bond. The derivatives synthesized were tested for their ability to form stable monomolecular films at the air/water interface by recording their surface pressure/molecular area compression isotherms. The inhibition of human pancreatic and gastric lipases by the bis-2-oxo amides was studied using the monolayer technique with mixed films of 1,2-dicaprin containing variable proportions of each inhibitor. The nature of the functional group (ester or ether), as well as the chain length, at the sn-2 position influenced the potency of the inhibition. Among the compounds tested, 2-[(2-oxohexadecanoyl)amino]-1-[[(2-oxohexadecanoyl)-amino]methyl]ethyl decanoate was the most potent inhibitor, causing a 50% decrease in HPL and HGL activities at 0.076 and 0.020 surface molar fractions, respectively.     

Sterically hindered triacylglycerol analogues as potent inhibitors of human digestive lipases

A novel class of inhibitors of human digestive lipases have been developed. Various sterically hindered triacylglycerols based on 2-methyl- and 2-butylglycerol, and/or 2-methyl fatty acids were synthesized. The triacylglycerol analogues were tested for their ability to form stable monomolecular films at the air/water interface by recording their surface-pressure/molecular-area compression isotherms. The inhibition of human pancreatic and gastric lipases by the sterically hindered triacylglycerol analogues was studied by using the monolayer technique with mixed films of 1,2-dicaprin, which contained variable proportions of each inhibitor. Triolein analogues that contain a butyl group at the 2-position of the glycerol backbone or methyl groups both at the 2-position of glycerol, and the alpha-position of each oleic acid residue were potent inhibitors; this caused a 50% decrease in HPL activity at 0.003 molar fraction.     

Scorpion digestive lipase: kinetic study using monomolecular film technique

Using the classical emulsified system and the monomolecular film technique, we compared the interfacial properties of the scorpion digestive lipase (SDL) with those of higher animals'. In the absence of bile slats, SDL does not hydrolyse efficiently pure tributyrin, as well as dicaprin films maintained at low surface pressure. The preincubation of bile salts with tributyrin seems to be a better substrate for SDL than the pure tributyrin. A kinetic study on the surface pressure dependency, stereospecificity and regioselectivity of SDL was performed using monomolecular films of either three dicaprin isomers or three pairs of didecanoyl-deoxyamino-O-methyl glycerol enantiomers (DDG) containing a single hydrolysable decanoyl ester bond. With all diacylglycerol isomers, SDL has a surface pressure threshold of about 15 m Nm(-1), below which enzymatic activity is undetectable. SDL seems to prefer vicinal ester groups of the diacylglycerol isomers, with preference for sn-1 position at both 15 and 23 m Nm(-1). Furthermore, the maximum SDL activity is measured with DDG having a primary ester bond (1,3DDG, SII). This shows that SDL has a preference for the sn-1 position of this diacylglycerol analogue. Moreover, this was in line with the fact that SDL is inactive on sn-2 position of both DDG isomers and a triacylglycerol. With diacylglycerol analogue isomers, SDL shows a preference for distal isomers contrary to what has been observed with diacylglycerol isomers. SDL interacts with egg-phosphatidyl choline (egg-PC) monomolecular films. The critical surface pressure value (13 m Nm(-1)) is comparable to those of pancreatic lipases.     

Synthesis of lipophilic aldehydes and study of their inhibition effect on human digestive lipases

[reaction: see text] Novel inhibitors of human digestive lipases, aldehyde dialkyl and alkyl-acyl glycerol analogues, were developed. The inhibitors were prepared starting from 3-(benzyloxy)-1,2-propanediol. The inhibition of human pancreatic and gastric lipases by the aldehyde derivatives was studied using the monolayer technique. (1R)-1-[(Dodecyloxy)methyl]-4-oxobutyl decanoate caused a 50% decrease in HPL and HGL activities at 0.100 and 0.053 molar fractions, respectively.     

Lipases and their industrial applications

Lipases (triacylglycerol acylhydrolase, EC 3.1.1.3) are part of the family of hydrolases that act on carboxylic ester bonds. The physiologic role of lipases is to hydrolyze triglycerides into diglycerides, monoglycerides, fatty acids, and glycerol. These enzymes are widely found throughout the animal and plant kingdoms, as well as in molds and bacteria. Of all known enzymes, lipases have attracted the most scientific attention. In addition to their natural function of hydrolyzing carboxylic ester bonds, lipases can catalyze esterification, interesterification, and transesterification reactions in nonaqueous media. This versatility makes lipases the enzymes of choice for potential applications in the food, detergent, pharmaceutical, leather, textile, cosmetic, and paper industries. The most significant industrial applications of lipases have been mainly found in the food, detergent, and pharmaceutical sectors. Limitations of the industrial use of these enzymes have mainly been owing to their high production costs, which may be overcome by molecular technologies, enabling the production of these enzymes at high levels and in a virtually purified form.     

Preparation, isolation, and characterization of Nalpha-Fmoc-peptide isocyanates: solution synthesis of oligo-alpha-peptidyl ureas

The N(alpha)-Fmoc-peptide isocyanates 3a-q, 4a-c, and 5a-c were prepared by the Curtius rearrangement of N(alpha)-Fmoc-peptide acid azides in toluene under thermal, microwave, and ultrasonic conditions. All the N(alpha)-Fmoc-oligo-peptide isocyanates made were isolated as stable crystalline solids with 71 to 94% yield and were fully characterized by 1H NMR, 13C NMR, and mass spectroscopy. Their utility for the synthesis of oligo-alpha-peptidyl ureas 7a-f and 8a-c by the divergent coupling approach was demonstrated. The coupling of N(alpha)-Fmoc-dipeptide isocyanates with amino acid ester or with N,O-bis(trimethylsilyl)amino acids resulted in N(alpha)-Fmoc-tripeptidyl urea ester and acids containing one each of peptide bond and urea bond. The divergent approach is extended to the synthesis of tetrapeptidyl ureas by the 2 + 2 strategy using bis-TMS-peptide acid as an amino component. To incorporate urea bonds in adjacent positions, N(alpha)-Fmoc-peptidyl urea isocyanates 9a-d were prepared and employed in the synthesis of three tetrapeptidyl ureas 10a-b and 11 containing one peptide bond and two urea bonds in series from the N-terminal end. The protocol was then employed for the synthesis of five urea analogues 13-15, 18, and 21 of [Leu5]enkephalin containing urea bonds at the 2, 3, 4 positions as well as at the 2, 4 and 2, 3, 4 positions. The analogue 2l was made by the convergent synthesis by the N --> C terminal chain extension. Finally, two urea analogues 22 and 23 of repeat units of bioelasto polymers, namely Val-Pro-Gly-Val-Gly-OH and Pro-Gly-Val-Gly-Val-OH, were synthesized incorporating the urea bond by the concomitant isocyanate generation and urea bond formation under thermal conditions.     

Identification of triacylglycerols containing two short-chain fatty acids at sn-2 and sn-3 positions from bovine udder by fast atom bombardment tandem mass spectrometry

Several triacylglycerol (TAG) molecular species, that contain two short-chain fatty acids (C4 to C8) at the sn-2 and sn-3 positions of the glycerol backbone, were isolated from bovine udder by using solvent extraction and silica gel column chromatography. Their structures were identified by fast atom bombardment (FAB) tandem mass spectrometry (MS/MS), based on the information obtained from collision-induced dissociation (CID) spectra of sodium-adducted molecules ([M + Na](+)) of model TAG compounds which had been synthesized from glycerol and appropriate fatty acids. For each species, the relative positions of the three fatty acids on the glycerol backbone, as well as fatty acid composition and double-bond position in the fatty acyl group, were determined. A majority of sodium-adducted molecules observed in the FAB mass spectrum were mixtures of at least two components that have different fatty acid composition but the same molecular mass. In addition, all the components present in mixtures of all the species contain a long-chain fatty acid (C12 to C18) at the sn-1 position, a short-chain fatty acid (C4 to C8) at the sn-2 position, and a butyric acid uniquely at the sn-3 position.     

Synthesis of alpha-fluorinated phosphonates from alpha-fluorovinylphosphonates: a new route to analogues of lysophosphatidic acid

A versatile, efficient method for the preparation of alpha-monofluoromethylene (-CHF-) phosphonates from alpha-fluorovinylphosphonate provides access to a class of lysophosphatidic acid (LPA) receptor-subtype agonists. In addition, sn-2 O-methylation of alpha-monofluoromethylene phosphonates using trimethylsilyldiazomethane generated sn-1-acyl, 2-O-methyl alpha-monofluoromethylene derivatives. Finally, a novel method for the selective etherification of 1,2-diols was developed and a new class of sn-1 O-methyl, 2-acyl alpha-monofluoromethylene LPA analogues was prepared. [reaction: see text]     

Synthesis and properties of aromatic poly(ester amide)s with pendant phosphorus groups

Two series of phosphorus‐containing aromatic poly(ester amide)s with inherent viscosities of 0.46–3.20 dL/g were prepared by low‐temperature solution polycondensation from 1,4‐bis(3‐aminobenzoyloxy)‐2‐(6‐oxido‐6H‐dibenz〈c,e〉〈1,2〉oxaphosphorin‐6‐yl)naphthalene and 1,4‐bis(4‐aminobenzoyloxy)‐2‐(6‐oxido‐6H‐dibenz〈c,e〉〈1,2〉oxaphosphorin‐6‐yl)naphthalene with various aromatic diacid chlorides. All the poly(ester amide)s were amorphous and readily soluble in many organic solvents, such as N,N‐dimethylformamide, N,N‐dimethylacetamide (DMAc), and N‐methyl‐2‐pyrrolidone (NMP). Transparent, tough, and flexible films of these polymers were cast from DMAc and NMP solutions. Their casting films had tensile strengths of 71–214 MPa, elongations to break of 5–10%, and initial moduli of 2.3–6.0 GPa. These poly(ester amide)s had glass‐transition temperatures of 209–239 °C (m‐series) and 222–267 °C (p‐series). The degradation temperatures at 10% weight loss in nitrogen for these polymers ranged from 462 to 489 °C, and the char yields at 800 °C were 55–63%. Most of the poly(ester amide)s also showed a high char yield of 35–45%, even at 800 °C under a flow of air. The limited oxygen indices of these poly(ester amide)s were 35–46. © 2002 John Wiley & Sons, Inc. J Polym Sci Part A: Polym Chem 40: 459–470, 2002; DOI 10.1002/pola.10129     

Synthesis of potential antitumor agents,dimeric and trimeric chlorins,from methylpheophorbide <Emphasis Type="Italic">a</Emphasis>

A series of dimeric and trimeric chlorins were synthesized from methylpheophorbide a. They are potential photosensitizers for photodynamic therapy in oncology. The macrocycles were conjugated due to the formation of ester and amide bonds. The carboxy groups were activated and catalytic transesterification was carried out to form the ester bond. The amide bond was formed using carboxy group activation; in several cases, amidation of the ester group in position 13(2) of the exocycle of methylpheophorbide α was carried out, which does not require activation.     

Lipids of Hibiscus cannabinus seeds

The neutral and polar lipids of kenaf seeds of the variety Uzbetskii-1574 from the 1983 crop have been studied. The fatty acid compositions of six acyl-containing classes of neutral lipids have been established for the first time. The acids in the sn-2 positions of the triacylglycerols and of the main classes of phospholipids have been determined. The main triacylglycerol species are those in which the sn-2 position is esterified by linoleic acid.     

Microwave-assisted domino access to C2-chain functionalized furans from tertiary propargyl vinyl ethers

Tertiary propargyl vinyl ethers armed with an electron-withdrawing group (amide or ester) at the tertiary propargylic position have been efficiently transformed into trisubstituted C(2)-chain functionalized furans. The metal-free domino transformation involves a microwave-assisted tandem [3,3]-propargyl Claisen rearrangement/5-exo-dig O-cyclization reaction. The manifold can be performed in a one-pot fashion from the primary components (1,2-ketoester/1,2-ketoamide or tertiary propargyl alcohols).     

Structural characterization of synthetic poly(ester amide) from sebacic acid and 4-amino-1-butanol by matrix-assisted laser desorption ionization time-of-flight/time-of-flight tandem mass spectrometry

Matrix-assisted laser desorption/ionization time-of-flight/time-of-flight tandem mass spectrometry (MALDI-TOF/TOF-MS/MS) was employed to analyze a poly(ester amide) sample (PEA-Bu) from the melt condensation of sebacic acid and 4-amino-1-butanol. In particular, we investigated the fragmentation pathways, the ester/amide bond sequences and the structure of species derived from side reactions during the synthesis. MALDI-TOF/TOF-MS/MS analysis was performed on cyclic species and linear oligomers terminated by dicarboxyl groups, carboxyl and hydroxyl groups and diamino alcohol groups. The sodium adducts of these oligomers were selected as precursor ions. Different end groups do not influence the fragmentation of sodiated poly(ester amide) oligomers and similar series of product ions were observed in the MALDI-TOF/TOF-MS/MS spectra. According to the structures of the most abundant product ions identified, the main cleavages proceed through a beta-hydrogen-transfer rearrangement, leading to the selective scission of the --O--CH2-- bonds. Abundant product ions originating from --CH2--CH2-- (beta-gamma) bond cleavage in the sebacate moiety were also detected. Their formation should be promoted by the presence of an alpha,beta-unsaturated ester or amide end group. MALDI-TOF/TOF-MS/MS provided structural information concerning the ester/amide sequences in the polymer chains. In the MALDI-TOF/TOF-MS/MS spectra acquired, using argon as the collision gas, of cyclic species and linear oligomers terminated by diamino alcohol groups, product ions in the low-mass range, undetected in the mass spectra acquired using air as the collision gas, proved to be diagnostic and made it possible to establish the presence of random sequences of ester and amide bonds in the poly(ester amide) sample. Furthermore, MALDI-TOF/TOF-MS/MS provided useful information to clarify the structures of precursor ions derived from side reactions during the synthesis.     

N[bond]C(alpha) bond dissociation energies and kinetics in amide and peptide radicals. Is the dissociation a non-ergodic process?

Dissociations of aminoketyl radicals and cation radicals derived from beta-alanine N-methylamide, N-acetyl-1,2-diaminoethane, N(alpha)-acetyl lysine amide, and N(alpha)-glycyl glycine amide are investigated by combined density functional theory and M?ller-Plesset perturbational calculations with the goal of elucidating the mechanism of electron capture dissociation (ECD) of larger peptide and protein ions. The activation energies for dissociations of N[bond]C bonds in aminoketyl radicals decrease in the series N[bond]CH(3) > N-CH(2)CH(2)NH(2) > N[bond]CH(2)CONH(2) approximately N[bond]CH(CONH(2))(CH(2))(4)NH(2). Transition state theory rate constants for dissociations of N[bond]C(alpha) bonds in aminoketyl radicals and cation-radicals indicate an extremely facile reaction that occurs with unimolecular rate constants >10(5) s(-1) in species thermalized at 298 K in the gas phase. In neutral aminoketyl radicals the N[bond]C(alpha) bond cleavage results in fast dissociation. In contrast, N[bond]C(alpha) bond cleavage in aminoketyl cation-radicals results in isomerization to ion-molecule complexes that are held together by strong hydrogen bonds. The facile N[bond]C(alpha) bond dissociation in thermalized ions indicates that it is unnecessary to invoke the hypothesis of non-ergodic behavior for ECD intermediates.     

Low background FRET-substrates for lipases and esterases suitable for high-throughput screening under basic (pH 11) conditions

FRET-based fluorogenic substrates for lipases and esterases were prepared in four steps from commercially available building blocks. The substrates are pyrenebutyric acid monoesters of aliphatic 1,2-diols bearing a dinitrophenylamino group as a quencher. The most enzyme-reactive substrate is ester 2a. The substrates do not show any measurable background reaction in the absence of enzyme even at pH 11, but react fast and specifically with lipases and esterases. These substrates offer an unprecedented and practical solution to the long-standing problem of a simple yet efficient high-throughput screening tool for lipase activities under basic conditions.     

Synthesis and nuclear magnetic resonance spectra of 2H-pyrido [1,2-a] pyrimidin-2-ones

Reaction of ethyl phenylpropiolate with several 2-aminopyridines afforded the corresponding 2H-pyrido[1,2-a] pyrimidin-2-ones in good yields. Analysis of the nmr spectra of these compounds and their hydrobromides were based on comparing their spectra with their 3-deuterated analogues. It was found that in these 2-oxo compounds, the proton at position-7 is shielded and absorbs together with the olefinie proton at position-3 in the region of 5 6.35-6.70 ppm. The latter proton could be used for differentiating these compounds from the corresponding 4-oxo isomers which have already been identified by the deshielded proton at pusition-6 near § 9.0 ppm. The ir and nmr spectral data for all these compounds are tabulated and discussed.     

Synthesis and Evaluation of Novel 2,2-Dimethylthiochromanones as Anti-Leishmanial Agents

Within this work, we describe the design and synthesis of a range of novel thiochromanones based on natural products reported to possess anti-leishmanial action, and their synthetic derivatives. All compounds were elaborated via the key intermediate 2,2,6-trimethoxythiochromanone, which was modified at the benzylic position to afford various ester, amine and amide analogues, substituted by chains of varying lipophilicity. Upon testing in Leishmania, IC₅₀ values revealed the most potent compounds to be phenylalkenyl and haloalkyl amides 11a and 11e, with IC₅₀ values of 10.5 and 7.2 μM, respectively.     

Direct separation of regio- and enantiomeric isomers of diacylglycerols by a tandem column high-performance liquid chromatography

A novel HPLC-based method for direct separation of the three isomers of mono-acid diacylglycerols (DAGs), i.e., 1,2-DAG, 2,3-DAG and 1,3-DAG, has been established. The method employs a tandem column system, in which two different columns (a conventional silica gel column and a chiral stationary phase column) are connected in series. Two isomeric mixtures of DAGs (i.e., dicapryloylglycerol and dioleoylglycerol) and lipase-catalyzed reaction mixtures were successfully resolved on the tandem column HPLC system without any derivatization prior to the analysis. According to the established analytical method, stereoselectivity of two lipases toward mono-acid triacylglycerols in ethanolysis reaction was investigated. The tested enzymes were immobilized Candida antarctica lipase B (CALB) and Rhizomucor miehei lipase (RML). Analyses of the enantiomeric purity of 1,2-DAG and 2,3-DAG, generated as intermediates during the reaction, revealed that CALB and RML have sn-3 and sn-1 stereopreference, respectively.     

Hydrolyzable nonionic surfactants: stability and physicochemical properties of surfactants containing carbonate, ester, and amide bonds

A linear and a branched nonionic cleavable surfactants containing a carbonate bond have been prepared from tetra(ethylene glycol) and an alkylchloroformate. The stability of these carbonate surfactants was determined by investigating their hydrolysis and biodegradability characteristics. The hydrolysis was catalyzed by alkali or enzymes (esterase from porcine liver and lipases from Mucor miehei and Candida antarctica B) and was monitored using 1H NMR. It was found that the stability toward alkali was higher for a carbonate surfactant than for a corresponding surfactant with an ester as weak bond. Biodegradation tests resulted in more than 60% degradation after 28 days for both carbonate surfactants. Physicochemical properties, such as critical micelle concentration (CMC), cloud point, area per molecule, and surface tension at the CMC, were determined and compared to those obtained from similar surfactants containing ester, amide, or ether bonds. It was found that the carbonate linkage is hydrophobic and that the oxycarbonyl part of the carbonate group is equivalent, in a formal sense, to an extra methylene group in the alkyl chain of the surfactant.     

Enantiomeric separation of asymmetric triacylglycerol by recycle high-performance liquid chromatography with chiral column

In our previous studies, we employed recycle HPLC for the separation of triacylglycerol (TAG)-positional isomers (PIs). In this study, a recycle HPLC system equipped with a polysaccharide-based chiral column was applied to the enantiomeric separation of some asymmetric TAGs having straight-chain C16-C18 acyl residues. As a result, 1,2-dipalmitoyl-3-oleoyl-rac-glycerol (rac-PPO), 1,2-dioleoyl-3-palmitoyl-rac-glycerol (rac-OOP), and 1,2-dipalmitoyl-3-linoleoyl-rac-glycerol (rac-PPL) were resolved into their respective enantiomers. However, neither 1,2-dioleoyl-3-linoleoyl-rac-glycerol (rac-OOL), consisting of only unsaturated fatty acids, nor 1,2-dipalmitoyl-3-stearoyl-rac-glycerol (rac-PPS), consisting of only saturated fatty acids, was resolved. These results suggest that the asymmetric TAGs, used in this study, having both a palmitic acid moiety and an oleic acid (or a linoleic acid) moiety at the sn-1 or sn-3 positions are resolved by the chiral column. This new chiral separation method can be used in combination with atmospheric pressure chemical ionization mass spectrometry to determine the sn-OOP/sn-POO ratio in palm oil. This method is applicable for the chiral separation of asymmetric TAGs in palm oil.