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.     

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.     

Alkylation of 1,2,3,9a-tetrahydro-9H-imidazo[1,2-a]indol-2-ones

Reaction of 1,2,3,9a-tetrahydro-9H-imidazo[1,2-a]indol-2-ones with dimethyl sulfate and haloalkanes in DMF or DMSO in the presence of potassium hydroxide gives the 1-substituted 1,2,3,9a-tetrahydro-9H-imidazo[1,2-a]indol-2-ones (N-substituted products) and 9,9a-dihydro-3H-imidazo[1,2-a]indoles (O-substituted products). The latter, on treatment with acids and bases, are converted into 1-alkoxycarbonylmethyl-2,3-dihydro-1H-indoles. 1-Ethoxycarbonyl-methyl-2,3-dihydro-1H-indoles on treatment with lithium aluminohydride undergoes cyclization to 2,3,9,9a-tetrahydrooxazolo[3,2-a]indole.Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 1, pp. 49–53, January, 1988.     

Solid-phase synthesis of sn-1,2- and sn-2,3-diacylglycerols via ring-opening of the glycidyl-bound resin

A general method developed for the parallel solid-phase synthesis of sn-1,2- and sn-2,3-diacyglycerol derivatives. The technique relies on the use of carboxylic acid-promoted epoxide ring-opening reactions of the glycidyl-bound resin 3. The polymer-bound monoacylglycerol 5, formed in this manner, is transformed to the respective polymer-bound diacylglycerols 7 and 9 by reaction of the free alcohol moiety with a second carboxylic acid under conditions that lead to retention or inversion of C-2 stereochemistry. The sequence is completed by BCl3-promoted cleavage of 7 and 9 to form the sn-1,2- and sn-2,3-diacylglycerols, respectively.     

Base- and Co(II)-catalyzed ring-opening reactions of perhydrooxireno[2,3-d][1,2]dioxines: an efficient route to 4-hydroxy-2,3-epoxy-ketones

A series of 3,4,6-substituted 3,6-dihydro-1,2-dioxines were epoxidized with m-chloroperbenzoic acid to furnish perhydrooxireno[2,3-d][1,2]dioxines (epoxy-1,2-dioxines) in yields ranging from 51% to 93% with de's from 26% to 100%. Unsymmetrical epoxy-1,2-dioxines were ring-opened using triethylamine to yield 4-hydroxy-2,3-epoxy-ketones quantitatively, and meso-epoxy-1,2-dioxines were ring-opened using Co(II) salen complexes to afford 4-hydroxy-2,3-epoxy-ketones in 77-98% yield. The first reported examples of the catalytic asymmetric ring-opening of meso-epoxy-1,2-dioxines using a range of chiral Co(II) salen and beta-ketoiminato complexes to afford highly enantio-enriched 4-hydroxy-2,3-epoxy-ketones are also presented.     

Heteroaromatic annulation of 2-methyl/2-cyanomethylbenzimidazole dianions with alpha-oxoketene dithioacetals: a highly regioselective synthetic protocol for 1,2- and 2,3-substituted/annulated pyrido[1,2-a]benzimidazoles

A highly efficient and regioselective annulation protocol for a series of linearly 2,3- and angularly 1,2-substituted and annulated pyrido[1,2-a]benzimidazoles involving [3 + 3] cyclocondensation of the dianions generated from 2-methyl (2A) and 2-cyanomethyl (3A) benzimidazoles with a variety of alpha-oxoketene dithioacetals has been reported. Thus the dianion 2A derived from 2-methylbenzimidazole has been shown to undergo regioselective 1,2-addition with various alpha-oxoketene dithioacetals derived from acyclic (4a-d) and cyclic ketones (13a,b, 20, 29 and 32) to afford various carbinol acetals which on intramolecular cyclocondensation in the presence of phosphoric acid furnish the corresponding 1-methylthio-2,3-substituted (5a-c) and 2,3-fused linear polycyclic (14a,b,21, 30, and 33) pyrido[1,2-a]benzimidazoles in high yields. Similarly the dianion 3A from 2-cyanomethylbenzimidazole undergoes one-pot conjugate addition-elimination and cyclocondensation with these alpha-oxoketene dithioacetals to give 4-cyano-3-(methylthio)-1(or 1,2-)-substituted (6a-d) and the corresponding angularly 1,2-fused (16a,b, 23, 31, and 34) polycylic analogues of pyrido[1,2-a]benzimidazoles in excellent yields.     

Triacylglycerol lipase from rape (Brassica napus L.) suitable for biotechnological purposes

Triacylglycerol lipase (EC 3.1.1.3) from rape (Brassica napus L. cv Ceres) is quite easily prepared from the 100,000 x g supernatant of cotyledon homogenates. The lipase is present in a high-molecular-mass fraction ( > 1.5 x 10⁶ dalton by gel filtration), but it can be rapidly extracted from the 100,000 xg supernatant by precipitation with polyethyleneglycol 8000 (4%, w/v) and MgCl₂ (40 mM) giving about a 10-fold purification. After delipidation, the lipase has an M_r of about 300,000. It hydrolyzes triacylglycerols to fatty acids and glycerol, although the fatty acids from the sn-1 or -3 positions are hydrolyzed first to yield 1,2(2,3)-diaclyglycerols. Lipase immobilized onto Celite by precipitation with acetone at-20°C catalyzes the esterification of oleic acid with butanol dissolved in hexane.     

Stereoselective synthesis of 2,6-disubstituted 3-piperidinols: application to the expedient synthesis of (+)-julifloridine

[reaction: see text] The asymmetric synthesis of 2,6-disubstituted 3-piperidinols having a 2,3-cis and 2,6-trans relative stereochemistry was accomplished in three steps using the following sequence: stereocontrolled nucleophilic addition of an organomagnesium reagent to a chiral pyridinium salt; monohydrogenation of the resulting 2-substituted 1,2-dihydropyridine; and a one-pot, highly diastereoselective epoxidation-nucleophilic addition with a heteroatom nucleophile or an organometallic reagent. This methodology was applied to the expedient asymmetric synthesis of (+)-julifloridine in four steps.     

Liquid chromatographic separation of the enantiomers of trans-chlordane, cis-chlordane, heptachlor, heptachlor epoxide and alpha-hexachlorocyclohexane with application to small-scale preparative separation

Analytical high-performance liquid chromatographic separations of the individual enantiomers of five polychlorinated compounds were obtained on polysaccharide stereoselective HPLC columns. The enantiomers of the pesticides trans-chlordane, cis-chlordane and heptachlor were separated on CHIRALCEL OD using a hexane mobile phase. The enantiomers of the heptachlor metabolite, heptachlor epoxide, were separated on CHIRALPAK AD using a methanol mobile phase. The enantiomers of alpha-hexachlorocyclohexane (alpha-HCH), were separated on CHIRALCEL OJ using a hexane/2-propanol mobile phase. Similar chromatographic conditions using preparative columns were used to isolate approximately 250 mg of each of the individual enantiomers. The purified individual enantiomers have been submitted for testing of their endocrine disruptor (ED) activity.     

High-performance liquid chromatographic separation of the enantiomers of organophosphorus pesticides on polysaccharide chiral stationary phases.

High-performance liquid chromatographic separation of the individual enantiomers of 12 organophosphorus pesticides (OPs) was obtained on polysaccharide enantioselective HPLC columns using alkane-alcohol mobile phase. The OP pesticides were crotoxyphos, dialifor, fonofos, fenamiphos, fensulfothion, isofenphos, malathion, methamidophos, profenofos, crufomate, prothiophos and trichloronate. The enantiomers of fenamiphos, fensulfothion, profenofos and crufomate were separated on CHIRALPAK AD; the enantiomers of fenamiphos were also separated on CHIRALPAK AS; the enantiomers of methamidophos, crufomate and trichloronate were separated on CHIRALCEL OD; the enantiomers of crotoxyphos, dialifor, fonofos, malathion, prothiophos and trichloronate were separated on CHIRALCEL OJ; and the enantiomers of isofenphos were separated on CHIRALCEL OG. Baseline or partial separation of the enantiomers of six of these OP pesticides was obtained on CHIRALCEL OJ. In continued method development, the separation of the enantiomers of the 12 OPs was investigated more extensively on CHIRALCEL OJ to determine whether the mobile phase composition, flow-rate and column temperature could be optimized to yield at least partial separation of the enantiomers. Chromatographic conditions were found that gave either baseline or near baseline separations of the enantiomers of the 12 OPs on the CHIRALCEL OJ column.     

Efficient two-step synthesis of 3-halo-3-enals or 2-halo-2-alkenyl ketones from propargylic bromides via a unique cationic 1,2-aryl or proton shift in electrophilic addition reaction of 2,3-allenols with X+

The reaction of readily available 1-substituted 2,3-allenols with Br2, NBS, or I2 afforded the not-easily-available but synthetically useful 3-halo-3-alkenals or 2-halo-2-alkenyl ketones in good yields via a sequential electrophilic interaction of X+ with the allene moiety , a 1,2-aryl or proton shift, and a H+-elimination process; the structures of the products were established by X-ray diffraction study.     

Analysis of triacylglycerols with non-aqueous reversed-phase liquid chromatography and positive ion electrospray tandem mass spectrometry

A non-aqueous reversed-phase liquid chromatographic method coupled to electrospray ionisation (ESI) tandem mass spectrometry was developed for the analysis of triacylglycerols (TGs). The synthetic TGs studied were separated according to their equivalent carbon number with a gradient of methanol (containing 0.01% (v/v) formate adjusted to pH 5.3 with ammonia) and chloroform. ESI mass spectra of TGs yielded positive ion current signals for [M + NH(4)](+) and [M + NH(4)-RCOONH(4)](+). The mass spectra also showed signals believed to arise from [M + K](+). Collision-induced dissociation (CID) of the [M + NH(4)](+) precursor ion yielded [M + NH(4) - RCOONH(4)](+), [RCO + 74](+) and [RCO](+) product ions as aids for the structural elucidation of the TGs. In addition, [RCO - 18](+) and small amounts of [RCO - 2](+) product ions were also found. The latter ions were observed only for TGs containing unsaturated fatty acids. CID of ammoniated 1-stearoyl-2-oleoyl-3-linoleoyl-glycerol (18:0/18:1/18:2) indicated that neutral loss of the sn-2 fatty acid was energetically less favourable than loss of the fatty acid from the sn-1 or sn-3 position.     

High-performance liquid chromatographic resolution of reverse isomers of 1,2-diacyl-rac-glycerols as 3,5-dinitrophenylurethanes

The resolution of reverse isomers remains a major unsolved problem in glycerolipid chromatography. We have investigated the separation of the reverse isomers of 1,2-diacyl-rac-glycerols under a variety of high-performance liquid chromatography (HPLC) conditions. The reverse isomers of diacylglycerols having various pairs of acyl groups including short and highly unsaturated chains, which were prepared by partial Grignard degradation of the corresponding triacylglycerols, were chromatographed as 3,5-dinitrophenylurethanes. Excellent resolution was achieved for the reverse isomers of very different pairs of acyl groups, such as acetate-palmitate and docosahexaenoate-palmitate, by chiral-phase HPLC on columns containing (R)- and (S)-1-(1-naphthyl)ethylamine polymeric phases, reversed-phase HPLC on a highly efficient C18 column (4 microm particle size) and silver ion HPLC on a silver loaded cation-exchange column. The chiral-phase HPLC also permitted complete enantiomer resolution for all the reverse isomers examined. No satisfactory resolution by any of the HPLC methods, however, was obtained for the reverse isomers possessing minor differences in chain lengths and degree of unsaturation, such as laurate-palmitate and oleate-linoleate. The limitations of resolution and characteristics of elution are described.     

A Concise Synthetic Method for Constructing 3-Substituted Piperazine-2-Acetic Acid Esters from 1,2-Diamines

We report a short synthetic route for synthesizing 2,3-substituted piperazine acetic acid esters. Optically pure amino acids were efficiently converted into 1,2-diamines that could be utilized to deliver the title 2,3-substituted piperazines in five steps with a high enantiomeric purity. The novel route facilitated, for the first time, the synthesis of 3-phenyl substituted-2-piperazine acetic acid esters that were difficult to achieve using other methods; however, in this case, the products underwent racemization.     

Reaction of 2-(2-oxo-2,3-dihydro-1<Emphasis Type="Italic">H</Emphasis>-indol-3-ylidene)acetic acid esters with benzene-1,2-diamine and 2-aminobenzenethiol

2-(2-Oxo-2,3-dihydro-1H-indol-3-ylidene)acetic acid esters reacted with benzene-1,2-diamine or 2-aminobenzenethiol to give (2-oxo-2,3-dihydro-1H-indol-3-yl)-substituted 3,4-dihydroquinoxalin-2(1H)-ones or 2H-1,4-benzothiazin-3(4H)-ones.     

Studies toward a synthesis of trilobatin B, a lignan from the liverwort Bazzania trilobata: asymmetric construction of the tetrahydrofuran segment

A novel and stereocontrolled process is described for the asymmetric synthesis of the tetrahydrofuran segment of a 2,3-dicarboxy-6,7-dihydroxy-1-(3′,4′-dihydroxyphenyl)-1,2- dihydronaphthalene mono-ester, trilobatin B, a lignan from the liverwort Bazzania trilobata. The key cis-substituted lactone ring was constructed in a stereoselective manner by Horner-Emmons reaction followed by the subsequent tandem Michael addition and cyclization of two types of lactol intermediates elaborated from natural sources.     

Tuning lipase enantioselectivity in organic media using solid-state buffers.

The enantioselectivity exhibited by Candida antarctica lipase B (CALB) in predominantly organic media has been studied for different enzyme protonation states. Alcoholysis of (+/-)-2-phenyl-4-benzyloxazol-5(4H)-one (1) using butan-1-ol as the nucleophile in low-water organic solvents was used as a model reaction. Using either organo-soluble bases or the newly introduced solid-state buffers of known pK(a), the protonation state of the lipase was altered. By choice of the appropriate solid-state buffer or organic base, the enantioselectivity could be selectively tuned. Both Et(3)N and the solid-state buffer pair CAPSO/CAPSO.Na were found to increase the enantioselectivity of the reaction catalyzed by CALB and that of another lipase (Mucor miehei). Significant differences to both the enantioselectivity and catalytic rate were observed, especially under hydrated conditions where byproduct acid was formed.     

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.     

Diastereoisomerism of thiol complexes of arsenic acids and pseudoasymmetry of arsenic: a 1H and 13C NMR study

Diastereoisomeric complexes of methylphenylarsinic acid and (L)-glutathione could be partially separated by HPLC, but the separated compounds rapidly racemized, presumably by pyramidal inversion at the arsenic atom. Hydrolysis of the diastereoisomeric complexes yielded methylphenylarsinous acid as a pair of enantiomers revealed by a 1H NMR study with an asymmetric lanthanide shift reagent. Methylphenylarsinous acid was also synthesized as an enantiomeric pair, shown by an asymmetric shift reagent experiment, by the hydrolysis of iodomethylphenylarsine. 1H and 13C NMR spectroscopy were used to demonstrate that complexing of phenylarsonic acid with (R,S)-3-mercapto-1,2-propanediol and with (R,S)-1-mercapto-2-propanol yielded, in each case, a pair of enantiomers, PhAs[(R)-ligand)]2, PhAs[(S)-ligand)]2, in which the homomorphic ligands were diastereotopic, and a pair of diastereoisomers, PhAs[(R)-ligand][(S)-ligand], which differed from each other in the configuration about the pseudoasymmetric arsenic atom.     

Studies on Cu(I)-catalyzed synthesis of simple 3-substituted 1,2-allenes and optically active 2-substituted secondary 2,3-allenols

The sequential treatment of terminal alkynes or propargylic alcohols with n-BuLi and MOMCl afforded the corresponding propargylic methyl ethers, which would react with primary alkyl Grignard reagents under the catalysis of CuBr to afford 3-substituted 1,2-allenes or 2-substituted secondary 2,3-allenols, respectively. The reaction may be applied to the synthesis of optically active 2-substituted secondary 2,3-allenols with up to >99% ee without any protection to the free hydroxyl group in the starting 4-hydroxy-2-alkynyl methyl ethers.