Neighboring-group participation in benzylidene acetal ring-opening of a 2-cyano-2-deoxypyranoside derivative by diethylaluminum cyanide

The oxirane ring-opening of an anhydro sugar with diethylaluminum cyanide (Et(2)AlCN) is a direct approach for obtaining a cyano derivative. Methyl 2,3-anhydro-4,6-O-benzylidene-alpha-D-allopyranoside showed anomalous chemical behavior when treated with Et(2)AlCN. The reaction afforded the corresponding beta-cyanohydrin as the minor component from a mixture of compounds resulting from the benzylidene acetal ring-opening caused by the attack of ethyl or cyano groups.     

Cis-9,trans-11-CLA exerts anti-inflammatory effects in human bronchial epithelial cells and eosinophils: comparison to trans-10,cis-12-CLA and to linoleic acid

Interaction of eosinophils and bronchial epithelial cells plays a pivotal role in maintaining inflammatory airway disease. Since conjugated linoleic acids (CLA) are suggested to exert anti-inflammatory effects, one purpose of this study was to compare cis-9,trans-11-CLA and trans-10,cis-12-CLA with regard to their influence on the stimulus-induced activation of eosinophils. ECP (eosinophil cationic protein) released in co-culture of stimulated and CLA-treated eosinophils with stimulated bronchial epithelial cells (BEAS-2B) was measured and cis-9,trans-11-CLA was found to be most potent in inhibiting ECP formation. Further, expression of the activation markers CD69 and CD13 induced by various stimuli (TNF-alpha, IL-5, IL-3) was significantly reduced in the presence of cis-9,trans-11-CLA. Subsequently, various concentrations of cis-9,trans-11-CLA vs. linoleic acid (LA, cis-9,cis-12-octadecadienoic acid) were tested for the effect on proliferative response and release of the pro-inflammatory cytokine IL-8 in stimulated BEAS-2B. Addition of cis-9,trans-11-CLA attenuated cell growth and significantly reduced IL-8 production at mRNA and protein levels. In contrast, LA had a slight stimulating effect on proliferation and was less effective in reducing the cytokine release. It was demonstrated that the inhibitory effect of cis-9,trans-11-CLA on IL-8 production is mediated through activation of the nuclear receptor PPARgamma, since blocking the receptor with a selective antagonist (GW9662) restored the stimulus-induced enhancement in IL-8 mRNA expression and protein secretion. PPARgamma has previously been shown to be closely involved in the downregulation of inflammation during hyperresponsiveness related to pulmonary immune responses. Thus, targeting PPARgamma, cis-9,trans-11-CLA might be of therapeutic value in the focus of airway disease while ameliorating inflammatory processes by affecting epithelial and eosinophil functions.     

Incorporation of cis-9,trans-11 or trans-10,cis-12 conjugated linoleic acid into plasma and cellular lipids in healthy men

This study investigated the incorporation of cis-9,trans-11 conjugated linoleic acid (c9,t11 CLA) and trans-10,cis-12-CLA (t10,c12 CLA) into plasma and peripheral blood mononuclear cell (PBMC) lipids when consumed as supplements highly enriched in these isomers. Healthy men (n = 49, age 31 +/- 8 years) consumed one, two, and four capsules containing approximately 600 mg of either c9,t11 CLA or t10,c12 CLA per capsule for sequential 8 week periods followed by a 6 week washout before consuming the alternative isomer. Both isomers were incorporated in a dose-dependent manner into plasma phosphatidylcholine (PC) (c9,t11 CLA r = 0.779, t10,c12 CLA r = 0.738; P < 0.0001) and cholesteryl ester (CE) (c9,t11 CLA r = 0.706, t10,c12 CLA r = 0.788; P < 0.0001). Only t10,c12 CLA was enriched in plasma nonesterified fatty acids. Both c9,t11 CLA and t10,c12 CLA were incorporated linearly into PBMC total lipids (r = 0.285 and r = 0.273, respectively; P < 0.0005). The highest concentrations of c9,t11 CLA and t10,c12 CLA in PBMC lipids were 3- to 4-fold lower than those in plasma PC and CE. These data suggest that the level of intake is a major determinant of plasma and PBMC CLA content, although PBMCs appear to incorporate both CLA isomers less readily.     

Formation of cis-3-hexenal,trans-2-hexenal and cis-3-hexenol in macerated Thea sinensis leaves

Thea sinensis; Theaceae; tea; cis-3-hexenal: leaf aldehyde; leaf alcohol; linolenic acid; biosynthesis of leaf alcohol.Linolenic acid and cis-3-hexenal were found in macerated leaves of Thea sinensis and this aldehyde may be produced from linolenic acid by an enzyme contained in macerated leaves in the presence of oxygen. This aldehyde was easily isomerized to trans-2-hexenal, and was converted to cis-3-hexenol by alcohol dehydrogenase. During maceration of freshly picked tea leaves, the amounts of trans-2-hexenal quickly increased and were influenced by maceration time, heating, oxygen and the pH. But in unpicked tea leaves the occurrence of trans-2-hexenal is extremely doubtful.     

Structure of cyclic dihydroxyacetone phosphate dimethyl acetal, a cyclic DHAP precursor, in the crystalline state

The six-membered cyclic phosphate diester, 5,5-dimethoxy-2-oxo-1,3,2-dioxaphosphorinane-2-ol, the dimethyl acetal of cyclic dihydroxyacetone phosphate, (MeO)(2)cDHAP, was obtained by the isolation of an intermediate in the basic hydrolysis of the cyclic triester derivative. The compound had been isolated in the form of the crystalline cyclohexylammonium (cha) salts: (cha)[(MeO)(2)cDHAP].3H(2)O (5a) and (cha)[(MeO)(2)cDHAP].H(2)O (5b), which were then converted into the free acid: (H(5)O(2))[(MeO)(2)cDHAP] (5c) and then into a series of different salts: Na[(MeO)(2)cDHAP].2H(2)O (5d), K[(MeO)(2)cDHAP].1.5H(2)O (5e), K[(MeO)(2)cDHAP].0.5H(2)O (5e'), Ca[(MeO)(2)cDHAP](2).2H(2)O (5f), CaK[(MeO)(2)cDHAP](3).2H(2)O (5g) and NH(4)[(MeO)(2)cDHAP] (5h). The synthesis of the compounds, their crystallization and crystal structures determined by X-ray crystallography are described. The most interesting structural feature observed in 5a-g anions in the crystalline state is the chair conformation of the P/O/C/C/C/O 1,3,2-dioxaphosphorinane ring, which is generally not flattened, in contrast to the deformations often observed in the analogous aryl derivatives (also in (MeO)(2)cDHAP(Ph); Slepokura, K.; Lis, T. Acta Crystallogr. Sect. C2004, 60, o315-o317). However, the anion in crystal 5h is disordered and exists in two conformations, 76% of which is the skew, S, conformation, not observed so far in the compounds of related structure.     

An archaeal endoribonuclease catalyzes cis- and trans- nonspliceosomal splicing in mouse cells

The tRNA endonuclease from the archaebacterium Methanococcus jannaschii (MJ endonuclease) can cleave RNAs forming specific bulge-helix-bulge (BHB) structures recognized by the enzyme. The resulting cleavage products are subsequently joined together by an endogenous ligase. We demonstrate the potential of using this strategy for repairing RNA in higher organisms by expressing the enzyme in mouse cells. Reporter target mRNAs modified with 17-nucleotide introns, flanked by sequences capable of forming BHB structures in cis, were expressed in mouse cells. RNA molecules that can form BHB substrates in trans with targeted mRNAs were also designed. Co-transfection of mouse cells with plasmids expressing these RNAs and the MJ endonuclease led to formation of RNA chimeras in which the target and exogenous RNA were recombined across the BHB. This technology is not limited to mRNA, but could in principle be used to destroy, modify or restore the function of a vast repertoire of RNA species or to join selectable tags to target RNAs.     

Commensurate molecules in isostructural crystals of cholesteryl cis- and trans-9-hexadecenoate

At 295 K, crystals of form I of cholesteryl cis-9-hexadecenoate (palmitoleate) and cholesteryl trans-9-hexadecenoate (palmitelaidate) are difficult to distinguish by X-ray diffraction. Both form monoclinic thin plates, space group P21 with two molecules (C43H74O2) A and B in the asymmetric unit. Unit cell dimensions for cholesteryl palmitelaidate (I) are a = 12.827(4), b = 9.075(4), c = 35.67(1) A, beta = 93.42(3) degrees, very similar to those of the palmitoleate crystals. Other crystals (form II) of the palmitelaidate ester are described. The crystal structure of form I of cholesteryl palmitelaidate has been determined from 3657 reflections (sin theta/lambda less than 0.46 A-1) measured at 295 K using CuK alpha X-radiation and refined to give Rw(F) = 0.095. The molecular packing arrangement is isostructural to that of the previously determined crystal structure of cholesteryl palmitoleate. In both crystals, the fatty acid chains of the A molecules are kinked at the double bond but are nearly straight. The chains of B molecules have more complicated dislocations and are bent. It is remarkable that, neglecting their detailed conformations, corresponding fatty acid chains in the two crystal structures have similar overall shapes, although palmitoleate chains have cis-ethylenic groups and palmitelaidate chains have trans groups.     

Quantitative gas chromatographic method for the analysis of cis-9, trans-11 and trans-10, cis-12 isomers of the conjugated linoleic acid in liver

A quantitative GC method for conjugated linoleic acid (CLA) isomers of physiological significance (cis-9, trans-11 CLA and trans-10, cis-12 CLA) as non-esterified fatty acids (NEFA) or triacylglycerols (TAG) was developed. Furthermore, the effect of the internal standard addition point (sample or fat extract) was studied. Response linearity, recovery and precision assays, detection and quantification limits were determined. Linearity was demonstrated over a range from 0.1 to 10 microg/mL. When CLA isomers were present as NEFA, the recovery significantly decreased (P< or =0.05) from 76% to 27.1% (cis-9, trans-11 CLA) and 28.5% (trans-10, cis-12 CLA) when the standards were added to the fat extract or to the initial tissue, respectively. As an application, liver samples from hamsters fed a diet supplemented with both CLA isomers were analyzed. The CLA isomers in liver samples were detected with reasonable reproducibility.     

Characterization of cis-9 trans-11 trans-15 C18:3 in milk fat by GC and covalent adduct chemical ionization tandem MS

Rumen biohydrogenation of dietary α-linolenic acid gives rise in ruminants to accumulation of fatty acid intermediates, some of which may be transferred into milk. Rumelenic acid [cis-9 trans-11 cis-15 C18:3 (RLnA)] has recently been characterized, but other C18:3 minor isomers are still unknown. The objective of this work was to identify a new isomer of octatridecenoic acid present in milk fat from ewes fed different sources of α-linolenic acid. Structural characterization of this fatty acid was achieved by GC-MS. Analysis of dimethyloxazoline and picolinyl ester derivatives allowed for location of the double bond positions. Covalent adduct chemical ionization tandem mass spectrometry confirmed the positional structure 9-11-15, identical to RLnA, and helped to establish double bond geometry (cis-trans-trans). This new C18:3 isomer could be formed by isomerization of cis-15 bond of RLnA and subsequently converted by hydrogenation to trans-11 trans-15 C18:2, an octadecadienoic acid also detected in this study.     

Different mechanisms of cis-9,trans-11- and trans-10,cis-12- conjugated linoleic acid affecting lipid metabolism in 3T3-L1 cells

Conjugated linoleic acid (CLA) has been shown to reduce body fat mass in various experimental animals. It is valuable to identify its influence on enzymes involved in energy expenditure, apoptosis, fatty acid oxidation and lipolysis. We investigated isomer-specific effects of high dose, long treatment of CLA (75.4 μmol/L, 8 days) on protein and gene expression of these enzymes in cultured 3T3-L1 cells. Proteomics identified significant up- or down-regulation of 52 proteins by either CLA isomer. Protein and gene expression of uncoupling protein (UCP) 1, UCP3, perilipin and peroxisome proliferator-activated receptor (PPAR) α increased whereas UCP2 reduced for both CLA isomers. And eight-day treatment of trans-10,cis-12 CLA, but not cis-9,trans-11 CLA, significantly up-regulated protein and mRNA levels of PKA (P<.05), CPT-1 and TNF-α (P<.01). Compared to protein expression, both isomers did not significantly influence the mRNA expression of HSL, ATGL, ACO and leptin. In conclusion, high-dose, long treatment of cis-9,trans-11 CLA did not promote apoptosis, fatty acid oxidation and lipolysis in adipocytes, but may induce an increase in energy expenditure. trans-10,cis-12 CLA exhibited greater influence on lipid metabolism, stimulated adipocyte energy expenditure, apoptosis and fatty acid oxidation, but its effect on lipolysis was not obvious.     

Substrate selectivity of various lipases in the esterification of cis- and trans-9-octadecenoic acid

The substrate selectivity of numerous commercially available lipases from microorganisms, plants and animal tissue towards 9-octadecenoic acids with respect to the cis/trans configuration of the CC double bond was examined by the esterification of cis- and trans-9-octadecanoic acid (oleic and elaidic acid respectively) with n-butanol in n-hexane. A great number of lipases studied, e.g. those from Pseudomonas sp., porcine pancreas or Carica papaya, were unable to discriminate between the isomeric 9-octadecenoic acids. However, lipases from Candida cylindracea and Mucor miehei catalysed the esterification of oleic acid 3–4 times faster than the corresponding reaction of elaidic acid and therefore have a high preference for the cis isomer. Of all biocatalysts examined, only recombinant lipases from Candidaantarctica favoured elaidic acid as substrate. While the preference of Candida antarctica lipase B for the trans isomer was quite low, Candida antarctica lipase A had an extraordinary substrate selectivity and its immobilized enzyme preparation [Chirazyme L-5 (3) from Boehringer] esterified elaidic acid about 15 times faster than oleic acid. Received: 29 October 1998 / Received revision: 18 December 1998 / Accepted: 21 December 1998     

Butters rich either in trans-10-C18:1 or in trans-11-C18:1 plus cis-9, trans-11 CLA differentially affect plasma lipids and aortic fatty streak in experimental atherosclerosis in rabbits

Dairy fat contains high amounts of saturated fatty acids (FA), which are associated with cardiovascular disease (CVD) risk. Manipulation of dairy cows nutrition allows to decrease the saturated FA content of milk fat, and is associated with increases either in conjugated linoleic acid (CLA) and trans-11-C18:1 contents, or in trans-10-C18:1 content. CLA putatively exhibits beneficial properties on CVD risk, whereas trans FA are suspected to be detrimental. The present study compared the effects of a trans-10-C18:1-rich butter (T10 butter), a trans-11-C18:1+CLA-rich butter (T11-CLA butter) and a standard butter (S butter) on lipid parameters linked to the CVD risk and fatty streaks. Thirty-six White New Zealand rabbits were fed one of the three butters (12% of the diet, plus 0.2% cholesterol) for 6 (experiment 1) or 12 (experiment 2) weeks. Liver lipids, plasma lipids and lipoprotein concentrations (experiments 1 and 2) and aortic lipid deposition (experiment 2) were determined. The T10 butter increased VLDL-cholesterol compared with the two others, and total and LDL-cholesterol compared with the T11-CLA butter ( P < 0.05). The T10 butter also increased non-HDL/HDL ratio and aortic lipid deposition compared with the T11-CLA butter ( P < 0.05). The T11-CLA butter non-significantly reduced aortic lipid deposition compared with the S butter, and decreased HDL-cholesterol and increased liver triacyglycerols compared with the two other butters (< 0.05). These results suggest that, compared with the S butter, the T10 butter had detrimental effects on plasma lipid and lipoprotein metabolism in rabbits, whereas the T11-CLA butter was neutral or tended to reduce the aortic lipid deposition.     

Metabolism of cis-12-octadecenoic acid and trans-9,trans-12-octadecadienoic acid and their influence on lipogenic enzyme activities in mouse liver

High carbohydrate (65% glucose) diets containing cis-12-octadecenoic acid (12c-18:1) or trans-9,trans-12-octadecadienoic acid (9t,12t-18:2) were fed to weanling mice to investigate the influence of fatty acid structure on six hepatic enzyme activities involved in lipid metabolism. Results with these diets were compared to those with diets containing no fatty acids, saturated fatty acids; cis-9-octadecenoic acid (9c-18:1) and cis-9,cis-12-octadecadienoic acid (9c,12c-18:2). These comparisons show saturated fatty acids, 9c-18:1, 12c-18:1, and 9t,12t-18:2, had little or no influence on the activity levels of fatty acid synthetase, malic enzyme (EC 1.1.1.40)citrate cleavage enzyme (EC 4.1.3.8), glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconate dehydrogenase (EC 1.1.1.44) and acetyl-CoA carboxylase (EC 6.4.1.2). Neither 12c-18:1 nor 9t,12t-18:2 produced the dramatic enzyme-lowering effect exhibited by the diet containing 9c,12c-18:2 when compared to the diet devoid of fat. Thus, both the 9 and 12 bonds must be present in the same molecule. Also, at least one and probably both bonds must be in the cis configuration to depress liver enzyme activities. Capillary gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) were both used for analysis of the methyl esters derived from the hepatic lipids. The GC and GC-MS data provided (a) direct evidence for incorporation of both isomers into hepatic lipids and (b) indirect evidence that 9t,12t-18:2 lowered liver delta 9-desaturase activity. In addition, since these products were found in the complex liver lipids, there is no doubt that the various enzymes concerned with activation and acylation utilize both of these isomeric fatty acids as substrates.     

Inhibition of stearoyl-CoA desaturase activity by the cis-9,trans-11 isomer and the trans-10,cis-12 isomer of conjugated linoleic acid in MDA-MB-231 and MCF-7 human breast cancer cells

Conjugated linoleic acid (CLA) is a collective term for a group of positional and geometric conjugated dienoic isomers of linoleic acid. CLA has been shown to have strong inhibitory effects on mammary carcinogenesis both in vitro and in vivo. In this study, we investigated the regulation of human stearoyl-CoA desaturase (SCD, EC 1.14.99.5) expression by CLA in human breast cancer cell lines, MDA-MB-231 and MCF-7. Treatment of the cells with the cis-9,trans-11 and trans-10,cis-12 CLA isomers (45 microM) did not repress SCD mRNA in both MDA-MB-231 and MCF-7 cells. However, the cis-9,trans-11 and trans-10,cis-12 CLA isomers significantly decreased SCD protein levels and SCD activity in MDA-MB-231 cells. In MCF-7 cells, both isomers did not affect protein levels, but they inhibited SCD activity. These results suggest that in MDA-MB-231 cells the cis-9,trans-11 and trans-10,cis-12 CLA isomers regulate human SCD by reducing SCD protein levels, while in MCF-7 cells both isomers have a direct inhibitory effect on SCD enzyme activity.     

Microsomal chemiluminescence of benzo[a]pyrene-7,8-dihydrodiol and its synthetic analogues trans- and cis-1-methoxyvinylpyrene

Low-density lipoproteins (LDL) have been shown to have a number of effects on the function of various cell types. To appreciate whether the in vitro effects of LDL have in vivo relevance, it is necessary to demonstrate that the biologic action described can be accounted for by native LDL and not by an alteration in the molecule or an addition to the preparation occurring during isolation. EDTA is frequently added to LDL during preparation to prevent oxidation. The effect of EDTA dialysis on LDL-mediated inhibition of lymphocyte responses was therefore examined. LDL alone did not inhibit mitogen-induced initial lymphocyte activation but rather suppressed lymphocyte DNA synthesis and subsequent proliferation in a transferrin-reversible manner. LDL dialysed with EDTA also inhibited lymphocyte responsiveness but the inhibition was not reversed by transferrin. Further experiments demonstrated that after dialysis EDTA in the LDL accounted for the change in its inhibitory effects. EDTA did not alter the lipoprotein but itself inhibited lymphocyte responses by chelating zinc necessary for DNA synthesis. These data indicate that LDL preparations may exhibit at least two separate effects on lymphocyte function. LDL is directly suppressive, while small amounts of contaminating EDTA can additionally be suppressive by chelating zinc. Thus, EDTA present in LDL preparations can alter their apparent biologic effects.     

Synthesis and reactivity of azobenzene-based bispropargyl sulfones: interesting comparison between cyclic and acyclic systems

Azobenzene-based bispropargyl bissulfone 3 containing stable E-azo moiety has been synthesized. Upon irradiation with long wavelength UV it isomerized to the Z-form 4, which can be thermally reisomerized to the E-isomer. Reactivity towards isomerization to the allenic system as well as DNA-cleaving efficiency under basic conditions was found to be significantly lower as compared to the previously synthesized cyclic sulfones 1 and 2. This lowering of reactivity can be explained in terms of low conversion to the allenic form and hence the lower extent of alkylation of DNA-bases, the only possible DNA-cleavage pathway for 3 and 4.     

In vivo distribution and turnover of trans- and cis-10-octadecenoic acid isomers in human plasma lipids

Triacylglycerols containing deuterium-labeled trans-10- and cis-10-octadecenoic acid (10t-18:1, 10c-18:1) plus the triacylglycerol of deuterated cis-9-octadecenoic acid (9c-18:1) were fed as a mixture to two young, adult male subjects. Analysis by mass spectroscopy of the labeled fats in blood samples collected periodically for 48 h allowed the uptake, distribution and turnover of both 10-octadecenoic acid isomers to be directly compared to 9c-18:1. A feature of this study is that actual weight data for the labeled fats were obtained. These data allowed plasma triacylglycerol turnover rates of 3.47-5.13 mg/min per kg to be estimated. Plasma and chylomicron triacylglycerol data also provided evidence that absorption of the deuterated fats mobilized 10-12 g of a triacylglycerol pool present in the intestinal cells. Other results are summarized as follows: the 10t-, 10c- and 9c-18:1 fatty acids were equally well absorbed, both delta 10-18:1 isomers were oxidized more rapidly than 9c-18:1, conversion of the delta 10-18:1 isomers into their corresponding 16:1 isomers was about 3-times faster than for 9c-18:1, the delta 10-18:1 isomers were preferentially incorporated at the 1-acyl and excluded from the 2-acyl position of phosphatidylcholine, esterification of cholesterol with the delta 10-18:1 fatty acids was 2.5-4.3-times slower than for 9c-18:1 and desaturation and elongation rates for the delta 10-18:1 acids were very low.