小花清风藤叶的化学成分研究

对于小花清风藤的化学成分和药理作用的研究目前较少报道,为了阐明小花清风藤的物质基础,该研究对小花清风藤(Sabia parviflora)的干燥叶,采用反复硅胶柱色谱、Sephadex LH-20柱色谱、制备薄层色谱及重结晶等手段进行分离纯化,运用化学分析和波谱学方法鉴定化合物的结构。结果表明:从小花清风藤干燥叶的甲醇超声提取物中进行分离共得到12个化合物,分别为N-反式阿魏酰酪胺(1)、N-顺式阿魏酰酪胺(2)、N-反式-对-香豆酰酪胺(3)、N-顺式-对-香豆酰酪胺(4)、N-反式-对-香豆酰章鱼胺(5)、N-顺式-对-香豆酰章鱼胺(6)、阿魏酸(7)、芹菜素(8)、木犀草素(9)、咖啡酸(10)、5-氧阿朴菲碱(11)、齐墩果酸(12)。其中,化合物2、4-9为首次从清风藤属植物中分离得到,化合物1、3、10为首次从该植物中分离得到。     

Dicarboxylic acid esters as transdermal permeation enhancers: Effects of chain number and geometric isomers

A series of transdermal permeation enhancers based on dicarboxylic acid esters was studied. Single-chain amphiphiles were markedly more effective than the double-chain ones. Monododecyl maleate, that is a cis derivative, was a more potent enhancer than its trans isomer, while the activity of succinates strongly depended on the donor vehicle. No difference between diastereoisomeric tartaric and meso-tartaric acid derivatives was found.     

Peroxidases and the metabolism of hydroxycinnamic acid amides in Poaceae

The arsenal of plants to fight off microorganisms and herbivores include hydroxycinnamic acid amides (HCAA) and their oxidation products. Hydroxycinnamic acid amides are widespread in the plant kingdom and in the recent years our knowledge of their biosynthesis and catabolism has increased substantially. Peroxidases are the primary candidates as the oxidative enzymes responsible for the turnover of hydroxycinnamic acid amide monomers. In barley, hydroxycinnamoylagmatine derivatives accumulate in young seedlings and in tissues infected with fungi. Hydroxycinnamoylagmatine is found as anti-fungal soluble dimers, called hordatines, and it is also a likely constituent of cell walls. Current evidence suggest that peroxidases are involved in the cross-linking of hydroxycinnamoylagmatine with cell wall components and possibly also in the synthesis of hordatines. Epidermal cell walls of barley respond to infection by the powdery mildew fungus with the deposition of polyphenolic material, that apparently contains hydroxycinnamic acid amides, at the site of attempted penetration. Accumulation of these compounds lowers the successful penetration by the fungus. The recent characterization of agmatine coumaroyl transferase (ACT), the N-hydroxycinnamoyltransferase responsible for the synthesis of hydroxycinnamoylagmatine in barley, has indicated that the production of these metabolites is widespread in the plant body and suggests multiple physiological functions for HCAA derivatives. The cloning of ACT has enabled the revelation of homologues genes in several monocots and the presence of a range of structurally diverse HCAAs in cereals suggests that their peroxidase-mediated metabolism is a common theme. The prospects for metabolic engineering of these pathways into other crops are discussed. Abbreviations: HCAA – hydroxycinnamic acid amide; HRPC – horseradish peroxidase C; ACT – agmatine coumaroyl transferase; THT – tyramine hydroxycinnamoyl transferase; HCBT – hydroxycinnamoyl/benzoyl-CoA:anthranilate N-hydroxycinnamoyl/benzoyl transferase; PHT – putrescine hydroxycinnamoyl transferase; SHT – spermidine/spermine hydroxycinnamoyl transferase; HHT – hydroxyanthranilate hydroxycinnamoyl transferase; p-CHA –p-coumaroyl hydroxyagmatine; p-CHDA –p-coumaroyl hydroxydehydroagmatine; PAL – phenylalanine ammonia lyase.     

Engineering of <Emphasis Type="Italic">Escherichia coli</Emphasis> for the synthesis of <Emphasis Type="Italic">N</Emphasis>-hydroxycinnamoyl tryptamine and serotonin

Plants synthesize various phenol amides. Among them, hydroxycinnamoyl (HC) tryptamines and serotonins exhibit antioxidant, anti-inflammatory, and anti-atherogenic activities. We synthesized HC–tryptamines and HC–serotonin from several HCs and either tryptamine or serotonin using Escherichia coli harboring the 4CL (4-coumaroyl CoA ligase) and CaHCTT [hydroxycinnamoyl-coenzyme A:serotonin N-(hydroxycinnamoyl)transferase] genes. E. coli was engineered to synthesize N-cinnamoyl tryptamine from glucose. TDC (tryptophan decarboxylase) and PAL (phenylalanine ammonia lyase) along with 4CL and CaHCTT were introduced into E. coli and the phenylalanine biosynthetic pathway of E. coli was engineered. Using this strategy, approximately 110.6 mg/L of N-cinnamoyl tryptamine was synthesized. By feeding 100 μM serotonin into the E. coli culture, which could induce the synthesis of cinnamic acid or p-coumaric acid, more than 99 μM of N-cinnamoyl serotonin and N-(p-coumaroyl) serotonin were synthesized.     

Evolution of rosmarinic acid biosynthesis

Rosmarinic acid and chlorogenic acid are caffeic acid esters widely found in the plant kingdom and presumably accumulated as defense compounds. In a survey, more than 240 plant species have been screened for the presence of rosmarinic and chlorogenic acids. Several rosmarinic acid-containing species have been detected. The rosmarinic acid accumulation in species of the Marantaceae has not been known before. Rosmarinic acid is found in hornworts, in the fern family Blechnaceae and in species of several orders of mono- and dicotyledonous angiosperms. The biosyntheses of caffeoylshikimate, chlorogenic acid and rosmarinic acid use 4-coumaroyl-CoA from the general phenylpropanoid pathway as hydroxycinnamoyl donor. The hydroxycinnamoyl acceptor substrate comes from the shikimate pathway: shikimic acid, quinic acid and hydroxyphenyllactic acid derived from l-tyrosine. Similar steps are involved in the biosyntheses of rosmarinic, chlorogenic and caffeoylshikimic acids: the transfer of the 4-coumaroyl moiety to an acceptor molecule by a hydroxycinnamoyltransferase from the BAHD acyltransferase family and the meta-hydroxylation of the 4-coumaroyl moiety in the ester by a cytochrome P450 monooxygenase from the CYP98A family. The hydroxycinnamoyltransferases as well as the meta-hydroxylases show high sequence similarities and thus seem to be closely related. The hydroxycinnamoyltransferase and CYP98A14 from Coleus blumei (Lamiaceae) are nevertheless specific for substrates involved in RA biosynthesis showing an evolutionary diversification in phenolic ester metabolism. Our current view is that only a few enzymes had to be “invented” for rosmarinic acid biosynthesis probably on the basis of genes needed for the formation of chlorogenic and caffeoylshikimic acid while further biosynthetic steps might have been recruited from phenylpropanoid metabolism, tocopherol/plastoquinone biosynthesis and photorespiration.     

The effect of trans-10, cis-12 conjugated linoleic acid on gene expression profiles related to lipid metabolism in human intestinal-like Caco-2 cells

We conducted an in-depth investigation of the effects of conjugated linoleic acid (CLA) on the expression of key metabolic genes and genes of known importance in intestinal lipid metabolism using the Caco-2 cell model. Cells were treated with 80 μmol/L of linoleic acid (control), trans-10, cis-12 CLA or cis-9, trans-11 CLA. RNA was isolated from the cells, labelled and hybridized to the Affymetrix U133 2.0 Plus arrays (n = 3). Data and functional analysis were preformed using Bioconductor. Gene ontology analysis (GO) revealed a significant enrichment (P < 0.0001) for the GO term lipid metabolism with genes up-regulated by trans-10, cis-12 CLA. Trans-10, cis-12 CLA, but not cis-9, trans-11 CLA, altered the expression of a number of genes involved in lipid transport, fatty acid metabolism, lipolysis, β-oxidation, steroid metabolism, cholesterol biosynthesis, membrane lipid metabolism, gluconeogenesis and the citrate cycle. These observations warrant further investigation to understand their potential role in the metabolic syndrome.     

13C-NMR of double and triple bond carbon atoms of unsaturated fatty acid methyl esters

The carbon magnetic resonance spectra of many fatty acid methyl esters with cis and trans double bonds and triple bonds at various positions and in many different combinations have been investigated.The influence of the ester group on double and triple bonds in the fatty acid chain depends strongly on the positions of these bonds. For a given position the influence is constant, even if one or more other double or triple bonds are present.Together with the evaluated chemical shift parameters for the effects of double and triple bonds on each other, complete assignments are possible and spectra of various types of unsaturated esters can be predicted with high accuracy (±0.1 ppm).     

Analysis of histidine and urocanic acid isomers by reversed-phase high-performance liquid chromatography

The qualitative separation performance of a C₁₈, C₈ and C₄ reversed-phase column was investigated for the separation of histidine and its metabolites histamine, 1-methyihistamine and trans- and cis-urocanic acid. Trans- and cis-urocanic acid were baseline separated from their precursor histidine on all three columns using isocratic elution with a mobile phase composed of 0.01 M aqueous TEAP pH 3.0 and acetonitrile at a ratio of 98:2 (v/v). However, histidine was not separated from histamine and 1-methyihistamine. Selecting the C₈ column and introducing 0.005 M of the ion pairing reagent 1-octanesulfonic acid sodium salt into the aqueous solution and acetonitrile at a ratio of 90:10 (v/v), significantly improved the separation. The separation was also followed by a change in the retention times and the order of elution. The sequence of elution was histidine, cis-urocanic acid, trans-urocanic acid, histamine and 1-methylhistamine with retention times of 5.58±0.07, 7.03±0.15, 7.92±0.18, 18.77±0.24 and 20.79±0.21 min (mean±SD; n=5). The separation on the C₈ column in the presence of the ion-pairing reagent was further improved with gradient elution that resulted in a reduction in the retention times and elution volumes of histamine and 1-methylhistamine. The detection limits of histidine and trans-urocanic acid at a wavelength of 210 nm and an injection volume of 0.05 ml were 5×10⁻⁸ mol l⁻¹ (n=3). The kinetic of the in-vitro conversion of trans- into the cis-isomer after UV irradiation was depending on the time of exposure and the energy of the light source. UVB light induced a significantly faster conversion than UVA light. TUCA and cUCA samples kept at −25°C were stable for up to 50 weeks. Samples, eluted from human skin showed various concentrations of histidine and trans- and cis-urocanic acid with an average of 1.69±0.33×10⁻⁵ mol l⁻¹, 1.17±0.43×10⁻⁵ mol l⁻¹ and 1.67±0.33×10⁻⁵ mol l⁻¹, respectively (n=8).     

A Novel Red Clover Hydroxycinnamoyl Transferase Has Enzymatic Activities Consistent with a Role in Phaselic Acid Biosynthesis

Red clover (Trifolium pratense) leaves accumulate several μmol g⁻¹ fresh weight of phaselic acid [2-O-(caffeoyl)-l-malate]. Postharvest oxidation of such o-diphenols to o-quinones by endogenous polyphenol oxidases prevents breakdown of forage protein during storage. Forage crops like alfalfa (Medicago sativa) lack both polyphenol oxidase and o-diphenols, and breakdown of their protein upon harvest and storage results in economic losses and release of excess nitrogen into the environment. Understanding how red clover synthesizes o-diphenols such as phaselic acid will help in the development of forage crops utilizing this natural system of protein protection. A possible pathway for phaselic acid biosynthesis predicts a hydroxycinnamoyl transferase (HCT) capable of forming caffeoyl and/or p-coumaroyl esters with malate. Genes encoding two distinct HCTs were identified in red clover. HCT1 shares more than 75% amino acid identity with a number of well-characterized shikimate O-HCTs implicated in monolignol biosynthesis. HCT2 shares only 34% amino acid sequence identity with HCT1 and has limited sequence identity to any previously identified HCT. Expression analyses indicate that HCT1 mRNA accumulates to 4-fold higher levels in stems than in leaves, whereas HCT2 mRNA accumulates to 10-fold higher levels in leaves than in stems. Activity assays of HCT1 and HCT2 proteins expressed in Escherichia coli indicate that HCT1 transfers caffeoyl or p-coumaroyl moieties from a coenzyme A-thiolester to shikimate but not malate, whereas HCT2 transfers caffeoyl or p-coumaroyl moieties from a coenzyme A-thiolester to malate but not shikimate. Together, these results indicate that HCT1 is involved in monolignol biosynthesis and HCT2 is a novel transferase likely involved in phaselic acid biosynthesis.In contrast to many other forage legumes (e.g. alfalfa [Medicago sativa]; Jones et al., 1995), red clover (Trifolium pratense) accumulates relatively high levels of the phenylpropanoid o-diphenol phaselic acid [2-O-(caffeoyl)-l-malic acid; hereafter referred to as caffeoyl-malate or phaselic acid] in its leaves (Hatfield and Muck, 1999; Winters et al., 2008). In red clover, upon cellular disruption, phaselic acid and other o-diphenols are readily oxidized by a soluble polyphenol oxidase (PPO) to produce their corresponding o-quinones (Hatfield and Muck, 1999; Sullivan et al., 2004). The formation of such o-quinones by PPO, and the subsequent secondary reactions of these quinones, are most often associated with browning of fresh fruits and vegetables (Steffens et al., 1994), which has a negative impact on perceived quality. When preserved by ensiling, however, oxidation of o-diphenols by PPO in red clover prevents degradation of protein during storage (Sullivan et al., 2004; Sullivan and Hatfield, 2006). Although alfalfa lacks significant levels of both PPO activity and o-diphenol compounds in its leaves, red clover''s natural system of protein protection has been transferred to this forage legume by expressing a red clover PPO transgene in alfalfa and exogenously adding o-diphenol PPO substrates to the resulting tissues or tissue extracts (Sullivan et al., 2004; Sullivan and Hatfield, 2006). Because ruminant animals poorly utilize degraded protein, adaptation of the PPO system to alfalfa and other forage crops would have substantial positive economic and environmental impacts (Sullivan and Hatfield, 2006). Unfortunately, lack of system components in these forage crops, especially the o-diphenol PPO substrates, presents a challenge to practical adaptation of this natural system of protein preservation. Consequently, understanding how red clover is able to accumulate o-diphenols such as phaselic acid will be a key step to adapt the PPO/o-diphenol system to a wide range of economically important forage crops.The biosynthetic pathways whereby red clover synthesizes and accumulates phaselic acid and other o-diphenols have not been defined. However, in the Brassicaceae, hydroxycinnamoyl esters with malic acid can be made via the action of sinapoyl-Glc:malate sinapoyltransferase (SMT; EC 2.3.1), which is capable of transferring a hydroxycinnamoyl moiety from a hydroxycinnamoyl-Glc ester to a malic acid acceptor. In Arabidopsis (Arabidopsis thaliana), SNG1 (for sinapoylglucose accumulator 1), which encodes the enzyme, has been shown to be responsible for the accumulation of sinapoylmalate in seeds and leaves (Lehfeldt et al., 2000). An SMT from radish (Raphanus sativus), presumably the homolog of the Arabidopsis SNG1 gene product, has been purified to apparent homogeneity and characterized (Grawe et al., 1992). The purified enzyme is capable of utilizing sinapoyl-, feruloyl-, caffeoyl-, and to a lesser extent p-coumaroyl-Glc esters to form the corresponding malic acid esters, suggesting that it is responsible for the accumulation of these esters in vivo. In contrast, in many plants, formation of certain hydroxycinnamoyl esters is often mediated by a member of the BAHD transferase family (D''Auria, 2006) that utilize a CoA thiolester hydroxycinnamoyl donor. Some of the best characterized of these hydroxycinnamoyl transferases (HCTs) are those associated with the biosynthesis of monolignols (Hoffmann et al., 2003, 2004; Shadle et al., 2007). These are capable of transferring p-coumaroyl or caffeoyl moieties from the respective CoA thiolesters to form 5-O-esters with shikimic acid or, to a lesser extent, 3-O-esters with quinic acid. Separable enzymatic activities capable of transferring a p-coumaroyl moiety to either shikimate/quinate or to 4′-hydroxyphenyllactate in basil (Ocimum basilicum) peltate gland extracts have been identified, although genes encoding these activities have not been cloned (Gang et al., 2002). Niggeweg et al. (2004) used gene-silencing experiments to definitively demonstrate that a hydroxycinnamoyl-CoA:quinate hydroxycinnamoyl transferase (HQT) is responsible for chlorogenic acid accumulation in the Solanaceae. Although phaselic acid biosynthesis in red clover could be via a pathway utilizing SMT, lack of an apparent SNG1 homolog in a collection of red clover EST sequences derived from leaves and young plants suggests otherwise (see “Discussion”). Therefore, pathways in red clover for the biosynthesis of phaselic acid utilizing one or more BAHD family transferase (Fig. 1) should be considered. In these proposed pathways, Phe would be converted to p-coumaroyl-CoA by the sequential action of Phe ammonia lyase (PAL), cinnamate-4-hydroxylase (C4H), and 4-coumarate:CoA ligase (4CL). The action of one or more specific HCTs and one or more p-coumarate 3′-hydroxylases (C3Hs) would then result in the formation of phaselic acid.Open in a separate windowFigure 1.Possible pathways for phaselic acid biosynthesis in red clover. Proposed pathway enzymes for the production of phaselic acid include PAL, 4CL, hydroxycinnamoyl:shikimate transferase (HCT-S), hydroxycinnamoyl:malate transferase (HCT-M), and C3H. The branch point at p-coumaroyl-CoA represents two alternative pathways. For simplicity, not all reactants and products are shown.Existing literature suggests that C3H enzymes, which are cytochrome P450 enzymes (CYP98A subfamily), do not directly hydroxylate p-coumaric acid to caffeic acid but rather act on p-coumaroyl ester derivatives. For example, the enzyme from Arabidopsis hydroxylates shikimic and quinic acid esters of p-coumaric acid but only poorly or not at all p-coumaric acid or its Glc or CoA esters (Schoch et al., 2001; Franke et al., 2002). Thus, one model of phaselic acid biosynthesis is the formation of 2-O-(p-coumaroyl)-l-malic acid (hereafter referred to as p-coumaroyl-malate) by a HCT and its subsequent hydroxylation by a C3H enzyme capable of utilizing the malic acid ester as a substrate (Fig. 1, bottom, red pathway). An alternative model would require at least two HCT activities for phaselic acid biosynthesis (Fig. 1, top, blue pathway). The first activity would form a substrate suitable for hydroxylation (e.g. p-coumaroyl-shikimate, since several characterized C3H enzymes appear to favor this substrate [Schoch et al., 2001; Franke et al., 2002; Gang et al., 2002; Morant et al., 2007]). Following hydroxylation to the caffeoyl derivative by a C3H, the first HCT activity could synthesize caffeoyl-CoA via its reverse reaction (Hoffmann et al., 2003; Niggeweg et al., 2004). A second HCT activity would then transfer the caffeoyl moiety to malic acid to form phaselic acid. Both pathways predict a transferase capable of transferring a hydroxycinnamoyl moiety (either p-coumaroyl or caffeoyl) to malic acid. Also, these pathways are consistent with the observation that, at least in vitro, several characterized HCT enzymes are capable of transfer reactions utilizing either p-coumaroyl- or caffeoyl-CoA (Hoffmann et al., 2003; Niggeweg et al., 2004). The identification and characterization of two distinct HCTs from red clover, one of which has properties consistent with a role in phaselic acid biosynthesis, are reported here.     

Cis-4-hydroxypipecolic acid and 2,4-cis-4,5-trans-4,5-dihydroxypipecolic acid from Calliandra

cis-4-Hydroxypipecolic acid and 2,4-cis-4,5-trans-4,5-dihydroxypipecolic acid were isolated from leaves of Calliandra pittieri. A system for resolving the eight imino acids isolated from Calliandra is described.     

13C-NMR of methyl,methylene and carbonyl carbon atoms of methyl alkenoates and alkynoates

The carbon magnetic resonance spectra of 102 fatty acid methyl esters with cis and trans double bonds and triple bonds at various positions and in many different combinations have been investigated. A comprehensive set of chemical shift parameters has been developed for the various substituents. With the aid of these parameters, the chemical shifts of all methyl, methylene carbonyl carbon atoms can be predicted with an accuracy of ±0.1 ppm or better.     

Geometric specificity of porcine liver carboxylesterase and its application for the production of cis-arylacrylic esters

Porcine liver carboxylesterase (carboxylic-ester hydrolase, EC 3.1.1.1) hydrolyses trans isomers of three different methyl 3-arylacrylates approximately one order of magnitude faster than the corresponding cis isomers. This phenomenon can be used for preparative production of cis esters from their trans counterparts as exemplified by methyl cinnamate. A solution of commercial, predominantly trans methyl cinnamate was irradiated by ultraviolet light and the resultant mixture of trans and cis esters was passed through a column packed with immobilized esterase. The effluent contained mainly trans cinnamic acid and cis methyl cinnamate. The latter was then extracted with methylene chloride, and the cis ester was isolated by evaporating the solvent. By esterifying the co-produced trans acid, the process can be made continuous.     

Formation of 12-oxo-trans-10-dodecenoic acid in chloroplasts from Thea sinensis leaves

Linolenic acid-[1-¹⁴C] was converted to 12-oxo-trans-10-dodecenoic acid, via 12-oxo-cis-9-dodecenoic acid by incubation with chloroplasts of Thea sinensis leaves. Thus, it was confirmed that linolenic acid is split into a C₁₂-oxo-acid, 12-oxo-trans-10-dodecenoic acid, and a C₆-aldehyde, trans-2-hexenal, leaf aldehyde, by an enzyme system in chloroplasts of tea leaves.     

Purification and Properties of Hydroxycinnamic Acid Ester Hydrolase from Aspergillus japonicus

Hydroxycinnamic acid ester hydrolase from the wheat bran culture medium of Aspergillus japonicus was purified 255-fold by ammonium sulfate fractionation, DEAE-Sephadex treatment and column chromatographies on DEAE-Sephadex, CM-Sephadex and various other Sephadexes. The purified enzyme was free from tannase and found to be homogeneous on polyacrylamide disc gel electrophoresis. Its molecular weight was estimated to be 150,000 by gel filtration and 142,000 by SDS-gel electrophoresis. The isoelectric point of the enzyme was pH 4.80. As to its amino acid composition, aspartic acid and glycine were abundant. The optimum pH and temperature for the enzyme reaction were, respectively, 6.5 and 55°C when chlorogenic acid was used as a substrate. The enzyme was stable between pH 3.0 to 7.5 and inactivated completely by heat treatment at 70°C for 10 min.

All metal ions examined did not activate the enzyme, while Hg⁺⁺ reduced its activity. The enzyme was markedly inhibited by diisopropylfluorophosphate and an oxidizing reagent, iodine, although it was not affected so much by metal chelating or reducing reagents. The purified enzyme hydrolyzed not only esters of hydroxycinnamic acids such as chlorogenic acid, caffeoyl tartaric acid and p-coumaroyl tartaric acid, but also ethyl and benzyl esters of cinnamic acid. However, the enzyme did not act on ethyl esters of crotonic acid and acrylic acid or esters of hydroxybenzoic acids.     

Seasonal variations in trans-2-hexenal and linolenic acid in homogenates of Thea sinensis leaves

The seasonal variations in the amounts of C₆-volatile components cis-3-hexenal trans-2-hexenal n-hexanal) and their precursors (linoleic and linolenic acid) in homogenates of Thea sinensis leaves were quantitatively analyzed throughout the year. Formation of trans-2-hexenal began in the middle of April and reached a maximum during July. Towards autumn the aldehyde gradually decreased and, in winter (December to March), was virtually absent. The levels of cis-3-hexenol remained constant during May–December. cis-3-Hexenal showed a similar variation pattern to that of trans-2-hexenal. The major fatty acids in the leaves were palmitic, palmitoleic, oleic, linoleic and linolenic acid, and occurred in non-ionic lipids and phospholipid fractions. The amounts of linoleic and linolenic acid did not show any marked variation except for a big peak in October.     

Phytochemical variation within populations of Echinacea angustifolia (Asteraceae)

Quantitative evaluation of phytochemical diversity in Echinacea angustifolia DC. populations from different natural geographic areas supports the existence of distinct natural chemotypes within the species. Consumers, growers and manufacturers of phytomedicines are interested in chemotype identification for prediction of phytochemical content in cultivar development. Six month old E. angustifolia roots, grown from nine different wild seed sources in a controlled environment, were extracted into 70% ethanol and 28 reported phytochemicals were measured by HPLC separation. Two-way ANOVA between the nine populations revealed quantitative differences (p<0.05) in the caffeic acid derivatives 2,3-O-dicaffeoyl tartaric acid (cichoric acid), 2-O-caffeoyl tartaric acid (caftaric acid), 1,3-dicaffeoyl-quinic acid (cynarin), echinacoside and ten reported alkamides. Canonical discriminant analysis determined the phytochemical variables which contributed the most towards chemotype distinction for five of the nine populations: undeca-2E,4Z-diene-8,10-diynoic acid-2-methylbutylamide¹, dodeca-2E,4E-dienoic acid isobutylamide¹, dodeca-2E-ene-8,10-diynoic acid isobutylamide⁷, hexadeca-2E,9Z-diene-12,14-diynoic acid isobutylamide¹, cichoric acid⁷, caftaric acid¹, and echinacoside⁷ (¹p<0.0001, ⁷p<0.05). Five of those compounds were also significantly associated with latitudinal variation by regression analyses (p<0.05).     

Cis-5-hydroxy-L-pipecolic acid from Morus alba and Lathyrus japonicus

cis-5-Hydroxy-L-pipecolic acid was isolated and characterized from the leaves of Morus alba and the seeds of Lathyrus japonicus. The trans-form was also obtained from the former.     

Accumulation of conjugated linoleic acid in <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> WU-P19 is enhanced by induction with linoleic acid and chitosan treatment

Production of conjugated linoleic acid (CLA) by the potential probiotic bacterium Lactobacillus plantarum WU-P19 was investigated with the aim of enhancing production. CLA produced using this bacterium may be used to supplement dietary intake. Cultures were fed linoleic acid for conversion to CLA and the CLA produced was measured. In some cases, chitosan was added to cultures to improve cellular uptake of linoleic acid. Under static conditions at 37 °C, the bacterium grew and produced CLA in the pH range of 5.5–6.5. At pH 6.0, a 36-h incubation period maximized the concentration of the dry biomass (0.82 g/L), the CLA content in the biomass (4.1 mg/g), and linoleic acid in the biomass (1.2 mg/g). In comparison with cultures grown without linoleic acid in the medium, supplementing the medium with linoleic acid at 600 μg/mL slowed the production of CLA, but the CLA content in the dry biomass increased to 12–14 mg/g and the linoleic acid content increased to 8–11 mg/g. Supplementing the culture medium with chitosan and linoleic acid enhanced production of CLA in the dry biomass to 21 mg/g within 36 h. Nearly 50% of the CLA was cis-9, trans-11-CLA, and the remainder was trans-10, cis-12-CLA. Linoleic acid content of the dry biomass was increased to 37 mg/g. Accumulation of CLA in the cells was enhanced by feeding linoleic acid. Supplementing the culture with linoleic acid and chitosan further increased accumulation of CLA.     

Epoxyoctadecadienoic acids from Crepis conyzaefolia seed oil

The seed oil of Crepis conyzaefolia (Gouan) Dalle Torre contains previously unidentified (±)-cis-12,13-epoxyoctadeca-trans-6-cis-9-dienoic (14%) and cis-12,13-epoxyoctadeca-cis-6-cis-9-dienoic (2%) acids and the more common vernolic [(±)-12,13-epoxyoctadec-cis-9-enoic] (32%) acid.     

Biosynthesis of 9-cis-Retinoic Acid from 9-cis-β-Carotene in Human Intestinal Mucosa in Vitro

The role of 9-cis-β-carotene (9-cis-β-C) as a potential precursor of 9-cis-retinoic acid (9-cis-RA) has been examined in human intestinal microcosa in vitro. By using HPLC, uv spectra, and chemical derivatization analysis, both 9-cis-RA and all-trans-retinoic acid (all-trans-RA) have been identified in the postnuclear fraction of human intestinal microcosa after incubation with 9-cis-β-C at 37°C. The biosynthesis of both 9-cis-RA and all-trans-RA from 9-cis-β-C was linear with increasing concentrations of 9-cis-β-C (2-30 μM) and was linear with respect to tissue protein concentration up to 0.75 mg/ml. Retinoic acid was not detected when a boiled incubation mixture was incubated in the presence of 9-cis-β-C. The rate of synthesis of 9-cis- and all-trans-RA from 4 μM 9-cis-β-C were 16 ± 1 and 18 ± 2 pmol/hr/mg of protein, respectively. However, when 2 μM all-trans-β-C was added to the 4 μM 9-cis-β-C, the rate of all-trans-RA synthesis was increased to 38 ± 6 pmol/hr/mg of protein, whereas the rate of 9-cis-RA synthesis remained the same. These results suggest that 9-cis-RA is produced directly from 9-cis-β-C. Furthermore, incubations of either 0.1 μM 9-cis- or all-trans-retinal under the same incubation conditions showed that 9-cis-RA could also arise through oxidative conversion of 9-cis-retinal. Although only 9-cis-RA was detected when 9-cis-RA was used as the substrate, the isomerization of the all-trans-RA to 9-cis-RA cannot be ruled out, since both all-trans-RA and trace amounts of 9-cis-RA were detected when all-trans-retinal was incubated as the substrate. These data indicate that 9-cis-β-C can be a source of 9-cis-RA in the human. This conversion may have a significance in the anticarcinogenic action of β-C.