Stereoselective synthesis of lanthionine derivatives in aqueous solution and their incorporation into the peptidoglycan of Escherichia coli

The three diastereoisomers—(R,R), (S,S) and meso—of lanthionine were synthesized in aqueous solution with high diastereoselectivity (>99%). The (S) and (R) enantiomers of two differently protected sulfamidates were opened by nucleophilic attack of (R) or (S)-cysteine. Acidification and controlled heating liberated the free lanthionines. Using the same chemistry, an α-benzyl lanthionine was also prepared. The proposed method, which avoids the need of enrichment by recrystallization, opens the way to the labelling of these compounds with ³⁵S. Furthermore, in vivo bioincorporation into Escherichia coli W7 was studied. No incorporation of α-benzyl lanthionine was observed. In contrast, meso-lanthionine can effectively replace meso-diaminopimelic acid in vivo, while in the presence of (R,R)-lanthionine the initial increase of bacterial growth was followed by cell lysis. In the future, meso-[³⁵S]lanthionine could be used to study the biosynthesis of peptidoglycan and its turnover in relation to cell growth and division.     

Isolation of a bacterium assimilating (R)-3-chloro-1,2-propanediol and production of (S)-3-chloro-1,2-propanediol using microbial resolution

A bacterium capable of assimilating 3-chloro-1,2-propanediol was isolated from soil by enrichment culture. The strain was identified as Alcaligenes sp. by taxonomic studies. The crude extracts of the cells had dehalogenating activities and converted various halohydrins to the corresponding epoxides. 3-Chloro-1,2-propanediol was degraded stereospecifically by the strain, liberating chloride ion. The residual isomer was found to be the (S)-form (99.4% enantiomeric excess). (S)-3-Chloro-1,2-propanediol was obtained from the racemate by use of this strain in 38% yield, and (S)-glycidol (99.4% enantiomeric excess) was subsequently synthesized from the obtained (S)-3-chloro-1,2-propanediol by alkaline treatment.     

Characterization of CHO-K1 cells stably expressing PDE-IV enzymes

A CHO-K1 cell line stably expressing a recombinant full-length human PDE-IVa (HSPDE4A4B) enzyme was established under hygromycin B selection. Full-length expression of the protein was determined by Western blot analysis, which revealed the presence of a 125-kDa immunoreactive band using rabbit anti-PDE-IVa antibodies. The potency of inhibitor compounds was examined by their ability to increase cAMP in the whole-cell, and by their ability to inhibit cAMP hydrolysis in a 100,000g supernatant (soluble enzyme preparation) obtained from the same cell line. Inhibition of the expressed PDE-IVa activity by selective PDE-IV inhibitors—(R) and (S)-rolipram, RS 14203, and CDP 840—at 100 nM substrate demonstrated that RS 14203 and CDP 840 were the most potent with IC₅₀=9 nM, followed by (R)-rolipram (IC₅₀=110 nM) and (S)-rolipram (IC₅₀=420 nM). The rank order of potencies of the inhibitors in elevating cAMP in the whole-cell assay was quite different from that on the soluble enzyme. RS 14203 was still the most potent compound in elevating cAMP. Moreover, the relative rank order of potencies between CDP 840 and (R)-rolipram changed dramatically, such that (R)-rolipram was more potent than CDP 840 = (S)-rolipram. An apparent 30-fold stereoselectivity between (R)- and (S)-rolipram was also noted. The whole-cell rank order of potencies was also maintained when PKA activity ratios were measured in place of cAMP levels. The ability of the compounds to elevate cAMP in the stable CHO-K1 cells appeared to track better with the potency of the compounds against the high-affinity (Sr) conformer of the enzyme rather than the low-affinity catalytic state.     

Substrate Specificity and Enantioselectivity of 4-Hydroxyacetophenone Monooxygenase

The 4-hydroxyacetophenone monooxygenase (HAPMO) from Pseudomonas fluorescens ACB catalyzes NADPH- and oxygen-dependent Baeyer-Villiger oxidation of 4-hydroxyacetophenone to the corresponding acetate ester. Using the purified enzyme from recombinant Escherichia coli, we found that a broad range of carbonylic compounds that are structurally more or less similar to 4-hydroxyacetophenone are also substrates for this flavin-containing monooxygenase. On the other hand, several carbonyl compounds that are substrates for other Baeyer-Villiger monooxygenases (BVMOs) are not converted by HAPMO. In addition to performing Baeyer-Villiger reactions with aromatic ketones and aldehydes, the enzyme was also able to catalyze sulfoxidation reactions by using aromatic sulfides. Furthermore, several heterocyclic and aliphatic carbonyl compounds were also readily converted by this BVMO. To probe the enantioselectivity of HAPMO, the conversion of bicyclohept-2-en-6-one and two aryl alkyl sulfides was studied. The monooxygenase preferably converted (1R,5S)-bicyclohept-2-en-6-one, with an enantiomeric ratio (E) of 20, thus enabling kinetic resolution to obtain the (1S,5R) enantiomer. Complete conversion of both enantiomers resulted in the accumulation of two regioisomeric lactones with moderate enantiomeric excess (ee) for the two lactones obtained [77% ee for (1S,5R)-2 and 34% ee for (1R,5S)-3]. Using methyl 4-tolyl sulfide and methylphenyl sulfide, we found that HAPMO is efficient and highly selective in the asymmetric formation of the corresponding (S)-sulfoxides (ee >99%). The biocatalytic properties of HAPMO described here show the potential of this enzyme for biotechnological applications.     

Enantioselective synthesis of (S)-phenylephrine by whole cells of recombinant Escherichia coli expressing the amino alcohol dehydrogenase gene from Rhodococcus erythropolis BCRC 10909

(R)-phenylephrine [(R)-PE] is an α₁-adrenergic receptor agonist that is widely used in over-the-counter drugs to treat the common cold. We found that Rhodococcus erythropolis BCRC 10909 can convert detectable level of 1-(3-hydroxyphenyl)-2-(methylamino) ethanone (HPMAE) to (S)-PE by high performance liquid chromatography tandem mass spectrometry analysis. An amino alcohol dehydrogenase gene (RE_AADH) which possesses the ability to convert HPMAE to (S)-PE was then isolated from R. erythropolis BCRC 10909 and expressed in Escherichia coli NovaBlue. The purified RE_AADH, tagged with 6×His, had a molecular mass of approximately 30 kDa and exhibited a specific activity of 0.19 μU/mg to HPMAE in the presence of NADPH, indicating this enzyme could be categorized as NADP⁺-dependent short-chain dehydrogenase reductase. E. coli NovaBlue cell expressing the RE_AADH gene was able to convert HPMAE to (S)-PE with more than 99% enantiomeric excess (ee), 78% yield and a productivity of 3.9 mmol (S)-PE/L h in 12 h at 30 °C and pH 7. The (S)-PE, recovered from reaction mixture by precipitation at pH 11.3, could be converted to (R)-PE (ee > 99%) by Walden inversion reaction. This is the first reported biocatalytic process for the production of (S)-PE from HPMAE.     

Glycosylation of 2-phenylpropionic acid and its ethyl ester in suspension cultures of Nicotiana tabacum,Dioscoreophyllum cumminsii and Aconitum japonicum

Suspension cultures of Nicotiana tabacum and Dioscoreophyllum cumminsii converted 2-(RS)-phenylpropionic acid and its ethyl ester into 2-(RS)-phenylpropionyl β-D-glucopyranoside, 2-(RS)-phenylpropionyl 6-O-β-D-glucopyranosyl-β-D-glucopyranoside and 6-O-β-D- glucopyranosyl-2-O-[2-(RS)-phenylpropionyl]-D-glucose which accumulated in the cells. A suspension culture of Aconitum japonicum converted these substrates into ethyl 6-O-[2-(RS)-phenylpropionyl]- β-D-glucopyranoside which was mostly excreted into the medium. The diastereomeric mixture of the glucosyl esters of 2-(RS)-phenylpropionic acid was resolved by HPLC to show the ratio of R:S was 1:1.     

Didymosphaeria igniaria: a new microorganism useful for the enantioselective reduction of aryl-aliphatic ketones

Didymosphaeria igniaria is a promising biocatalyst in asymmetric reductions of prochiral aromatic-aliphatic ketones such as acetonaphthones, acetophenones, and acetylpyridines. The organism converted the substrates mainly to (S)-alcohols. Excellent results in terms of conversion and enantioselectivity (100% yield, >99% ee) were obtained with acetonaphthones. In case of acetyl pyridines, the optical purity of the product depended on the position of the carbonyl group on the pyridine ring and followed the order 2-acetyl ? 4-acetyl > 3-acetyl-pyridine. Transformation of o-methoxy-acetophenone gave optically pure (S)-(-)-1-(2-methoxyphenyl)-ethanol in 95% yield. The transformation of para-methyl ketone gave (R)-alcohol (81% ee), whereas para-bromo ketone gave (S)-alcohol (98% ee). Monitoring of the biotransformation of these substrates over time led to the conclusion that for both substrates, non-selective carbonyl group reduction occurred in the first step, followed by selective oxidation of the (R)-isomer of p-bromo-phenylethanol and selective oxidation of the (S)-isomer of p-methyl-phenylethanol. D. igniaria exhibited poor enantioselectivity in the reduction of bicyclic aryl-aliphatic ketones such as 1- and 2-tetralones. Only (S)-5-methoxy-1-tetralol was obtained in optically pure (>99% ee) form.     

Microbial Production of Aliphatic (S)-Epoxyalkanes by Using Rhodococcus sp. Strain ST-10 Styrene Monooxygenase Expressed in Organic-Solvent-Tolerant Kocuria rhizophila DC2201

We describe the development of biocatalysis for producing optically pure straight-chain (S)-epoxyalkanes using styrene monooxygenase of Rhodococcus sp. strain ST-10 (RhSMO). RhSMO was expressed in the organic solvent-tolerant microorganism Kocuria rhizophila DC2201, and the bioconversion reaction was performed in an organic solvent-water biphasic reaction system. The biocatalytic process enantioselectively converted linear terminal alkenes to their corresponding (S)-epoxyalkanes using glucose and molecular oxygen. When 1-heptene and 6-chloro-1-hexene were used as substrates (400 mM) under optimized conditions, 88.3 mM (S)-1,2-epoxyheptane and 246.5 mM (S)-1,2-epoxy-6-chlorohexane, respectively, accumulated in the organic phase with good enantiomeric excess (ee; 84.2 and 95.5%). The biocatalysis showed broad substrate specificity toward various aliphatic alkenes, including functionalized and unfunctionalized alkenes, with good to excellent ee. Here, we demonstrate that this biocatalytic system is environmentally friendly and useful for producing various enantiopure (S)-epoxyalkanes.     

Condensed tannins. Optically active diastereoisomers of (+)-mollisacacidin by epimerization

1. (+)-Mollisacacidin [(+)-3′,4′,7-trihydroxy-2,3-trans-flavan-3,4-trans- diol] is converted by autoclaving into the optically active free phenolic 2,3-trans-3-4-cis (12% yield), 2,3-cis-3,4-trans (11%) and 2,3-cis-3,4-cis (2·8%) diastereoisomers through epimerization at C-2 and C-4. 2. The relative configurations of the epimeric forms were determined by nuclear-magnetic-resonance spectrometry and paper ionophoresis in comparison with synthetic reference compounds, and was confirmed by chemical interconversions. 3. From this a scheme of epimerization is inferred and their absolute configurations are assigned as (2R:3S:4S), (2S:3S:4R) and (2S:3S:4S) respectively from the known absolute configuration (2R:3S:4R) of (+)-mollisacacidin.     

Rhodococcus erythropolis DCL14 Contains a Novel Degradation Pathway for Limonene

Strain DCL14, which is able to grow on limonene as a sole source of carbon and energy, was isolated from a freshwater sediment sample. This organism was identified as a strain of Rhodococcus erythropolis by chemotaxonomic and genetic studies. R. erythropolis DCL14 also assimilated the terpenes limonene-1,2-epoxide, limonene-1,2-diol, carveol, carvone, and (−)-menthol, while perillyl alcohol was not utilized as a carbon and energy source. Induction tests with cells grown on limonene revealed that the oxygen consumption rates with limonene-1,2-epoxide, limonene-1,2-diol, 1-hydroxy-2-oxolimonene, and carveol were high. Limonene-induced cells of R. erythropolis DCL14 contained the following four novel enzymatic activities involved in the limonene degradation pathway of this microorganism: a flavin adenine dinucleotide- and NADH-dependent limonene 1,2-monooxygenase activity, a cofactor-independent limonene-1,2-epoxide hydrolase activity, a dichlorophenolindophenol-dependent limonene-1,2-diol dehydrogenase activity, and an NADPH-dependent 1-hydroxy-2-oxolimonene 1,2-monooxygenase activity. Product accumulation studies showed that (1S,2S,4R)-limonene-1,2-diol, (1S,4R)-1-hydroxy-2-oxolimonene, and (3R)-3-isopropenyl-6-oxoheptanoate were intermediates in the (4R)-limonene degradation pathway. The opposite enantiomers [(1R,2R,4S)-limonene-1,2-diol, (1R,4S)-1-hydroxy-2-oxolimonene, and (3S)-3-isopropenyl-6-oxoheptanoate] were found in the (4S)-limonene degradation pathway, while accumulation of (1R,2S,4S)-limonene-1,2-diol from (4S)-limonene was also observed. These results show that R. erythropolis DCL14 metabolizes both enantiomers of limonene via a novel degradation pathway that starts with epoxidation at the 1,2 double bond forming limonene-1,2-epoxide. This epoxide is subsequently converted to limonene-1,2-diol, 1-hydroxy-2-oxolimonene, and 7-hydroxy-4-isopropenyl-7-methyl-2-oxo-oxepanone. This lactone spontaneously rearranges to form 3-isopropenyl-6-oxoheptanoate. In the presence of coenzyme A and ATP this acid is converted further, and this finding, together with the high levels of isocitrate lyase activity in extracts of limonene-grown cells, suggests that further degradation takes place via the β-oxidation pathway.     

2(S),4(R)-4-(β-d-Galactopyranosyloxy)-4-isobutyl-glutamic acid: A new amino acid in Reseda odorata

2(S),4(R)-4-(β-d-Galactopyranosyloxy)-4-isobutylglutamic acid (I) has been isolated from the flowers of Reseda odorata, wherein it occurs in substantial quantity. Hydrolysis of I gives d-galactose, 2(S),4(R)-4-hydroxy-4-isobutylglutamic acid (II) and 3(R),5(S)-3-hydroxy-3-isobutyl-2-pyrrolidone-5-carboxylic acid (III) and its treatment with nitrous acid yields a galactoside of a non-nitrogenous hydroxy acid lactone (IV). The structures of I and its degradation products are supported by PMR, ¹³C-NMR and other spectroscopic methods. ¹³C-NMR spectroscopy of the model compound 2-(β-d-galactopyranosyloxy)isobutyric acid confirmed the structure of the natural product. The S- (or l-) configuration at C(2) in the amino acid moiety of I has been established by the use of the Clough—Lutz—Jirgenson rule and the R-configuration at C(4) of the same unit has been assigned tentatively. I represents the first example of a glycoside of a higher plant amino acid in which the carbohydrate residue is linked to an aliphatic hydroxy group.     

The conversion of (24S)-24-ethylcholesta-5,22,25-trien-3β-ol into poriferasterol,both in vivo and with a cell-free homogenate of the alga Trebouxia sp.

[6-³H₁] (24S)-24-Ethylcholesta-5,22,25-trien-3β-ol added to the growth medium of a culture of Trebouxia sp. 213/3 was efficiently taken-up by the cells and converted into (24R)-24-ethylcholesta-5,22-dien-3β-ol (poriferasterol) which is one of the major sterols of this alga. A cell-free homogenate was obtained from Trebouxia which catalysed the NADPH-dependent reduction of [6-³H₁] (24S)-24-ethylcholesta-5,22,25-trien-3β-ol to yield poriferasterol. The δ²⁵-sterol reductase was found to be mainly localized in the microsomal fraction of the homogenate.     

Biochemical Characterization of the Oxygenation of Unsaturated Fatty Acids by the Dioxygenase and Hydroperoxide Isomerase of Pseudomonas aeruginosa 42A2

We have studied oxygenation of fatty acids by cell extract of Pseudomonas aeruginosa 42A2. Oleic acid ((9Z)-18:1) was transformed to (10S)-hydroperoxy-(8E)-octadecenoic acid ((10S)-HPOME) and to (7S,10S)-dihydroxy-(8E)-octadecenoic acid (7,10-DiHOME). Experiments under oxygen-18 showed that 7,10-DiHOME contained oxygen from air and was formed sequentially from (10S)-HPOME by isomerization. (10R)-HPOME was not isomerized. The (10S)-dioxygenase and hydroperoxide isomerase activities co-eluted on ion exchange chromatography and on gel filtration with an apparent molecular size of ∼50 kDa. 16:1n-7, 18:2n-6, and 20:1n-11 were also oxygenated to 7,10-dihydroxy fatty acids, and (8Z)-18:1 was oxygenated to 6,9-dihydroxy-(7E)-octadecenoic acid. A series of fatty acids with the double bond positioned closer to ((6Z)-18:1, (5Z,9Z)-18:2) or more distant from the carboxyl group ((11Z)-, (13Z)-, and (15Z)-18:1) were poor substrates. The oxygenation mechanism was studied with [7S-²H]18:1n-9, [7R-²H]18:2n-6, and [8R-²H]18:2n-6 as substrates. The pro-R hydrogen at C-8 was lost in the biosynthesis of (10S)-HPODE, whereas the pro-S hydrogen was lost and the pro-R hydrogen was retained at C-7 during biosynthesis of the 7,10-dihydroxy metabolites. Analysis of the fatty acid composition of P. aeruginosa revealed relatively large amounts of (9E/Z)-16:1 and (11E/Z)-18:1 and only traces of 18:1n-9. We found that (11Z)-18:1 (vaccenic acid) was transformed to (11S,14S)-dihydroxy-(12E)-octadecenoic acid and to a mixture of 11- and 12-HPOME, possibly due to reverse orientation of (11Z)-18:1 at the active site compared with oleic acid. The reaction mechanism of the hydroperoxide isomerase suggests catalytic similarities to cytochrome P450.     

Stereoselectivity of the interactions of thioglucoside glucohydrolase and epithiospecifier protein from various sources

(S)-2-Hydroxy-3-butenylglucosinolate (epi-progoitrin) was converted to erythro- and threo-1-cyamo-2-hydroxy-3,4-epithiobutanes by combinations of thioglucoside glucohydrolase and epithiospecifier protein (ESP) from various sources. In all combinations studied the erythro/threo product ratio was about 1.4, except when the thioglucoside glucohydrolase was from Sinapis alba L. and the ESP from Brassica campestris in which case the ratio was about 1.8.     

Formation of (-⊃-1′,2′-epi-2-cis-xanthoxin acid from a precursor of abscisic acid

Tomato shoots and avocado mesocarp supplied with (±)-[2-¹⁴C]-5-(1,2-epoxy-2,6,6-trimethylcyclohexyl)-3-methylpenta-cis-2-trans-4-dienoic acid metabolize it into (+)-abscisic acid and a more polar material that was isolated and identified as (?)-epi-1′(R),2′(R)-4′(S)-2-cis-xanthoxin acid. The (+)-1′(S),2′(S)-4′(S)-2-cis-xanthoxin acid recently synthesized from natural violaxanthin, has the 1′,2′-epoxy group on the opposite side of the ring to that of the 4′(S)-hydroxyl group and the compound is rapidly converted into (+)-abscisic acid. The 1′,2′-epoxy group of (?)-1′,2′-epi-2-cis-xanthoxin acid is on the same side of the ring as the 4′(S) hydroxyl group: the compound is not metabolized into abscisic acid. The configuration of the 1′,2′-epoxy group probably controls whether or not the 4′(S) hydroxyl group can be oxidized. (+)-2-cis-Xanthoxin acid is probably not a naturally occurring intermediate because a ‘cold trap’, added to avocado fruit forming [¹⁴C]-labelled abscisic acid from [2-¹⁴C]mevalonate, failed to retain [¹⁴C] label.     

Structure and functions of γ-dodecalactone isolated from Antrodia camphorata for NK cell activation

The preserved fungal species Antrodia camphorata has diverse health-promoting effects and has been popularly used in East Asia as a traditional herb. We isolated a volatile compound from the culture medium of A. camphorata and identified it as γ-dodecalactone (γ-DDL). Cytomic screening for immune-modulating activity revealed that γ-DDL can activate human NK cells to express the early activation marker CD69. Further experiments showed that γ-DDL not only can induce NK cells to express CD69 but also stimulate NK cells to secrete cytotoxic molecules (FasL and granzyme B) and Th1 cytokines (TNF-α and INF-γ).Measuring the distribution of γ-DDL in the subcellular compartments of NK cells revealed that γ-DDL has been converted to 4-hydroxydodecanoic acid (an acyclic isomer of γ-DDL) in a time-dependent manner in the cytoplasm.Synthetic (R,S)-4-hydroxydodecanoic acid activated NK cells to express CD69 mRNA within 10 min, in contrast to γ-DDL, which activated NK cells to express CD69 within 50 min. This faster activation suggests that γ-DDL has converted to 4-hydroxydodecanoic acid and to stimulate the NK cells to express CD69.Optically pure (R)-(+)-4-hydroxydodecanoic acid and (S)-(?)-4-hydroxydodecanoic acid were obtained via: (1) synthesis of its diastereomeric esters of (R,S)-4-hydroxydodecanoic (R)-(?)-2-phenylpropionate; (2) separation of diastereomers via preparative HPLC, and (3) subsequent hydrolysis of the obtained optical pure ester of (R)-(+)-4-hydroxydodecanoic acid (R)-(?)-2-phenylpropionate and (R)-(?)-4-hydroxydodecanoic acid (R)-(?)-2-phenylpropionate, respectively. Further assays of NK cells activation using each enantiomer showed that only the (R)-(+)-4-hydroxydodecanoic acid can activate NK cells.     

Oxidative decarboxylation of mandelic acid derivative by recombinant Escherichia coli: a novel method of ethyl vanillin synthesis

The benzoylformate decarboxylase gene (mdlC) from Pseudomonas putida was expressed in Escherichia coli BL21(DE3). The recombinant strain together with E. coli/pET30a-mdlB converted (S)-3-ethoxy-4-hydroxymandelic acid (S-EMA) into ethyl vanillin without ethyl vanillin degradation. 4 g ethyl vanillin/l was obtained from 10 g EMA/l within 12 h at 30 °C. This is the first report on the biotransformation of (S)-EMA to ethyl vanillin.     

Stereochemical Control in the Still-Wittig Rearrangement Synthesis of Cyclohexyl (Z)-Alkene Inhibitors of Pin1

Three stereoisomeric inhibitors of Pin1: (2R,5S)-, (2S,5R)- and (2S,5S)-Ac–pSer–Ψ[(Z)CH = C]–pipecolyl(Pip)–2-(2-naphthyl)ethylamine 1, that mimic L-pSer–D-Pro, D-pSer–L-Pro, and D-pSer–D-Pro amides respectively, were synthesized by a 13-step route. The newly formed stereogenic centers in the pipecolyl ring were introduced by Luche reduction, followed by stereospecific [2,3]-Still-Wittig rearrangement. The (Z)- to (E)-alkene ratio in the rearrangements were consistently 5.5 to 1. The stereochemistry at the original Ser α-carbon controlled the stereochemistry of the Luche reduction, but it did not affect the stereochemical outcome of the rearrangement, which consistently gave the (Z)-alkene. The epimerized by-product, (2S,5S)-10, resulting from the work-up after Na/NH₃ debenzylation of (2S,5R)-9, was carried on to the (2S,5S)-1 isomer. Compound (2S,5S)-10 was resynthesized from the Luche reduction by-product, (2R,3R)-3, and the stereochemistry was confirmed by comparison of the optical rotations. The IC₅₀ values for (2R,5S)-1, (2S,5R)-1 and (2S,5S)-1 Pin1 inhibition were: 52, 85, and 140 μM, respectively.     

Preparation of optically-active 3-pyrrolidinol and its derivatives from 1-benzoylpyrrolidinol by hydroxylation with Aspergillus sp. and stereo-selective esterification by a commercial lipase

An effective preparation scheme for optically-active 3-pyrrolidinol and its derivatives based on biological transformation is proposed. Aspergillus sp. NBRC 109513 hydroxylated 1-benzoylpyrrolidine, yielding (S)-1-benzoyl-3-pyrrolidinol with 66 % ee. Kinetic resolution of 1-benzoyl-3-pyrrolidinol by Amano PS-IM lipase formed optically-active 1-benzoyl-3-pyrrolidinol with >99 % ee. (S)-1-Benzoyl-3-pyrrolidinol was successfully converted to 3-pyrrolidinol and its derivatives with by chemical reactions (>99 % ee).     

Occurrence of 15-cis-violaxanthin in Viola tricolor

A new cis isomer in the violaxanthin series has been isolated from the blossoms of Viola tricolor and identified by MS, IR and UV as the central-monocis form. It was converted to all-trans-violaxanthin by stereomutation. The CD correlation between 15-cis-violaxanthin and natural violaxanthin (5,6,5′,6′-diepoxy-5,6,5′,6′-tetrahydro- β,β-caroten-3,3′-diol) provided the basis for assignment of the absolute configurations 3S, 5R, 6S, 3′S, 5′R, 6′S. Trans—cis isomerization of all-trans-violaxanthin also resulted in 15- cis-violaxanthin. In addition a quantitative determination of the carotenoids was conducted.