Tetra-acylated cyanidin 3-sophoroside-5-glucosides from the flowers of Iberis umbellata L. (Cruciferae)

The structures of 11 acylated cyanidin 3-sophoroside-5-glucosides (pigments 1-11), isolated from the flowers of Iberis umbellata cultivars (Cruciferae), were elucidated by chemical and spectroscopic methods. Pigments 1-11 were acylated with malonic acid, p-coumaric acid, ferulic acid, sinapic acid and/or glucosylhydroxycinnamic acids.Pigments 1-11 were classified into four groups by the substitution patterns of the linear acylated residues at the 3-position of the cyanidin. In the first group, pigments 1-3 were determined to be cyanidin 3-O-[2-O-(2-O-(acyl)-β-glucopyranosyl)-6-O-(trans-p-coumaroyl)-β-glucopyranoside]-5-O-[6-O-(malonyl)-β-glucopyranoside], in which the acyl moiety varied with none for pigment 1, ferulic acid for pigment 2 and sinapic acid for pigment 3. In the second one, pigments 4-6 were cyanidin 3-O-[2-O-(2-O-(acyl)-β-glucopyranosyl)-6-O-(4-O-(β-glucopyranosyl)-trans-p-coumaroyl)-β-glucopyranoside]-5-O-[6-O-(malonyl)-β-glucopyranoside], in which the acyl moiety varied with none for pigment 4, ferulic acid for pigment 5 and sinapic acid for pigment 6. In the third one, pigments 7-9 were cyanidin 3-O-[2-O-(2-O-(acyl)-β-glucopyranosyl)-6-O-(4-O-(6-O-(trans-feruloyl)-β-glucopyranosyl)-trans-p-coumaroyl)-β-glucopyranoside]-5-O-[6-O-(malonyl)-β-glucopyranoside], in which the acyl moiety varied with none for pigment 7, ferulic acid for pigment 8, and sinapic acid for pigment 9. In the last one, pigments 10 and 11 were cyanidin 3-O-[2-O-(2-O-(acyl)-β-glucopyranosyl)-6-O-(4-O-(6-O-(4-O-(β-glucopyranosyl)-trans-feruloyl)-β-glucopyranosyl)-trans-p-coumaroyl)-β-glucopyranoside]-5-O-[6-O-(malonyl)-β-glucopyranoside], in which acyl moieties were none for pigment 10 and ferulic acid for pigment 11.The distribution of these pigments was examined in the flowers of four cultivars of I. umbellata by HPLC analysis. Pigment 1 acylated with one molecule of p-coumaric acid was dominantly observed in purple-violet cultivars. On the other hand, pigments (9 and 11) acylated with three molecules of hydroxycinnamic acids were observed in lilac (purple-violet) cultivars as major anthocyanins. The bluing effect and stability on these anthocyanin colors were discussed in relation to the molecular number of hydroxycinnamic acids in these anthocyanin molecules.     

Bilirubin conjugates of human bile. The excretion of bilirubin as the acyl glycosides of aldobiouronic acid,pseudoaldobiouronic acid and hexuronosylhexuronic acid,with a branched-chain hexuronic acid as one of the components of hexuronosylhexuronide

Structure elucidations have been performed on the bilirubin conjugates isolated from human hepatic bile as the phenylazo derivatives. The major bilirubin conjugates are excreted, not as was formerly thought in the form of glucuronides, but as the acyl glycosides of aldobiouronic acid, pseudoaldobiouronic acid and hexuronosylhexuronic acid. The isolated aldobiouronides are proposed to have the structures of an acyl 6-O-hexopyranosyluronic acid-hexopyranoside, an acyl 4-O-hexofuranosyluronic acid-d-glucopyranoside, and an acyl 4-O-β-d-glucofuranosyluronic acid-d-glucopyranoside respectively, with the acyl radicals being those of the phenylazo derivative of bilirubin. The pseudoaldobiouronide is suggested to be the acyl 4-O-α-d-glucofuranosyl-β-d -glucopyranosiduronic acid, with the acyl radical being that of the phenylazo derivative of vinylneoxanthobilirubinic acid. The hexuronosylhexuronide presumably is the acyl 4-O-(3-C-hydroxymethylribofuranosyluronic acid)-β-d-glucopyranosiduronic acid, with the acyl radical being that of the phenylazo derivative of bilirubin. The 3-C-hydroxymethylriburonic acid, isolated as one of the components of the hexuronosylhexuronide, is the first natural branched-chain hexuronic acid to be detected, and the first branched-chain sugar ever detected in humans.     

Mononuclear and tetranuclear palladacycles with terdentate [C,N,N] and [C,N,O] Schiff base ligands. C-H versus C-Br activation reactions

Reaction of the Schiff base ligands 2-Br-4,5-(OCH₂O)C₆H₂C(H)NCH₂CH₂NMe₂ (a) and 4,5-(OCH₂CH₂)C₆H₃C(H)NCH₂CH₂NMe₂ (b) with Pd(OAc)₂ or K₂[PdCl₄] leads to the mononuclear cyclometallated compounds [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)NCH₂CH₂NMe₂-C6,N,N}(OCOMe)] (1a) and [Pd{4,5-(OCH₂CH₂)C₆H₂C(H)NCH₂CH₂NMe₂-C6,N,N}(Cl)] (1b), derived from C-H activation at the C6 carbon. Treatment of a with Pd₂(dba)₃ gave [Pd{4-5-(OCH₂O)C₆H₂C(H)NCH₂CH₂NMe₂-C2,N,N}(Br)] (2a), via C-Br activation.The metathesis reaction of 1a with aqueous sodium chloride gave [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)NCH₂CH₂NMe₂-C6,N,N}(Cl)] (3a), with exchange of the acetate group by a chloride ligand. Treatment of the cyclometallated monomers 1a-3a with PPh₃ in a 1:1 molar ratio yielded the mononuclear complexes [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)NCH₂CH₂NMe₂-C6,N}(L)(PPh₃)] (L: OAc, 4a; Cl, 5a) and [Pd{4-5-(OCH₂O)C₆H₂C(H)NCH₂CH₂NMe₂-C2,N}(Br)(PPh₃)] (6a), with Pd-NMe₂ bond cleavage. However, treatment of a solution of 3a or 2a with silver trifluoromethanesulfonate, followed by reaction with PPh₃ in acetone yielded the cyclometallated complexes [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)NCH₂CH₂NMe₂-C6,N,N}(PPh₃)][CF₃SO₃] (7a) and [Pd{4-5-(OCH₂O)C₆H₂C(H)NCH₂CH₂NMe₂-C2,N,N}(PPh₃)][CF₃SO₃] (8a), respectively, where the Pd-NMe₂ bond was retained.The reaction of the ligands 2-Br-4,5-(OCH₂O)C₆H₂C(H)N(2′-OH-5′-^tBuC₆H₃) (c) and 4,5-(OCH₂CH₂)C₆H₃C(H)N(2′-OH-5′-^tBuC₆H₃) (d) with Pd(OAc)₂ gave the tetranuclear complexes [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)N(2′-O-5′-^tBuC₆H₃)-C6,N,O}]₄ (1c) and [Pd{4,5-(OCH₂CH₂)C₆H₂C(H)N(2′-O-5′-^tBuC₆H₃)-C6,N,O}]₄ (1d), respectively. Treatment of 1c with PPh₃ in 1:4 molar ratio, gave the mononuclear species [Pd{2-Br-4,5-(OCH₂O)C₆HC(H)N(2′-(O)-5′-^tBuC₆H₃)-C6,N,O}(PPh₃)] (2c) with opening of the polynuclear structure after P-O_bridging bond cleavage.The structure of compounds 2a, 1c and 1d has been determined by X-ray diffraction analysis.     

Isomerization of 1-O-Indol-3-Ylacetyl-β-d-Glucose : Enzymatic Hydrolysis of 1-O, 4-O, and 6-O-Indol-3-YlacetyL-β-d-Glucose and the Enzymatic Synthesis of Indole-3-Acetyl Glycerol by a Hormone Metabolizing Complex

The first compound in the series of reactions leading to the ester conjugates of indole-3-acetic acid (IAA) in kernels of Zea mays sweet corn is the acyl alkyl acetal, 1-O-indol-3-ylacetyl-β-d-glucose (1-O-IAGlu). The enzyme catalyzing the synthesis of this compound is UDP-glucose:indol-3-ylacetate glucosyl-transferase (IAGlu synthase). The IAA moiety of the high energy compound 1-O-IAGlu may be enzymatically transferred to myo-inositol or to glycerol or the 1-O-IAGlu may be enzymatically hydrolyzed. Alternatively, nonenzymatic acyl migration may occur to yield the 2-O, 4-O, and 6-O esters of IAA and glucose. The 4-O and 6-O esters may then be enzymatically hydrolyzed to yield free IAA and glucose. This work reports new enzymatic activities, the transfer of IAA from 1-O-IAGlu to glycerol, and the enzymecatalyzed hydrolysis of 4-O- and 6-O-IAGlu. Data is also presented on the rate of non-enzymatic acyl migration of IAA from the 1-O to the 4-O and 6-O positions of glucose. We also report that enzymes catalyzing the synthesis of 1-O-IAGlu and the hydrolysis of 1-O, 4-O, and 6-O-IAGlu fractionate as a hormone metabolizing complex. The association of synthetic and hydrolytic capabilities in enzymes which cofractionate may have physiological significance.     

Enzymatic synthesis of lactosylated and sialylated derivatives of epothilone A

Epothilone A is a derivative of 16-membered polyketide natural product, which has comparable chemotherapeutic effect like taxol. Introduction of sialic acids to these chemotherapeutic agents could generate interesting therapeutic glycoconjugates with significant effects in clinical studies. Since, most of the organisms biosynthesize sialic acids in their cell surface, they are key mediators in cellular events (cell-cell recognition, cell-matrix interactions). Interaction between such therapeutic sugar parts and cellular polysaccharides could generate interesting result in drugs like epothilone A. Based on this hypothesis, epothilone A glucoside (epothilone A 6-O-β-D-glucoside) was further decorated by conjugating enzymatically galactose followed by sialic acids to generate epothilone A 7-O-β-D-glucopyranosyl, 4′-O-α-D-galactoside i.e., lactosyl epothilone A (lac epoA) and two sialosides of epothilone A namely epothilone A 7-O-β-D-glucopyranosyl, 4′-O-α-D-galactopyranosyl 3″-O-α-N-acetyl neuraminic acid and epothilone A 7-O-β-D-glucopyranosyl, 4′-O-α-D-galactopyranosyl 6″-O-α-N-acetylneuraminic acid i.e., 3′sialyllactosyl epothilone A: 3′SL-epoA, and 6′sialyllactosyl epothilone A: 6′SL-epoA, respectively. These synthesized analogs were spectroscopically analyzed and elucidated, and biologically validated using HUVEC and HCT116 cancer cell lines.     

Acetylated allose-containing flavonoid glucosides from Stachys anisochila

In continuation of our chemosystematic study of Stachys (Labiatae) we have isolated the previously reported isoscutellarein 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-β-D-glucopyranoside] (1) and 3′-hydroxy-4′-O-methylisoscutellarein 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-β-D-glucopyranoside] (4) and four new allose-containing flavonoid glycosides from S. anisochila. The new glycosides are hypolaetin 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-β-D-glucopyranside] (6) as well as the three corresponding diacetyl analogues of 1, 4 and 6, isoscutellarein 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-6″-O-acetyl-β-D-glucopyranoside], 3′-hydroxy-4′-O-methylisoscutellarein 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-6″-O-acetyl-β-D-glucopyranoside] and hypolaetin 7-O-[6″'-O-acetyl-β-D-allopyranosyl-(1 → 2)-6″-O-acetyl-β-D-glucopyranoside]. Extensive two-dimensional NMR studies (proton-carbon correlations, COSY experiments) allowed assignment of all ¹H NMR sugar signals and a correction of the ¹³C NMR signal assignments for C-2 and C-3 of the allose.     

Novel stable five-coordinate diorgano cobalt(III) complexes: Formation, structure, and reaction with carbon monoxide

A series of new five-coordinate acyl vinyl cobalt(III) complexes Co{η¹-C(CCPh)CHPh}[C(O)CCO] L₂(L = PMe₃) (6-10) were prepared via formal insertion of diphenylbutadiyne into Co-H function of mer-octahedral hydrido-acyl(phenolato)-cobalt(III) complexes. The complexes are diamagnetic. One square pyramidal structure of complex 6 was confirmed by X-ray diffraction analysis. These complexes are stable in solid state. In solution, six-coordinate acyl vinyl carbonyl cobalt(III) complex 11 is approved through the reaction of complex 7 with CO and the structure of complex 11 was determined by X-ray method.     

Structure and Hypotensive Effect of Flavonoid Glycosides in Kinkan (Fortunella japonica) Peelings

An attempt was made to isolate the hypotensive substances from a hot water extract of kinkan. Eight flavonoid glycosides were isolated by repeated chromatography and by gel filtration after extracting with n-butanol and treating with lead subacetate. Their structures were established to be 6,8-di-C-glucosylapigenin (1), 3,6-di-C-glucosylacacetin (2), 2″-O-α-l-rhamnosyl-4′-O-methyl-vitexin (3), 2″-O-α-l-rhamnosyl-4′-O-methylisovitexin (4), 2″-O-α-l-rhamnosylvitexin (5), 2″-O-α-l-rhamnosylorientin (6), 2″-O-α-l-rhamnosyl-4′-O-methylorientin (7) and ponicilin (8) by UV. MS, ¹-NMR and ¹³C-NMR spectroscopy, and by sugar analysis. Each component was intravenously injected in SHR-SP (0.5 ~ 1.0 mg/100 g of body weight), 1, 2, 5 and 6 were found to lower the rat blood pressure.

Among these compounds, 2, 3, 4, 6 and 7 were new flavone glycosides.     

Synthesis and Biological Activity of the Lipophilic Derivatives of N- Acetyl-1-thiomuramoyl-l-alanyl-d-isoglutamine

A variety of the lipophilic derivatives at C-1 and C-6 in N-[2-O-(2-acetamido-2,3-dideoxy-1-thio-β-d-glucopyranose-B-yl)-d-lactoy]-l-alanyl-(N¹-fatty acyl)-d-isoglutamine methyl esters were synthesized from 2N-acetyl-1-S-acetyl-4,6-O-isopropylidene-1-thiomuramoyl-l-alanyl-d-isogluta-mine methyl ester. Their immunoadjuvant activity in guinea-pigs, and the protective effect in mice infected with Escherichia coli (E-77156) were examined.     

p-Hydroxybenzoyl-Glucose Is a Zwitter Donor for the Biosynthesis of 7-Polyacylated Anthocyanin in Delphinium

The blue color of delphinium (Delphinium grandiflorum) flowers is produced by two 7-polyacylated anthocyanins, violdelphin and cyanodelphin. Violdelphin is derived from the chromophore delphinidin that has been modified at the 7-position by Glc and p-hydroxybenzoic acid (pHBA) molecules. Modification of violdelphin by linear conjugation of Glc and pHBA molecules to a Glc moiety at the 7-position produces cyanodelphin. We recently showed that anthocyanin 7-O-glucosylation in delphinium is catalyzed by the acyl-Glc–dependent anthocyanin glucosyltransferase (AAGT). Here, we sought to answer the question of which enzyme activities are necessary for catalyzing the transfer of Glc and pHBA moieties to 7-glucosylated anthocyanin. We found that these transfers were catalyzed by enzymes that use p-hydroxybenzoyl-Glc (pHBG) as a bifunctional acyl and glucosyl donor. In addition, we determined that violdelphin is synthesized via step-by-step enzymatic reactions catalyzed by two enzymes that use pHBG as an acyl or glucosyl donor. We also isolated a cDNA encoding a protein that has the potential for p-hydroxybenzoylation activity and two AAGT cDNAs that encode a protein capable of adding Glc to delphinidin 3-O-rutinoside-7-O-(6-O-[p-hydroxybenzoyl]-glucoside) to form violdelphin.     

Switching between penta- and hexacoordination with salen-silicon-complexes

The reaction between phenyltrichlorosilane and the tetradentate ligands o-HO-C₆H₄-C(CH₃)N-(CH₂)_n-NC(CH₃)-o-C₆H₄-OH (n = 2, 3, 4), supported by an amine base, yields pentacoordinate silicon complexes (C₆H₅)Si-[o-O-C₆H₄-C(CH₃)N-(CH₂)_n-N-C(CH₂)-o-C₆H₄-O] with enamine functionalized ligands. This reaction pattern can be transferred onto various ligands of 2-iminomethylphenolate-type. The resulting pentacoordinate silicon complexes react with a variety of Brønsted acids HY to yield hexacoordinate salen silicon complexes (C₆H₅)(Y)Si-[o-O-C₆H₄-C(CH₃)N-(CH₂)_n-NC(CH₃)-o-C₆H₄-O] (Y = benzoate, picrate, 8-oxyquinolinate, 2-oxy-1,4-naphthoquinonate, p-tert-butylphenolate, (5-phenyltetrazol)-2-ide, fluoride, tetrafluoroborate). Hexacoordination of their Si-atoms was confirmed by ²⁹Si NMR spectroscopy and, in some cases, by X-ray crystal structure analysis. Examples for similarities and differences in the coordination behavior of the silicon atom and its heavier congeners (Ge, Sn) in the salen-type coordination sphere as well as data regarding the nucleophilicity of some of these novel enamine complexes are presented.     

Synthesis and solid-state structures of (triphos)iridacyclopentadiene complexes as models for vinylidene intermediates in the [2 + 2 + 1] cyclotrimerization of alkynes

The iridium 1,1,1-tris(diphenylphosphinomethyl)ethane (triphos) complexes [{κ²(C¹,C⁴)-CRCRCRCR}{CH₃C(CH₂PPh₂)₃}Ir(NCMe)]BF₄ (2-NCMe, R = CO₂Me) and [{κ²(C¹,C⁴)-CRCRCRCR}{CH₃C(CH₂PPh₂)₃}Ir(CO)]BF₄ (2-CO, R = CO₂Me) serve as models for proposed iridium-vinylidene intermediates of relevance to the [2 + 2 + 1] cyclotrimerization of alkynes. The solid-state structures of 2-NCMe, 2-CO, and [κ²(C¹,C⁴)-CRCRCRCR]{CH₃C(CH₂PPh₂)₃}Ir(Cl) (2-Cl), were determined by X-ray crystallography.     

Modification of aminoallenylidene complexes of chromium via exchange of the C3-bound amino group

The aminoallenylidene(pentacarbonyl)chromium complexes [(CO)₅CrCCC(NR¹R²)Ph] (1a-c) react with dimethylamine by addition of the amine to the C1C2 bond of the allenylidene ligand to give alkenyl(amino)carbene complexes [(CO)₅CrC(NMe₂)CHC(NR¹R²)Ph] (2a-c) (R¹ = Me: R² = Me (a), Ph (b); R¹ = Et: R² = Ph (c)). In contrast, addition of a large excess (usually 20 equivalents) of ammonia or primary amines, H₂NR, to solutions of [(CO)₅CrCCC(NMe₂)Ph] (1a) affords the aminoallenylidene complexes [(CO)₅CrCCC(NHR)Ph] (1d-w) in which the dimethylamino group is replaced by NH₂ or NHR, respectively. In addition to simple amines such as methylamine, butylamine, and aniline, amines carrying a functional group (allylamine, propargylamine) and amino acid esters as well as amino terpenes and amino sugars can be used to displace the NMe₂ substituent. Usually the Z isomer (with respect to the partial C3-N double bond) is formed exclusively. Products derived from addition of H₂NR to the C1C2 bond of 1a are not observed. The amino group in 1d-w is rapidly deprotonated by excess of amine to form iminium alkynyl chromates [1d-w]⁻, thus protecting 1d-w from addition of free amine to either C3 or across the C1C2 bond. The iminium alkynyl chromates are readily reprotonated by acids or by chromatography on wet SiO₂ to reform 1d-w.     

Novel rhodium complexes containing a bulky iminophosphine ligand and their use as catalysts for the hydroboration of vinylarenes

Addition of o-Ph₂PC₆H₄CHN-2,6-ⁱPr₂C₆H₃ (1) to [RhCl(coe)₂]₂ (coe = cis-cycloctene) gave several new iminophosphino rhodium(I) complexes including [Rh(κ²-o-Ph₂PC₆H₄CHN-2,6-ⁱPr₂C₆H₃)(μ-Cl)]₂ (2). Addition of 1 to Rh(acac)(coe)₂ (acac = acetylacetonato) gave [Rh(acac)(κ²-o-Ph₂PC₆H₄CHN-2,6-ⁱPr₂C₆H₃)] (3) in yields of up to 75%. Complex 3 has been examined for its ability to catalyze the hydroboration of a series of vinyl arenes. Reactions using catecholborane and pinacolborane seem to proceed largely through a dehydrogenative borylation mechanism to give a number of boronated products.     

Synthesis, structure, and characterization of some ruthenium arene complexes of N-(arylmethylene)-2-(methylthio)anilines and 2-(methylthio)aniline

A series of cationic, half-sandwich ruthenium complexes with the general formula [(η⁶-p-cymene)RuCl(MeSC₆H₄2-NCHAr)][PF₆] (3a-h), have been prepared from the reaction of [(η⁶-p-cymene)RuCl₂]₂ with various N,S-donor Schiff base ligands derived from 2-(methylthio)aniline and several substituted benzaldehydes. The related aniline complex [(η⁶-p-cymene)RuCl(MeS-C₆H₄-2-NH₂)][PF₆] (4) was synthesized from 2-(methylthio)aniline. All of the ruthenium complexes were characterized by IR, ¹H NMR, and UV/Vis spectroscopies. The molecular structure of complex 4 was determined by X-ray crystallography.     

Crystal Structures of β-Primeverosidase in Complex with Disaccharide Amidine Inhibitors

β-Primeverosidase (PD) is a disaccharide-specific β-glycosidase in tea leaves. This enzyme is involved in aroma formation during the manufacturing process of oolong tea and black tea. PD hydrolyzes β-primeveroside (6-O-β-d-xylopyranosyl-β-d-glucopyranoside) at the β-glycosidic bond of primeverose to aglycone, and releases aromatic alcoholic volatiles of aglycones. PD only accepts primeverose as the glycone substrate, but broadly accepts various aglycones, including 2-phenylethanol, benzyl alcohol, linalool, and geraniol. We determined the crystal structure of PD complexes using highly specific disaccharide amidine inhibitors, N-β-primeverosylamidines, and revealed the architecture of the active site responsible for substrate specificity. We identified three subsites in the active site: subsite −2 specific for 6-O-β-d-xylopyranosyl, subsite −1 well conserved among β-glucosidases and specific for β-d-glucopyranosyl, and wide subsite +1 for hydrophobic aglycone. Glu-470, Ser-473, and Gln-477 act as the specific hydrogen bond donors for 6-O-β-d-xylopyranosyl in subsite −2. On the other hand, subsite +1 was a large hydrophobic cavity that accommodates various aromatic aglycones. Compared with aglycone-specific β-glucosidases of the glycoside hydrolase family 1, PD lacks the Trp crucial for aglycone recognition, and the resultant large cavity accepts aglycone and 6-O-β-d-xylopyranosyl together. PD recognizes the β-primeverosides in subsites −1 and −2 by hydrogen bonds, whereas the large subsite +1 loosely accommodates various aglycones. The glycone-specific activity of PD for broad aglycone substrates results in selective and multiple release of temporally stored alcoholic volatile aglycones of β-primeveroside.     

Oxidovanadium(IV) complexes containing ligands derived from dithiocarbazates - Models for the interaction of VO with thiofunctional ligands

The tetragonal-pyramidal VO²⁺ complexes [VO{(RSC-S)N-NX}₂] (1-6) were synthesised by the reactions of VO(OCHMe₂)₃ with the dithiocarbazate ligands RSC(S)-NH-NX, where X = cyclo-pentyl, cyclo-hexyl or 4-Me₂N-C₆H₄-CH, and R = CH₃ or CH₂C₆H₅. The compounds were characterised by elemental analysis, IR- and mass spectrometries, and in cases of compounds 1, 3, 4 and 5, by X-ray diffraction. The chiral compound 4 (X = cyclo-hexyl, R = CH₂C₆H₅) crystallises in the C configuration. In compound 5, the VO moiety is disordered (83.3:16.7%) with respect to the plane spanned by the four equatorial ligand functions.     

Regioselective acylation of several polyhydroxylated natural compounds by Candida antarctica lipase B

Regioselective acylation of four polyhydroxylated natural compounds, deacetyl asperulosidic acid (1), asperulosidic acid (2), puerarin (3) and resveratrol (4) by Candida antarctica Lipase B in the presence of various acyl donors (vinyl acetate, vinyl decanoate or vinyl cinnamoate) was studied. Compounds 1, 2 and 4 were regioselectively acetylated with vinyl acetate to afford products, 3′-O-acetyl-10-O-deacetylasperulosidic acid (1a), 3′,6′-O-diacetyl-10-O-deacetylasperulosidic acid (1b), 3′-O-acetylasperulosidic acid (2a), 3′,6′-O-diacetylasperulosidic acid (2b), 4′-O-acetylresveratrol (4a), respectively, with yields of 22 to 50%, while reactions with vinyl decanoate and vinyl cinnamoate were slow with lower yields. Compound 3 was readily acylated with all three acyl donors and quantitatively converted to products 6″-O-acetylpuerarin (3a), 6″-O-decanoylpuerarin (3b), 6″-O-cinnamoylpuerarin (3c), respectively. The structures of these acylated products were determined by spectroscopic methods (MS and NMR).     

Purification and Characterization of a Novel Galloyltransferase Involved in Catechin Galloylation in the Tea Plant (Camellia sinensis)

Catechins (flavan-3-ols), the most important secondary metabolites in the tea plant, have positive effects on human health and are crucial in defense against pathogens of the tea plant. The aim of this study was to elucidate the biosynthetic pathway of galloylated catechins in the tea plant. The results suggested that galloylated catechins were biosynthesized via 1-O-glucose ester-dependent two-step reactions by acyltransferases, which involved two enzymes, UDP-glucose:galloyl-1-O-β-d-glucosyltransferase (UGGT) and a newly discovered enzyme, epicatechin:1-O-galloyl-β-d-glucose O-galloyltransferase (ECGT). In the first reaction, the galloylated acyl donor β-glucogallin was biosynthesized by UGGT from gallic acid and uridine diphosphate glucose. In the second reaction, galloylated catechins were produced by ECGT catalysis from β-glucogallin and 2,3-cis-flavan-3-ol. 2,3-cis-Flavan-3-ol and 1-O-galloyl-β-d-glucose were appropriate substrates of ECGT rather than 2,3-trans-flavan-3-ol and 1,2,3,4,6-pentagalloylglucose. Purification by more than 1641-fold to apparent homogeneity yielded ECGT with an estimated molecular mass of 241 to 121 kDa by gel filtration. Enzyme activity and SDS-PAGE analysis indicated that the native ECGT might be a dimer, trimer, or tetramer of 60- and/or 58-kDa monomers, and these monomers represent a heterodimer consisting of pairs of 36- or 34- of and 28-kDa subunits. MALDI-TOF-TOF MS showed that the protein SCPL1199 was identified. Epigallocatechin and epicatechin exhibited higher substrate affinities than β-glucogallin. ECGT had an optimum temperature of 30 °C and maximal reaction rates between pH 4.0 and 6.0. The enzyme reaction was inhibited dramatically by phenylmethylsulfonyl fluoride, HgCl₂, and sodium deoxycholate.