Isolation and characterization of rat intestinal bacteria involved in biotransformation of (−)-epigallocatechin

Two intestinal bacterial strains MT4s-5 and MT42 involved in the degradation of (?)-epigallocatechin (EGC) were isolated from rat feces. Strain MT4s-5 was tentatively identified as Adlercreutzia equolifaciens. This strain converted EGC into not only 1-(3, 4, 5-trihydroxyphenyl)-3-(2, 4, 6-trihydroxyphenyl)propan-2-ol (1), but also 1-(3, 5-dihydroxyphenyl)-3-(2, 4, 6-trihydroxyphenyl)propan-2-ol (2), and 4′-dehydroxylated EGC (7). Type strain (JCM 9979) of Eggerthella lenta was also found to convert EGC into 1. Strain MT42 was identified as Flavonifractor plautii and converted 1 into 4-hydroxy-5-(3, 4, 5-trihydroxyphenyl)valeric acid (3) and 5-(3, 4, 5-trihydroxyphenyl)-γ-valerolactone (4) simultaneously. Strain MT42 also converted 2 into 4-hydroxy-5-(3, 5-dihydroxyphenyl)valeric acid (5), and 5-(3, 5-dihydroxyphenyl)-γ-valerolactone (6). Furthermore, F. plautii strains ATCC 29863 and ATCC 49531 were found to catalyze the same reactions as strain MT42. Interestingly, formation of 2 from EGC by strain MT4s-5 occurred rapidly in the presence of hydrogen supplied by syntrophic bacteria. Strain JCM 9979 also formed 2 in the presence of the hydrogen or formate. Strain MT4s-5 converted 1, 3, and 4 to 2, 5, and 6, respectively, and the conversion was stimulated by hydrogen, whereas strain JCM 9979 could catalyze the conversion only in the presence of hydrogen or formate. On the basis of the above results together with previous reports, the principal metabolic pathway of EGC and EGCg by catechin-degrading bacteria in gut tract is proposed.     

Enzymatic activity of cell-free extracts from Burkholderia oxyphila OX-01 bio-converts (+)-catechin and (−)-epicatechin to (+)-taxifolin

This study characterized the enzymatic ability of a cell-free extract from an acidophilic (+)-catechin degrader Burkholderia oxyphila (OX-01). The crude OX-01 extracts were able to transform (+)-catechin and (?)-epicatechin into (+)-taxifolin via a leucocyanidin intermediate in a two-step oxidation. Enzymatic oxidation at the C-4 position was carried out anaerobically using H₂O as an oxygen donor. The C-4 oxidation occurred only in the presence of the 2R-catechin stereoisomer, with the C-3 stereoisomer not affecting the reaction. These results suggest that the OX-01 may have evolved to target both (+)-catechin and (?)-epicatechin, which are major structural units in plants.     

Conversion of (−)-Carvone and (+)-Carvone by a Strain of Aspergillus niger

The conversion of (?)-carvone and (+)-carvone by a strain of Aspergillus niger was studied as one of the series of biochemical reduction of terpenes.

(?)-Carvone was found to be reduced essentially to (+)-neodihydrocarveol, although (+)-dihydrocarvone and (+)-isodihydrocarvone were also formed in small amounts, whereas (+)-carvone was converted to (?)-isodihydrocarvone, (?)-isodihydrocarveol, (?)-neoisodihydrocarveol, (?)-dihydrocarvone, (?)-neodihydrocarveol, and (+)-dihydrocarveol, of which the former three were the major products.

The metabolic pathways for (?)-carvone and (+)-carvone by the strain of Aspergillus niger are discussed and the results on microbial and chemical reductions of carvone and dihydrocarvone are summarized.     

Biotransformation of (+)-carvone and (−)-carvone using human skin fungi: A green method of obtaining fragrances and flavours

The synthesis of optically pure compounds is increasingly in demand among the pharmaceutical, fine chemical and agro-food industries, while the importance of chirality in the activity and biological properties of many compounds has previously been established. The aim of the present study was therefore to evaluate the biotransformation capacities of (+)-carvone and (?)-carvone using the fungi Scolecobasidium sp, three lines of Cladosporium sp, Phoma sp, Aureobasidium sp and Epicoccum sp, all obtained from human skin. The seven fungi evaluated were capable of hydrogenating the activated alkene, followed by the reduction of ketone to chiral alcohol, with conversions between 9.5 and 100%, and with diastereomer excess (d.e.) of over 89% of dihydrocarveol when (+)-carvone was used as a substrate. These results demonstrate that the filamentous fungi of human skin are potential biocatalytic tools for obtaining chiral alcohols.     

Conversion of (−)-Carvotanacetone and (+)-Carvotanacetone by Pseudomonas ovalis,Strain 6–1

As a part of series on the biochemical reduction of terpenes, the conversion of (?)-carvotanacetone (I) and (+)-carvotanacetone (II) by Pseudomonas ovalis, strain 6–1, has been studied.

By the action of the microorganism, I was reduced to give (+)-carvomenthone (III), (+)-neocarvomenthol (IV), and (?)-carvomenthol (V), whereas II was also reduced to (?)-isocarvomenthone (VI), (?)-carvomenthone (VII), (?)-isocarvomenthol (VIII), and (?)-neoisocarvomenthol (IX); of which III, VI and IX are the major products.

The metabolic pathways of I and II and mechanism of stereospecific hydrogenation are also discussed.     

Myeloperoxidase-mediated LDL oxidation and endothelial cell toxicity of oxidized LDL: attenuation by (−)-epicatechin

Recent data suggest an inverse epidemiological association between intake of flavanol-rich cocoa products and cardiac mortality. Potential beneficial effect of cocoa may be attributed to flavanol-mediated improvement of endothelial function, as well as to enhancement of bioavailability and bioactivity of nitric oxide in vivo. ( ? )-Epicatechin is one bioactive flavanol found in cocoa. This review deals with protective actions of ( ? )-epicatechin on two key processes in atherogenesis, oxidation of LDL and damage to endothelial cell by oxidized LDL (oxLDL), with emphasis on data from this laboratory. ( ? )-Epicatechin not only abrogates or attenuates LDL oxidation but also counteracts deleterious actions of oxLDL on vascular endothelial cells. These protective actions are only partially shared by other vasoprotective agents such as vitamins C and E or aspirin. Thus, ( ? )-epicatechin appears to be a pleiotropic protectant for both LDL and endothelial cells.     

Biological activities of (−)-epicatechin and (−)-epicatechin-containing foods: Focus on cardiovascular and neuropsychological health

Recent studies have suggested that certain (?)-epicatechin-containing foods have a blood pressure-lowering capacity. The mechanisms underlying (?)-epicatechin action may help prevent oxidative damage and endothelial dysfunction, which have both been associated with hypertension and certain brain disorders. Moreover, (?)-epicatechin has been shown to modify metabolic profile, blood's rheological properties, and to cross the blood-brain barrier. Thus, (?)-epicatechin causes multiple actions that may provide unique synergy beneficial for cardiovascular and neuropsychological health. This review summarises the current knowledge on the biological actions of (?)-epicatechin, related to cardiovascular and brain functions, which may play a remarkable role in human health and longevity.     

Functional beta cell regeneration in the islets of pancreas in alloxan induced diabetic rats by (−)-epicatechin

(?)-Epicatechin (1), a naturally occuring flavonoid compound was found to have reversed the diabetogenic action of alloxan in albino rats (2). (?)-Epicatechin administration in doses of 30 mg/kg (i.p) twice daily for 4–5 days in alloxan induced (150 mg/kg, i.p.) diabetic albino rats (either sex), has brought down the high blood sugar levels to normal values. Concurrent histological studies of the pancreas of these animals showed regeneration of the beta-cell population of the islets which were earlier necrosed by alloxan. Immunoreactive insulin (IRI) studies showed that the regenerated beta-cells of the islets of pancreas are functional in nature.     

Effects of (−)-epicatechin on molecular modulators of skeletal muscle growth and differentiation

Sarcopenia is a notable and debilitating age-associated condition. Flavonoids are known for their healthy effects and limited toxicity. The flavanol (−)-epicatechin (Epi) enhances exercise capacity in mice, and Epi-rich cocoa improves skeletal muscle structure in heart failure patients. (−)-Epicatechin may thus hold promise as treatment for sarcopenia.We examined changes in protein levels of molecular modulators of growth and differentiation in young vs. old, human and mouse skeletal muscle. We report the effects of Epi in mice and the results of an initial proof-of-concept trial in humans, where muscle strength and levels of modulators of muscle growth were measured. In mice, myostatin and senescence-associated β-galactosidase levels increase with aging, while those of follistatin and Myf5 decrease. (−)-Epicatechin decreases myostatin and β-galactosidase and increases levels of markers of muscle growth. In humans, myostatin and β-galactosidase increase with aging while follistatin, MyoD and myogenin decrease. Treatment for 7 days with (−)-epicatechin increases hand grip strength and the ratio of plasma follistatin/myostatin.In conclusion, aging has deleterious effects on modulators of muscle growth/differentiation, and the consumption of modest amounts of the flavanol (−)-epicatechin can partially reverse these changes. This flavanol warrants its comprehensive evaluation for the treatment of sarcopenia.     

Immunohistochemical localization of β-glucosidases in lignin and isoflavone metabolism in Cicer arietinum L. seedlings

Coniferin specific- and isoflavone 7-glucoside specific -glucosidases have been localized in stem and root sections of chick pea (Cicer arietinum L.) seedlings by the indirect immunofluorometrical method. The coniferin specific -glucosidase has been found in the cell walls of the tracheary elements and of the endo-, epi-, and exodermis. All these tissues are known to contain either lignin or polymers, like suberin and cutin, which consist partially of phenylpropanoid elements. The localization of this -glucosidase is therefore in agreement with its postulated relationship to the phenylpropanoid metabolism. The isoflavone 7-glucoside specific -glucosidase, on the other hand, is predominantly located in the parenchymatic cortex cells, and obviously in the cytoplasm. These cells are known to contain the isoflavone formononetin, which has been shown to undergo turnover in chick pea seedlings. We therefore have good reason to assume that this -glucosidase is involved in the metabolism of the 7-glucoside of this isoflavone.Abbreviations SDS sodium dodecylsulfate - PBS physiological phosphate saline The results are part of the thesis of Gerd Burmeister, 1980, University of Münster     

The lack of effectiveness of (−)-epicatechin against alloxan induced diabetes in Wistar rats

(?)-Epicatechin, a naturally occurring flavonoid, has been reported to protect pancreatic beta cells from alloxan-induced diabetes (1) and to stimulate beta cell regeneration when given after alloxan administration (2,3). However, in the present study, we have not been able to confirm these findings. Administration of (?)-epicatechin (100 mg/kg/day for 15 days) beginning three days after alloxan administration (140 mg/kg) had no significant effect on blood glucose levels when compared to alloxan control animals. Another study which failed to demonstrate an antidiabetic effect of (?)-epicatechin (4) has been criticized for not using fresh solutions of (?)-epicatechin because of its instability in aqueous solution (5). We have found, however, that (?)-epicatechin is stable for at least five days based on thin-layer chromatographic analysis and optical rotation measurements.     

Biotransformation of (−)-codeinone to (−)-codeine byPapaver somniferum cells immobilized in reticulate polyurethane foam

Papaver somniferum cells immobilized in reticulate-polyurethane foam biotransformed (–)-codeinone to (–)-codeine. A biotransformation ratio of 79% was found in immobilized cells whereas a ratio of 57% was found in suspended cells. Of the total amount of codeine formed only 12.2% was detected inside the cells, most of the product (87.3%) being released into the medium. When immobilized cells were cultivated in the absence of nitrate, only 40% of the cells remained alive and the biotransformation of codeinone was strongly reduced. When orthophosphate was omitted from the growth medium a bioconversion ratio of 86% was achieved.     

Effects of (−)-epicatechin and derivatives on nitric oxide mediated induction of mitochondrial proteins

Impaired mitochondrial function represents an early manifestation of endothelial dysfunction and likely contributes to the development of cardiovascular diseases (CVD). The stimulation of mitochondrial function and/or biogenesis is seen as a means to improve the bioenergetic and metabolic status of cells and thus, reduce CVD. In this study we examined the capacity of the flavanol (?)-epicatechin and two novel derivatives to enhance mitochondrial function and protein levels in cultured bovine coronary artery endothelial cells. As nitric oxide production by endothelial cells is suspected in mediating mitochondria effects (including biogenesis), we also examined the dependence of responses on this molecule using an inhibitor of nitric oxide synthase. Results indicate that the flavanol (?)-epicatechin and derivatives are capable of stimulating mitochondrial function as assessed by citrate synthase activity as well as induction of structural (porin, mitofilin) and oxidative phosporylation protein levels (complex I and II). Effects were blocked by the use of the chemical inhibitor of the synthase thus, evidencing a role for nitric oxide in mediating these effects. The results observed indicate that the three agents are effective in enhancing mitochondria function and protein content. The effects noted for (?)-epicatechin may serve to explain the healthy effects on cardiometabolic risk ascribed to the consumption of cocoa products.     

(+)-Catechin is more bioavailable than (−)-catechin: Relevance to the bioavailability of catechin from cocoa

Catechin is a flavonoid present in fruits, wine and cocoa products. Most foods contain the (+)-enantiomer of catechin but chocolate mainly contains ( ? )-catechin, in addition to its major flavanol, ( ? )-epicatechin. Previous studies have shown poor bioavailability of catechin when consumed in chocolate. We compared the absorption of ( ? ) and (+)-catechin after in situ perfusion of 10, 30 or 50 μmol/l of each catechin enantiomer in the jejunum and ileum in the rat. We also assayed 23 samples of chocolate for (+) and ( ? )-catechin. Samples were analyzed using HPLC with a Cyclobond I-2000 RSP chiral column. At all concentrations studied, the intestinal absorption of ( ? )-catechin was lower than the intestinal absorption of (+)-catechin (p < 0.01). Plasma concentrations of ( ? )-catechin were significantly reduced compared to (+)-catechin (p < 0.05). The mean concentration of ( ? )-catechin in chocolate was 218 ± 126 mg/kg compared to 25 ± 15 mg/kg (+)-catechin. Our findings provide an explanation for the poor bioavailability of catechin when consumed in chocolate or other cocoa containing products.     

Cyclohexene epoxides, (+)-pandoxide, (+)β-senepoxide and (−)-pipoxide,from Uvaria pandensis

The new cyclohexene epoxide (+)-pandoxide along with (+)-β-senepoxide and (−)-pipoxide were isolated from the stembark, roots and leaves of the new Uvaria species U. pandensis. Their structures and stereochemical configurations were determined by spectroscopic methods. The pipoxide has an absolute configuration opposite to the one previously isolated from Uvaria species. The absolute stereochemical configuration of (+)-β-senepoxide was assigned unambiguously by an X-ray diffraction analysis.     

A New Keto-alcohol, (−)-Mintlactone, (+)-isoMintlactone and Minor Components in Peppermint Oil

Eighty-one constituents were newly identified from the oil of Mentha piperita L., including a new keto-alcohol, (?)-mintlactone and (+)-isomintlactone. They were determined by spectral data and syntheses to be 4-hydroxy-4-methyl-2-cyclohexen-1-one (8), (6R, 7aR) (10) and (6R, 7aS)-3,6-dimethyl-5,6,7,7a-tetrahydro-2(4H)-benzofuranone (11), respectively.     

Enantiomer and conformer recognition of (+) and (−)-disparlure and their analogs by the pheromone binding proteins of the gypsy moth,Lymantria dispar

Adult female gypsy moths produce a sex pheromone (+)-(7R,8S)-2-methyl-7,8-epoxyoctadecane, (+)-disparlure, to attract male gypsy moths. To better understand the recognition of (+)-disparlure by the male’s olfactory system, we synthesized racemic and enantiopure oxa and thia analogs of (+)-disparlure (ee > 98%). Ab initio calculations of the conformeric landscapes around the dihedral angles C_{5–6–7–8} and C_{7–8–9–10} of (+)-disparlure and corresponding dihedral angles of analogs revealed that introduction of the heteroatom changes the conformeric landscape around these important epitopes. The energy difference between HOMO and LUMO decreased after oxygen or sulfur was introduced into the backbone. Consistent with this, an enhancement of binding affinity between sulfur analogs and the pheromone-binding proteins (PBPs) was observed in vitro. Docking of the pheromone and analogs onto models of the two known PBPs of the gypsy moth revealed that the internal binding pocket of PBP1 showed higher selectivity than that of PBP2, consistent with in vitro binding assays. Further energy analysis revealed that enantiomers adopted different conformations with different energies when docked in the internal binding pocket of PBPs, resulting in enantiomer discrimination of PBPs towards disparlure and its analogs.     

Microbial transformations of (−)-α- and (+)-β-thujone by fungi of Absidia species

The microbial transformations of (−)-α- and (+)-β-thujone (1a and 1b) in cultures of Absidia species: Absidia coerulea AM93, Absidia glauca AM254 and Absidia cylindrospora AM336 were studied. The biotransformations of (−)-α-thujone (1a), by these fungi strains, afforded mixtures of 4-hydroxy- and 7-hydroxy-α-thujone (2 and 3). Aforementioned fungi strains were also able to hydroxylate of (+)-β-thujone at C-7 position. Only A. glauca AM254 transformed 1b to 8-hydroxy-β-thujone (7) and (2S)-2-hydroxyneoisothujol (6). The (4R)-4-hydroxyisothujole (5) was identified as one of the major metabolite of (+)-β-thujone (1b) in culture of A. cylindrospora AM336. This strain was also able to introduce hydroxy group to C-4 position in 1b without reduction of carbonyl group at C-3. The absolute configuration of all chiral centers of new (4R)-4-hydroxyisothujol (5) and (2S)-2-hydroxyneoisothujol (6) were established taking into account the configuration of (+)-β-thujone (1b) and their spectral data.