Secoisolariciresinol dehydrogenase purification, cloning, and functional expression. Implications for human health protection. (1/17)

Matairesinol is a central precursor in planta in the biosynthesis of numerous lignans, including that of the important antiviral and anticancer agent, podophyllotoxin. In this study, the approximately 32-kDa NAD-dependent secoisolariciresinol dehydrogenase, which catalyzes the enantiospecific conversion of (-)-secoisolariciresinol into (-)-matairesinol in Forsythia intermedia, was purified >6,000-fold to apparent homogeneity. The 831-base pair cDNA clone encoding this 277-amino acid protein was next obtained from a library constructed from F. intermedia stem tissue, whose fully functional recombinant protein, produced by expression of this cDNA in Escherichia coli, catalyzed the same enantiospecific conversion via the corresponding lactol intermediate. A homologous secoisolariciresinol dehydrogenase gene was also isolated from a Podophyllum peltatum rhizome cDNA library, whose 834-base pair cDNA clone encoded a 278-amino acid protein with a calculated molecular mass of approximately 32 kDa. Expression of this protein in E. coli produced a fully functional recombinant protein that also catalyzed the enantiospecific conversion of (-)-secoisolariciresinol into (-)-matairesinol via the intermediary lactol. Various kinetic parameters were defined and established conversion of the intermediary lactol as being rate-limiting. With this overall enzymatic conversion now unambiguously defined, the entire biochemical pathway to the lignans, secoisolariciresinol and matairesinol, has been elucidated. Last, both secoisolariciresinol and matairesinol are metabolized in the gut of mammals, following digestion of high fiber dietary grains, seeds, and berries, into the so-called "mammalian" lignans, enterodiol and enterolactone, respectively; these in turn confer significant protection against the onset of breast and prostate cancers.  (+info)

Three new glycosides from Sinopodophyllum emodi (Wall.) Ying. (2/17)

Tow new aryltetralin-type lignan glycosides: methyl epipodophyllate 7'-O-beta-D-glucopyranoyl-(1-->6)-beta-D-glucopyranoside (1), 4-demethylepipodophyllotoxin 7'-O-beta-D-glucopyranoside (2), and a new phenyl ethanol glycoside: phenyl ethanol 4-O-beta-D-xylopyranosyl-(1-->6)-beta-D-glucopyranoside (3), along with three known compounds: junipetriolosides (4), 3,4-dihydroxy-phenyl ethanol (5), and 4-hydroxy-phenyl ethanol (6) were isolated and identified from the n-butanol extract of the roots and rhizomes of Sinopodophyllum emodi (Wall.) Ying. The structures of the above were established by means of spectral data and chemical methods.  (+info)

New lignan glycosides from Chinese medicinal plant, Sinopodophillum emodi. (3/17)

Two new podophillotoxin glucosides, L-picropodophillotpxin 7'-O-(beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranoside) (2) and L-picropodophillotpxin 7'-O-beta-D-glucopyranoside (3), were isolated from Chinese medicinal plant, Sinopodophillum emodi, together with 4 known compounds, podophillotoxin (1), podorhizol 4'-O-beta-D-glucopyranoside (4), deoxypodophillotoxin (5), and dehydropodophillotoxin (6). The structures of 2 and 3 were finally determined by the extensive decouping and nuclear Overhauser effect (NOE) experiments in NMR spectra and circular dichroism (CD) spectra. Compound 2 is the second example of podophillotoxin diglucoside, and both the first one and 2 were isolated from S. emodi. X-ray crystal structure analysis of 1, 5, and 6 was carried out. Compounds 1 and 5 showed the different conformations from those reported.  (+info)

No effect of a homeopathic preparation on neonatal calf diarrhoea in a randomised double-blind, placebo-controlled clinical trial. (4/17)

A double-blind, placebo-controlled clinical trial of a homeopathic treatment of neonatal calf diarrhoea was performed using 44 calves in 12 dairy herds. Calves with spontaneously derived diarrhoea were treated with either the homeopathic remedy Podophyllum (D30) (n = 24) or a placebo (n = 20). No clinically or statistically significant difference between the 2 groups was demonstrated. Calves treated with Podophyllum had an average of 3.1 days of diarrhoea compared with 2.9 days for the placebo group. Depression, inappetence and fever were presented equally in the 2 groups. These results support the widely held opinion that scientific proof for the efficacy of veterinary homeopathy is lacking. In the European Union this implies a considerable risk for animal welfare, since in some countries priority is given to homeopathic treatments in organic farming.  (+info)

The effect of podophyllotoxin, colchicine, urethane, and nitrogen mustard on the respiration of normal and suprarenalectomized rat lymphatic tissue. (5/17)

The injection of podophyllotoxin, colchicine, and a nitrogen mustard derivative, methyl-bis(beta-chloroethyl)amine, into normal animals causes a reduction of the respiration of the lymphatic tissues, spleen, and/or thymus. No effect was demonstrable on kidney, a representative tissue of non-lymphatic origin. The degree of inhibition was considerably less in suprarenalectomized animals, suggesting that the inhibition is mediated by the suprarenal gland.  (+info)

Crystal structures of apo-form and binary/ternary complexes of Podophyllum secoisolariciresinol dehydrogenase, an enzyme involved in formation of health-protecting and plant defense lignans. (6/17)

(-)-Matairesinol is a central biosynthetic intermediate to numerous 8-8'-lignans, including the antiviral agent podophyllotoxin in Podophyllum species and its semi-synthetic anticancer derivatives teniposide, etoposide, and Etopophos. It is formed by action of an enantiospecific secoisolariciresinol dehydrogenase, an NAD(H)-dependent oxidoreductase that catalyzes the conversion of (-)-secoisolariciresinol. Matairesinol is also a plant-derived precursor of the cancer-preventative "mammalian" lignan or "phytoestrogen" enterolactone, formed in the gut following ingestion of high fiber dietary foodstuffs, for example. Additionally, secoisolariciresinol dehydrogenase is involved in pathways to important plant defense molecules, such as plicatic acid in the western red cedar (Thuja plicata) heartwood. To understand the molecular and enantiospecific basis of Podophyllum secoisolariciresinol dehydrogenase, crystal structures of the apo-form and binary/ternary complexes were determined at 1.6, 2.8, and 2.0 angstrom resolution, respectively. The enzyme is a homotetramer, consisting of an alpha/beta single domain monomer containing seven parallel beta-strands flanked by eight alpha-helices on both sides. Its overall monomeric structure is similar to that of NAD(H)-dependent short-chain dehydrogenases/reductases, with a conserved Asp47 forming a hydrogen bond with both hydroxyl groups of the adenine ribose of NAD(H), and thus specificity toward NAD(H) instead of NADP(H). The highly conserved catalytic triad (Ser153, Tyr167, and Lys171) is adjacent to both NAD(+) and substrate molecules, where Tyr167 functions as a general base. Following analysis of high resolution structures of the apo-form and two complex forms, the molecular basis for both the enantio-specificity and the reaction mechanism of secoisolariciresinol dehydrogenase is discussed and compared with that of pinoresinol-lariciresinol reductase.  (+info)

Anti-inflammatory effects of Podophyllum hexandrum (RP-1) against lipopolysaccharides induced inflammation in mice. (7/17)

PURPOSE: Down-regulation of lipopolysaccharide (LPS) induced hyper-inflammatory response by non-toxic pharmacological agents acquires paramount importance for countering bacterial sepsis. Anti-inflammatory potential of aqueous extract of Podophyllum hexandrum, a plant well documented in Ayurvedic literature for various therapeutic purposes, was investigated. METHODS: In vivo studies were performed on Balb/c mice pre-treated with supra-lethal dose of LPS endotoxin (E.coli 055:B5) with or without treatment with P. hexandrum extract (RP-1). Mouse peritoneal macrophage cultures were used to understand ex vivo effects of RP-1 on LPS generated nitric oxide (NO), secretion of IFN-gamma, IL-6 and TNF-alpha. Griess assay and sandwich ELISA method were used to quantify inducible NO and cytokines respectively. RESULTS: Minimal dose of LPS that rendered 100% mortality to mice was found to be 450 microg/kg b.w. Administration of RP-1 (200 mg/kg b.w., i.p.) one hour before lethal LPS treatment (0.5 mg/kg b.w.) rendered maximum (78%) survival. Ex vivo study revealed that RP-1 (50 microg/ml) treatment to peritoneal macrophages inhibited LPS (5 microg/ml) induced nitrite generation to 37%, IFN-gamma secretion to 5%, IL-6 secretion to 50% and TNF-alpha secretion to 50 % of LPS treated control values. CONCLUSION: This study has demonstrated anti-inflammatory potential of aqueous extract of P. hexandrum.  (+info)

Characterization of two epimers, 4alpha and 4beta, of a novel podophyllotoxin-4-O-(D)-6-acetylglucopyranoside from Podophyllum hexandrum by LC-ESI-MS-MS. (8/17)

High-performance liquid chromatography (HPLC) with diode array detection interfaced to electrospray ionization (ESI) mass spectrometry (MS) is applied to identify the two epimers of a novel and minor constituent, podophyllotoxin-4-O-(D)-6-acetylglucopyraniside from high-altitude Podophyllum hexandrum for the first time. This is done by matching the structural information from the tandem MS data with the reported lignan markers. The results show that LC-MS-MS is the method of choice for fast detection and detailed chemical analysis of mixtures in the crude extracts of Podophyllum. The method can be employed in the absence of reference standards for the markers and is particularly useful in view of the scarcity of these rare chemical standards.  (+info)

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