(−)-Pisatin, an induced pterocarpan metabolite of abnormal configuration from Pisum sativum

Banks, Simon R.

doi:10.1016/0031-9422(82)85025-5

Cited by 3 publications

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“…( )! _E(1 -E)Nn (1) where N is the number of labelable atoms and E is the overall enrichment of the label (6). The observed distribution was in excellent agreement with the prediction based on three labelable atoms (Table I).…”

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Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Matthews¹,

Weiner²,

Matthews³

et al. 1987

Plant Physiol.

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Some isolates of the plant pathogen Nectria haematococca detoxify the isoflavonoid phytoalexin (-)maackiain by hydroxylation at carbon 6a. Precursor feeding studies strongly suggest that the penultimate step in (+)pisatin biosynthesis by Pisum sativum is 6a-hydroxylation of (+)maackiain. We have used "O labeling to test the involvement of oxygenases in these two reactions. When fungal metabolism of maackiain took place under '"02, the product was labeled with 99% efficiency; no label was incorporated by metabolism in H210. Pisatin synthesized by pea pods in the presence of 1802 or H2180 was a mixture of molecules containing up to three labeled oxygen atoms. Primary mass spectra of such mixtures were complex but were greatly simplified by tandem MS. This analysis indicated that the 6a oxygen of pisatin was derived from H20 and not from 02. Labeling patterns for the other five oxygen atoms were consistent with the proposed pathway for biosynthesis of pisatin and related isoflavonoids. We conclude that the fungal hydroxylation of maackiain is catalyzed by an oxygenase, but the biosynthetic route to the 6a hydroxyl of pisatin is unknown.Several pterocarpan phytoalexins are known to be detoxified by Nectria haematococca or by related fungi with the same imperfect state, Fusarium solani. Pisatin is demethylated by a Cyt P-450 monooxygenase (16). Phaseollin is hydroxylated to la-hydroxyphaseollone by a reaction which also appears to be oxygenase-mediated, since 180 labeling has shown that the new oxygen atom is derived from 02 (15). Maackiain and medicarpin are metabolized by one or more of three different initial reactions, depending on the fungal isolate; all three involve the net addition of one oxygen atom (I1, 12). One of these reactions produces the la-hydroxydienone analogous to the phaseollin metabolite. Another, the hydroxylation at carbon 6a, also involves incorporation of an atom from 02, as reported below.Oxygenases are also thought to be responsible for several reactions in the biosynthesis of these phytoalexins. In particular, the 6a-hydroxylation of maackiain has been proposed to be a step in pisatin biosynthesis by Pisum sativum (Fig. 1) (2-4). This reaction is not identical with the fungal conversion, since the fungus metabolized only (-)maackiain (23) whereas the expected precursor of (+)pisatin, the isomer normally produced by pea, would be the enantiomer (+)maackiain. larity between these two reactions, which appear to play such opposed roles in the interaction between host and parasite, prompted us to examine both of them in this study.MATERIALS AND METHODS Biological Materials. Nectria haematococca Berk. and Br. mating population VI isolate T-95, from our culture collection (22, 23), was maintained on V-8 juice agar and grown in glucoseasparagine (GA) liquid medium as described (25). Pisum sativum L. cultivar Alaska was grown in a greenhouse.6a-Hydroxylation of Maackiain. A 3-d culture of N. haematococca in GA medium was filtered, weighed, and suspended in double strength GA medium at 60 m...

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confidence: 99%

Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Matthews¹,

Weiner²,

Matthews³

et al. 1987

Plant Physiol.

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“…The same authors found that when CuC12-treated seedlings were fed (-)maackiain or (-)HMK, a substantial proportion of the pisatin synthesized was the (-)isomer, whereas none of this metabolite was detected in the absence of these precursors (1,3). Since the quantity of (-)pisatin obtained was much greater than could be accounted for by incorporation of the radiolabeled precursors, it was concluded that a new enzymic pathway leading to (-)pisatin had been induced, a pathway not present in seedlings treated with CuC12 alone.…”

Section: Cuci2-treated Seedlingsmentioning

confidence: 95%

Synthesis of the Phytoalexin Pisatin by a Methyltransferase from Pea

Matthews¹,

VanEtten²

1986

Plant Physiol.

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Previous labeling studies in vivo suggest that the terminal step of (+)pisatin biosynthesis in Pisum sativum L. is methylation of the phenol (+)6a-hydroxymaackiain (HMK). We have found that extracts from pea seedlings perform this reaction, using S-adenosylmethionine as the methyl donor. The enzyme activity was induced by microbial infection or treatment with CuC12, which elicit pisatin synthesis, though some activity was also present in healthy tissues. It has been reported that CuCl2-treated pea tissue provided with (-)HMK or (-)maackiain can synthesize (-)pisatin. Our extract showed no methyltransferase activity dependent on either of these substrates. Methylation of (+)maackiain was detectable, but much slower than that of (+)HMK.Pisatin is one of a large group of low mol wt toxic compounds known to be synthesized de novo by plant tissues in response to certain kinds of stress, including microbial infection. Such compounds, called phytoalexins, are of interest in part because of their possible role as a determinant ofdisease resistance in plants. In addition, phytoalexin synthesis has been a useful subject for studies on the regulation of gene expression in higher plants. These studies have shown that the accumulation of isoflavonoid phytoalexins in legumes is preceded by increases in the levels of some of the biosynthetic enzymes (4, 10); where examined, the increases have been found to be mediated primarily by higher concentrations of the corresponding mRNAs (15,16).The enzymes whose regulation has been examined in most detail are phenylalanine ammonia-lyase and chalcone synthase, enzymes required not only for phytoalexin synthesis but also for other important pathways of secondary metabolism. These enzymes are also inducible by some stimuli which do not elicit phytoalexin synthesis (8, 12), so it seems likely that steps in the later, committed portion of the phytoalexin pathway are important in controlling the plant's phytoalexin response. Unfortunately, the enzymes catalyzing these late reactions are largely unknown. Recently two such enzymes were obtained from soybean. Levels of these enzyme activities, a monooxygenase (9) and a dimethylallyltransferase (11), were also increased by conditions which elicited phytoalexin production.We now report that Pisum sativum possesses an inducible methyltransferase which catalyzes the terminal step in the biosynthesis of (+)pisatin, the major phytoalexin produced by this plant. MATERIALS AND METHODS Plant Material. Pea seeds (cv Alaska) were treated for 15 min with a 1:20 dilution of commercial bleach (5.25% NaOCl) in 80% ethanol, and maintained under sterile conditions until harvested. The seeds were rinsed with H20, spread on paper toweling, and imbibed overnight with 4 ml H20/g seeds at 18°C in the dark. Excess water was removed and germination was continued for 3 d, at which time the radicles were 2 to 4 cm long. The seedlings were covered with 5 mM CuCl2 (control seedlings with H20) for 1 h, drained, and incubated further until used for enzyme preparati...

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The Chemical and Biologic Profile of a Red Clover (Trifolium pratense L.) Phase II Clinical Extract

Booth

Overk

Yao

et al. 2006

The Journal of Alternative and Complementary Medicine

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The major and minor chemical and active estrogenic components of a preformulated phase II red clover clinical extract were identified, quantitatively measured, and the final capsule doses were calculated. The extract is currently under evaluation in a year-long clinical study for the alleviation of menopausal hot flashes. This is the first report to thoroughly summarize the chemistry and biology of all major peaks observed in the HPLC-UV chromatogram of a clinical red clover dietary supplement.

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(−)-Pisatin, an induced pterocarpan metabolite of abnormal configuration from Pisum sativum

Cited by 3 publications

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Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Synthesis of the Phytoalexin Pisatin by a Methyltransferase from Pea

The Chemical and Biologic Profile of a Red Clover (Trifolium pratense L.) Phase II Clinical Extract

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