この論文は国立情報学研究所の学術雑誌公開支援事業により電子化されました

SHIMADA, Mikio

SUZUKI, Shiro

UMEZAWA, Toshiaki

Kyoto University Research Information Repository

Title<Preliminary>Stereochemical Selectivity inSecoisolariciresinol Formation by Cell-free Extracts fromArctium lappa L. Ripening SeedsAuthor(s)SUZUKI, Shiro; UMEZAWA, Toshiaki; SHIMADA, MikioCitationWood research : bulletin of the Wood Research Institute KyotoUniversity (1999), 86: 37-38Issue Date1999-09-30URL http://hdl.handle.net/2433/53159RightType Departmental Bulletin PaperTextversionpublisherKyoto UniversityPrelimina.!L.Stereochemical Selectivity in SecoisolariciresinolFormation by Cell-free Extracts fromArctium lappa L. Ripening Seeds*lShiro SUZUKI*2, Toshiaki UMEZAWA*2 and Mikio SHIMADA*2(Received May 31, 1999)Keywords: Arctium lappa, ripening seeds, enantioselective, stereochemistry, lignan, biosynthesisLignans are a class of secondary metabolites found as 8,8'-linked dimers of two phenylpropane units l ,2). Theenantiomeric compositions of a particular lignan can varywith plant species, and the predominant enantiomer candiffer among the plant sources3- 7). For example, (-)-matairesinol was isolated from Forsythia intermedia, whereasthe (+ )-antipode was isolated from Wikstroemia siko-kiana6 ,11). In addition, lignans have various biologicalactivities such as antitumor, antimiotic and antiviral, andsome of them accumulate in large amounts in heartwood ofwoody plants7,8).The biosynthetic mechanism of lignans has long beenmarked among phytochemists because of their abovefeatures. In 1990, U mezawa et al. firstly demonstratedthe enantioselective formation of optically pure (-)-secoisolariciresinol from achiral coniferyl alcohol with cell-free extracts from F. intermedia in the presence ofNAD(P)Hand H 20 29 ). Since then, many reports have beenpublished on the enzyme systems involved in lignanbiosynthesis in Forsythia plants as enzyme sources, mostlyby Lewis and co-workers 10-20). As a whole, these studiesrevealed enzymatic formation of the naturally occurringenantiomers of Forsythia lignans and each enzymaticconversion was controlled strictly with respect tostereochemis try.On the other hand, there are many examples of plantswhich produce the opposite enantiomers to those of thelignans occurring in Forsythia Spp.3-7,21). Using Arctiumlappa petioles as an enzyme source, Umezawa and Shimadareported an enzyme activity that catalyzed the formation ofthe opposite enantiomer of secoisolariciresinol to thatformed by Forsythia enzyme preparation22). 1n contrast tocell-free extracts from Forsythia plants, the enzymepreparation from A. lappa petioles catalyzed theenantioselective formation of (+ )-secoisolariciresinol (ca.20% enantiomer excess (e.e.)) from coniferyl alcohol in thepresence of NADPH and H 20 2. The (+ )-enantiomer ofsecoisolariciresinol was the predominant antipode ofsecoisolariciresinol isolated from A. lappa petioles (78%e.e.).The absolute configuration at C8 position of (+)-secoisolariciresinol is, however, opposite to that of (-)-arctigenin, which is isolated in significant amounts fromthe seeds23,24). Thus, the stereochemical mechanism forproducing (+ )-secoisolariciresinol and (- )-arctigenin inA. lappa awaits further experimental works.This paper describes the formation of (- )-seco-isolariciresinol with cell-free extracts from ripening seeds ofA. lappa, demonstrating that the different organs of A. lappaproduced enzymes with the opposite stereochemicalselectivity in secoisolariciresinol formation.A preliminary GC-MS analysis indicated that theproduction of lignans in A. lappa seeds was devel-opmentally regulated. Trace amounts of lignans(pinoresinol, lariciresinol and secoisolariciresinol) werepresent in both the flower buds and flowers in bloom of A.lappa, while the amount was significantly greater inripening seeds ca. 6 days after the start ofblooming. Next,cell-free extracts were prepared from ripening seeds of A.lappa as described previousll2,25). When [9, 9-2H2,OC2H 3Jconiferyi alcohol was incubated with the cell-freeextracts In the presence of NADPH and H 20 2,~~ sOH H20,I ...0 OCH3OH HOConiferylalcoholOCH3Pinoresinol LariciresinolH3COOCH3 HONADPH ..H3COO~~I 88' 0HCO .03 H......_ [~O.0 OCHOH 3(-)-Arcti geninFig. 1. A possible biosynthetic pathway of lignans in Arctium lappa seeds.*' A part of this work was presented in the 48th annual meeting ofJapan Wood Research Society, April 4, 1998, Shizuoka, Japan.*2 Laboratory of Biochemical Control.-37-WOOD RESEARCH No.86 (1999)eH 10] pinoresinol, [2H 10] lariciresinol and [2H 10] seco-isolariciresinol were formed. They were identified bycomparing their mass spectra and retention times on GCwith those of authentic samples. The enantiomericcompositions of the lignans formed enzymatically from [9,9-2H2, OC 2H 3]coniferyl alcohol were analyzed by chiralHPLC and GC-MS· as described previousl/2,25). Theenantiomeric composi tion of [2H lO]secoisolariciresinol was38% e.e. [(-»(+)].This result, therefore, strongly suggests that ( - )-secoisolariciresinol was enantioselectively formed fromconiferyl alcohol via pinoresinol and lariciresinol (Fig. I).Next we incubated (± )-[9,9,9',9'-2 H4]pinoresinols and(± )-[9, 9, 9', 9'-2H4]lariciresinols individually with thecell-free extracts in the presence ofNADPH. The cell-freeextracts catalyzed the selective formation of (-)-[2H4]secoisolariciresinol in both assays. Furthermore,the (- )-enantiomer of secoisolariciresinol which wasformed selectively in the enzymatic reaction was alsopredominant in this lignan isolated from A. lappa seeds(65% e.e.).Thus, this is the first report that indicates clearly thattwo enzyme preparations from different organs of a singleplant species catalyzed the selective formation of differentenantiomers of lignans, and strongly suggests that therewere organ-specific isozymes that catalyzed the seco-isolariciresinol formation with different enantioselectivityin A. lappa. This view is in good accordance with thestudy by Fujita et al. 26) ; they reported the clonings of twoputative pinoresinolliariciresinol reductase cDNAs fromthe cDNA library of Thuja plicata, and E. coli-expression ofthe recombinant proteins. The two putative reductases~howed different selectivity in secoisolariciresinol for-mation in terms of the substrates. Thus, one catalyzedthe formation of optically pure (- )-secoisolariciresinolfrom (+ )-pinoresinol, while the other catalyzed theformation of optically pure (+ )-secoisolariciresinol from( - )-pinoresinol.We have recen tly reported tha t there is stereochemicaldiversity in lignan biosynthesis among various plantspecies3,27). The present results go further in suggestingthat different stereochemical mechanisms in lignanformation can operate even in different organs of a singleplant species.ReferencesI) D.C. AYRES and J.D. LOIKE: "Lignans: Chemical,biological and clinical properties", Cambridge UniversityPress (1990).2) W.D. MACRAE and G.H.N. TOWERS: Phytochemistry, 23,1207-1220 (1984).3) T. UMEZAWA, T. OKUNISHI and M. SHIMADA: WoodResearch, 84, 62-75 (1997).4) T. OKUNISHI, T. UMEZAWA and M. SHIMADA: WoodResearch, 84, 25-27 (1997).5) T. OKUNISHI, T. UMEZAWA and M. SHIMADA:J. Wood. Sci.,in press.6) T. UMEZAWA and M. SHIMADA: Mokuzai Gakkaishi, 42,180-185 (1996).7) T. UMEZAWA and M. SHIMADA: Mokuzai Gakkaishi, 42,911-920 (1996).8) KP. SWAN, K.S.JIANG andJ.A.F. GARDNER: Phytochemistry,8, 345-351 (1969).9) T. UMEZAWA, L.B. DAVIN and N.G. LEWIS: Biochem.Biophys. Res. Commun., 171, 1008-1014 (1990).10) T. UMEZAWA, L.B. DAVIN, K YAMAMOTO, D.G.I. KINGSTONand N.G. LEWIS: J. Chem. Soc., Chem. Commun., 1405-1408(1990).II) T. UMEZAWA, L.B. DAVIN and N.G. LEWIS: J. BioI. Chem.,266, 10210-10217 (1991).12) L.B. DAVIN, D.L. BEDGAR, T.KATAYAMA and N.G. LEWIS:Phytochemistry, 31, 3869-3874 (1992).13) T. KATAYAMA, L.B. DAVIN and N.G. LEWIS: Phytochemistry,31, 3875-3881 (1992).14) T. KATAYAMA, L.B. DAVIN, A. CHU and N.G. LEWIS:Phytochemistry, 33, 581-591 (1993).15) A. CHU, A. DINKOVA, L.B. DAVIN, D.L. BEDGAR and N.G.LEWIS: J. Bio!. Chem., 268, 27026-27033 (1993).16) S. OZAWA, L.B. DAVIN and N.G. LEWIS: Phytochemistry, 32,643-652 (1993).17) T. UMEZAWA, H. KURODA, T. ISOHATA, T. HIGUCHI and M.SHIMADA: Biosci. Biotech. Biochem., 58, 230-234 (1994).18) P.W. PARE, H.-B. WANG, L.B. DAVIN and N.G. LEWIS:Tetrahedron Lett., 35, 4731-4734 (1994).19) AT. DINKOVA-KoSTOVA, D.R. GANG, L.B. DAVIN, D.L.BEDGAR, A. CHU and N.G. LEWIS: J. Bio!. Chem., 271,29473-29482 (1996).20) L.B. DAVIN, H.-B. WANG, A.L. CROWELL, D.L. BEDGAR,D.M. MARTIN, S. SARKANEN and N.G. LEWIS: Science, 275,362-366 (1997).21) C.B.S. RAO: "Chemistry of Lignans", Andhra UniversityPress (1978).22) T. UMEZAWA and M. SHIMADA: Biosci. Biotech. Biochem., 60,736-737 (1996).23) J. SHINODA and M. KAWAGOYE: Yakugaku Zasshi, 49,565-575 (1929).24) T. OMAKI: Yakugaku Zasshi, 55, 816-827 (1935).25) S. SUZUKI, T. UMEZAWA and M. SHIMADA: Biosci. Biotech.Biochem., 62, 1468-1470 (1998).26) M. FUJITA, D.R. GANG, L.B. DAVIN and N.G. LEWIS: J.Bio!. Chem., 274, 618-627 (1999).27) T. UMEZAWA, T. OKUNISHI and M. SHIMADA: "ACSSymposium Series Volume 697, Lignin and LignanBiosynthesis," eds. by N.G. Lewis and S. Sarkanen,American Chemical Society, Washington D.C., pp.377-388 (1998).- 38-

<Preliminary>Stereochemical Selectivity in Secoisolariciresinol Formation by Cell-free Extracts from Arctium lappa L. Ripening Seeds

http://repository.kulib.kyoto-u.ac.jp/dspace/bitstream/2433/53159/1/KJ00000018048.pdf

<Preliminary>Stereochemical Selectivity in Secoisolariciresinol Formation by Cell-free Extracts from Arctium lappa L. Ripening Seeds

Abstract

Similar works

Full text

Available Versions

Kyoto University Research Information Repository