*Corresponding author. Tel.:#56-45-325429; fax:#56-45-325440.
E-mail address:fpardo@ufro.cl (F. Pardo)
Biochemical Systematics and Ecology 28 (2000) 911}913
Stigmast-4-ene-3,6-dione an unusual phytotoxic
sterone from the roots of
Echium
v
ulgare
L.
Fernando Pardo
!
,
*
, Fernando Perich
!
, Rene
H
Torres
"
,
Franco Delle Monache
#
!Departamento de Ciencias Qun&micas, Universidad de La Frontera, Casilla 54-D, Temuco, Chile "Departamento de Ciencias Qun&micas y Biologn&a, Universidad de Santiago de Chile, Casilla 44, Correo 33,
Santiago, Chile
#Centro Chimica Recettori (CNR), Largo Francesco Vito 1, 00168, Roma, Italy
Received 11 May 1999; accepted 6 December 1999
Keywords: Echiumvulgare; Boraginaceae; Germination inhibitors; Weeds; Sterones
1. Subject and source
Echiumvulgare L. (blueweed, Boraginaceae) a weed indigenous to Europe which commonly grows in cereal crops in Southern Chile (Espinoza, 1988); source: roots of
Echium vulgare was collected in CajoHn, Temuco, in December 1997; a voucher specimen (FPD 30) is deposited in the Department of Chemical Sciences, Universidad de La Frontera, Temuco.
2. Previous work
Echium vulgare: quinoid pigments and phenolic compounds (Sherbanivs'kii, 1971; Delorme et al., 1977; Fedorev et al., 1979; Huizing et al., 1981), pyrrolizidine
alkaloids (Man'ko, 1964; Pedersen, 1975; Kamirov et al., 1975; Delorme et al.,
1977; Huizing et al., 1981; El-Shazly et al., 1996) andc-linolenic acid (Penkova et al.,
1990).
3. Present study
In a previous study the ethanolic extract of the roots of E. vulgare had been
shown to inhibit barley (Hordeum vulgare) germination (Pardo et al., 1998).
Further fractionation of the ethanolic extract on a silica gel #ash column eluted
sequentially with CHCl
3, Et2O, EtOAc and Me2CO, led to an active Et2O fraction,
which was re-chromatographed over silica gel (hexane-C
6H6/2 : 1), and the combined
active fractions were puri"ed by preparative TLC (Merck 60 F
254) using
hexane-EtOAc (10 : 1.5). Two major active compounds,b-sitosterol (Rf 0.25) and the
sterone stigmast-4-ene-3,6-dione (Rf 0.40) were isolated and further identi"ed by
spectroscopic studies. This sterone was previously isolated from Sambucus ebulus
(Tunman and Grimm, 1974) and NMR data obtained here are reported for the"rst
time. NMR spectra were recorded in CDCl
3 on a Varian XL-300 spectrometer,
operating at 300 MHz for1H and 75 MHz for 13C nuclei. The chemical shifts are
reported in d (ppm), with TMS as an internal standard. Multiplicity and
coupling constants (J/Hz) are given in parenthesis. 13C NMR(CDCl
3):
202.4(C-6), 199.5(C-3), 161.1(C-5), 125.4(C-4), 56.5(C-14), 55.8(C-17), 51.0(C-9), 46.8(C-7), 45.8(C-24), 42.5(C-13), 39.8(C-10), 39.1(C-12), 36.0(C-20), 35.5(C-1), 34.2(C-8), 34.0(C-2) 33.8(C-22), 29.1(C-25), 28.0(C-16), 26.0(C-23), 24.0(C-15), 23.0(C-234.2(C-8),
20.9(C-11), 19.8(Me-27), 19.0(Me-26), 18.7(Me-21), 17.5(Me-19), 12.0(Me-29),
11.9(Me-18).1H NMR(CDCl
3): 6.17(brd,J"0.8 Hz; H-4), 2.68 and 2.02(H2-2), 2.52
and 2.14(H2-7), 1.17(s,Me-18), 0.93(d, J"6.4 Hz;Me-21), 0.85-0.80(Me-26, Me-27,
Me-29), 0.72(s,Me-19). INEPT experiments: Irradiation at delta 6.17(H-4) gave re-sponse for C-6(202.4) and C-10(39.8); irr.at delta 1.17(Me-18) g.r. for C-5(161.1), C-9(50.9), C-10(39.8) and C-1(35.5); irr.at delta 0.72(Me-19) g.r. for C-14(56.5) and C-13(42.5).
In the barley bioassay, the sterone inhibited germination by 40% at a concentration
of 3 mM, and also showed a moderate stimulatory e!ect on root growth at
concentra-tions below 0.1 mM. b-Sitosterol showed lesser stimulatory as well as inhibitory
e!ects in the bioassays.
4. Chemotaxonomic signi5cance
The presence of the sterone stigmast-4-ene-3,6-dione is reported for the"rst time in
the genus Echium. This sterone could be assumed to be a biogenetic precursor of
brassinosteroids, which represent steroidal plant growth regulators with high plant growth-promoting and anti-stress activity.
Acknowledgements
This research was supported by DIDUFRO 9914, DICYT and Programa de
CooperacioHn CientmH "ca Internacional, FundacioHn Andes/CONICYT.
References
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