ANTIFUNGAL ACTIVITY OF FLAVONOIDS ISOLATED FROM ULEX SPECIES
P. Máximo, A. Lourenço, S. Feio* and J. Roseiro*
Departamento de Química, Centro de Química Fina e Biotecnologia, Faculdade de Ciências e Tecnologia
Universidade Nova de Lisboa, Quinta da Torre, 2825-114 Caparica, Portugal.
*Instituto Nacional de Engenharia e Tecnologia Industrial IBQTA, Laboratório de Microbiologia Industrial
Azinhaga dos Lameiros, 1699 Lisboa Codex, Portugal.
ACTIVE COMPOUNDS
_____________________________
R2
R1O
O
INTRODUCTION
R2
Isoflavonoids are described in Leguminosae as
phytoallexins and defence agents against insects. There
are different structural factors in isoflavonoid molecules
that are important for their antimicrobial activity. The
presence of isoprenoid groups proved to increase activity
in isoflavons and pterocarpans and the relative acidity
and number of hydroxyl groups are important in
structure-activity relations [1,2].
R1
OH
1
2
3
OH
R3
O
O
O
OH
R1 = CH3
O
O
R2 =
R1 = H R2 =
O
5
R1 = R2 = OCH3
6
R1 = R2 = R3 = OCH3
7
R1 = OCH3 R2 = OH
8
R1 = H R2 = OH R3 = OCH3
9
R1 = R3 = H R2 = OH
R1 = R2 = H
R3 = H
OH
R3 = H
O
11
O
OH
HO
10
O
OH
O
HO
O
O
RESULTS AND DISCUSSION
O
OH
O
12
4
The fungicidal activity of 18 isoflavonoids together with 2 chalcones, 1 flavone and 1
isoflavanone, isolated from Ulex species, was tested by the bioautographic method against
Cladosporium cucumerinum. Most of the test compounds showed antifungal activity and
inhibited the fungal growth at 100 µg concentration. From the experimental results some
structure-activity comments can be drawn.
Among the 11 isoflavones tested (1-4,13-17,20,21) the structures with an open chain
isoprenoid substituent at C-8 were active (1,2,4). The C-6 substituted compounds (1315,17) had no activity, except for one dimethylpyrano substituted compound 21 that partially
inhibited the fungal growth. All the pterocarpans studied (5-9,12,22) have an 8,9methylenodioxy group and their structural differences are in ring A substitution. The
presence of a C-2 hydroxyl group imparts lack (22) or only partial fungicidal activity (18).
The presence of hydroxyl groups seems to play an important role for antifungal behaviour of
this kind of compounds.
O
H3CO
O
NON ACTIVE COMPOUNDS
_______________________________________
HO
O
HO
OH
R
O
13
OH
O
16
O
OH
15
R=
OH
OCH3
HO
OH
OH
O
O
O
O
HO
17
O
EXPERIMENTAL
O
18
OH
Cell suspension for thin-layer chromatography (TLC) bioassay
Cladosporium cucumerinum CCMI 206 was grown on malt extract agar at 25ºC in pyrex Petri dishes for ten days.
O
O
The mycelium was harvested from the agar plates in a small volume of fresh Homans and Fuchs nutrient broth
[3], filtered through four layers of sterilised gauze and diluted in nutrient broth in order to obtain 106 cell ml-1.
Bioautographic TLC bioassay
Aliquots of the test compounds (100 mg) were spotted, in quadruplicate, on silica gel 60 F254 TLC plates (Merck
5554), which were eluted with the appropriate eluent for each sample (CHCl3-MeOH 0.6% - 1.25% mixts).
Developed chromatograms were dried and the spots of each compound were marked under 254 nm UV light. A 20
ml sample of the cell suspension was sprayed evenly over each plate, in a glove box. Plates were incubated in
closed pyrex trays lined with moist paper at 25ºC for two-three days, protected from light. Bioautograms were
evaluated by clear spots, indicating zones of inhibition.
O
O
OH
O
19
O
SLIGHTLY ACTIVE COMPOUNDS
______________________________________________
HO
O
OH
21
H3CO
OH
R=
O
O
H3CO
O
O
OH
14
O
O
O
O
O
20
O
O
HO
O
O
22
Acknowledgments
We wish to thank the people from Herbário, Museu, Jardim Botânico,
Faculdade de Ciências, Universidade de Lisboa (Portugal) for collecting plant material.
One of us (P. M.) wishes to thank FCT (Portugal) for a PRAXIS XXI fellowship.
References
[1] Tahara, S. and Ibrahim, R.K. (1995) Phytochemistry 38, 1073.
[2] Tahara, S., Katagiri, Y., Ingham, J.L. and Mizutani, (1994) Phytochemistry 36, 1261.
[3] Homans A.L. and Fushs A. (1970) J. Chromatogr. 51, 327-329.