INVESTIGADORES
PARAJE Maria Gabriela
congresos y reuniones científicas
Título:
Mode of action of extracts from a phototoxic plant on fungal biofilms.
Autor/es:
JULIANA MARIONI, JULIO E ARCE MIRANDA, LAURA R COMINI, JOSÉ L CABRERA, SUSANA C NÚÑEZ MONTOYA, MARÍA G PARAJE.
Lugar:
Rosario
Reunión:
Congreso; X Congreso Argentino de Microbiología General (SAMIGE); 2013
Institución organizadora:
SAMIGE
Resumen:
In recent years, different species from Candida no albicans are becoming the main pathogens causing
nosocomial infections in our country, especially because of their ability to form biofilms.
Heterophyllaea pustulata Hook. f. (Rubiaceae) is a phototoxic vegetal species from NW Argentina and
Bolivia, popularly known as ??cegadera?. Previously, only the anthraquinones (AQs)-rich extracts have
been shown to have in vitro antibacterial and antifungal effects against planktonic microorganisms,
with a low acute toxicity in vivo. The aim of this study was to evaluate the activity of different polarity
extracts from H. pustulata against biofilms formed by a Candida no albicans strain, and to investigate if
any effects could be enhanced by light, quantifying the reactive oxygen species (ROS) and reactive
nitrogen intermediate (RNI) generation.
Four extracts were obtained: hexane (Ext-H), benzene (Ext-B), ethyl acetate (Ext-AE), and ethanol
(Ext-E) from the aerial parts of this plant. Biofilm quantification was performed by the O?Toole & Kolter
method, using an ATCC strain of C. no albicans. Sensitivity to the extracts was determined following
the protocols of the Clinical and Laboratory Standards Institute, at three concentrations (0.2, 0.1 and
0.05 mg/ml) in triplicate. The assay was carried out under two conditions: darkness and irradiation,
simultaneously. The irradiation system comprised a 20W Phillips actinic lamp (380? 480 nm, 0.65
mW/cm2) with a maximum at 420 nm, placed inside a black box at 20 cm above the samples. The
supernatant was separated by measuring the superoxide anion (O2
·- ) production by the reduction of
the nitro-blue tetrazolium (NBT) reaction; the nitric oxide (NO) generation was evaluated by Griess
reagent; and the total system antioxidant capability was determined through FRAP assay (Ferrous
Reduction Antioxidant Potency).
No extracts tested resulted active under darkness, while under irradiation Ext-B, AE y E showed the
ability to decrease biofilms with the most active being Ext-E (biofilm reduction of 34.8±2.3 % at 0, 2
mg/ml). Furthermore, when the system was irradiated, O2
·- and NO were generated. The total
system´s antioxidant capability was higher with respect to darkness.
We established that the ability of the extracts obtained from H. pustulata to inhibit the biofilm growth of
ATCC C. no albicans was correlated with the irradiation effect that caused the generation of two
reactive toxic species: O2
·- and NO. These results encouraged us to evaluate the qualitative and
quantitative bioactive extract composition, try to explain the observed effect. Since Candida biofilms
are resistant to the antifungal agents used in medicine, the natural compounds found to be active
against this form of growth, warrants the continuity of our research, in order to assess the effects of
the pure compounds both isolated and purified from extracts of H. pustulata.