Aspergillus flavus as a Model System to Test the Biological Activity of Botanicals: An Example on Citrullus colocynthis L. Schrad. Organic Extracts

Abstract

Citrullus colocynthis L. Schrader is an annual plant belonging to the Cucurbitaceae family, widely distributed in the desert areas of the Mediterranean basin. Many pharmacological properties (anti-inflammatory, anti-diabetic, analgesic, anti-epileptic) are ascribed to different organs of this plant; extracts and derivatives of C. colocynthis are used in folk Berber medicine for the treatment of numerous diseases-such as rheumatism arthritis, hypertension bronchitis, mastitis, and even cancer. Clinical studies aimed at confirming the chemical and biological bases of pharmacological activity assigned to many plant/herb extracts used in folk medicine often rely on results obtained from laboratory preliminary tests. We investigated the biological activity of some C. colocynthis stem, leaf, and root extracts on the mycotoxigenic and phytopathogenic fungus Aspergillus flavus, testing a possible correlation between the inhibitory effect on aflatoxin biosynthesis, the phytochemical composition of extracts, and their in vitro antioxidant capacities.

Keywords: Aspergillus flavus; Citrullus colocynthis; HPLC-MS/MS; antimycotoxigenic activity; model system.

Figure 1

DPPH scavenging activity of Citrullus colocynthis root, leaf, and stem extracts. Data are means of three replicates ± S.D. Same letters indicate absence of statistically significant differences (p < 0.05).

Figure 2

Activity on toxin accumulation and mycelium growth. Aflatoxin accumulation (reported as fluorescence arbitrary units) in A. flavus six-days after CCM cultures treated with stem (A), leaf (B), and root (C) extracts. (D) Early mycelium growth inhibition of A. flavus conidia treated with 500 µg/mL extracts, measured 48 h after inoculum by optical density increasing, and expressed as percentage in respect to control. Error bars refer to mean values of four replicates ± S.D.

Figure 3

Aflatoxin time-course accumulation. Effect of stem (A), leaf (B), and root (C) extracts on AF time-course accumulation in A. flavus. (D) Comparison between 500 µg/mL stem, leaf, and root CHL extract; α-lipoic acid 1 mM is used as a reference. Error bars refer to mean values of four replicates ± SD.

Figure 4

Effect of 500 µg/mL CHL (A) and EA (B) root extract over time on aflatoxin production. Extracts were added to conidia of A. flavus inoculated in CCM after 65, 72, and 86 h of incubation. Error bars refer to mean values of four replicates ± SD.

Figure 5

Effect of 500 µg/mL root, stem, leaf (R, S, L) MET and CHL extracts on conidia production (A) and conidiophores morphology (B) (left: Control; right: R-CHL treated cultures) in A. flavus 96-wells CCM cultures. Values are reported as inhibition percentages in respect to control; Error bars refer to mean values of four replicates ± S.D. Different letters over the bars indicate the differences that were statistically significant (p < 0.05).