The Mechanism of Antifungal Action of Essential Oil from Dill (Anethum graveolens L.) on Aspergillus flavus

The essential oil extracted from the seeds of dill (Anethum graveolens L.) was demonstrated in this study as a potential source of an eco-friendly antifungal agent. To elucidate the mechanism of the antifungal action further, the effect of the essential oil on the plasma membrane and mitochondria of Aspergillus flavus was investigated. The lesion in the plasma membrane was detected through flow cytometry and further verified through the inhibition of ergosterol synthesis. The essential oil caused morphological changes in the cells of A. flavus and a reduction in the ergosterol quantity. Moreover, mitochondrial membrane potential (MMP), acidification of external medium, and mitochondrial ATPase and dehydrogenase activities were detected. The reactive oxygen species (ROS) accumulation was also examined through fluorometric assay. Exposure to dill oil resulted in an elevation of MMP, and in the suppression of the glucose-induced decrease in external pH at 4 µl/ml. Decreased ATPase and dehydrogenase activities in A. flavus cells were also observed in a dose-dependent manner. The above dysfunctions of the mitochondria caused ROS accumulation in A. flavus. A reduction in cell viability was prevented through the addition of L-cysteine, which indicates that ROS is an important mediator of the antifungal action of dill oil. In summary, the antifungal activity of dill oil results from its ability to disrupt the permeability barrier of the plasma membrane and from the mitochondrial dysfunction-induced ROS accumulation in A. flavus

Anti-Nociceptive and Anti-Inflammation Effect Mechanisms of Mutants of Syb-prII, a Recombinant Neurotoxic Polypeptide

Syb-prII, a recombinant neurotoxic polypeptide, has analgesic effects with medicinal value. Previous experiments indicated that Syb-prII displayed strong analgesic activities. Therefore, a series of in vivo and vitro experiments were designed to investigate the analgesic and anti-inflammatory properties and possible mechanisms of Syb-prII. The results showed that administered Syb-prII-1 and Syb-prII-2 (0.5, 1, 2.0 mg/kg, i.v.) to mice significantly reduced the time of licking, biting, or flicking of paws in two phases in formalin-induced inflammatory nociception. Syb-prII-1 inhibited xylene-induced auricular swelling in a dose-dependent manner. The inhibitory effect of 2.0 mg/kg Syb-prII-1 on the ear swelling model was comparable to that of 200 mg/kg aspirin. In addition, the ELISA and Western blot analysis suggested that Syb-prII-1 and Syb-prII-2 may exert an analgesic effect by inhibiting the expression of Nav1.8 and the phosphorylation of ERK, JNK, and P38. Syb-prII-1 markedly suppressed the expression of IL-1β, IL-6, and TNF-α of mice in formalin-induced inflammatory nociception. We used the patch-clamp technique and investigated the effect of Syb-prII-1 on TTX-resistant sodium channel currents in acutely isolated rat DRG neurons. The results showed that Syb-prII-1 can significantly down regulate TTX-resistant sodium channel currents. In conclusion, Syb-prII mutants may alleviate inflammatory pain by significantly inhibiting the expression of Nav1.8, mediated by the phosphorylation of MAPKs and significant inhibition of TTX-resistant sodium channel currents

In Vitro and In Vivo Activities of Essential Oil from the Seed of Anethum graveolens L. against Candida spp.

The essential oil produced from the seed of Anethum graveolens L. (Umbelliferae) was tested in vitro and in vivo anti-Candida activity. The microbroth dilution method was used in the minimal inhibitory concentration (MIC), according to M27-A3 of the guidelines of the Clinical and Laboratory Standard Institute (CLSI). And then, efficacy evaluation of essential oil in the prophylaxis and treatment of experimental vaginal candidiasis was performed in immunosuppressed mice. The anti-Candida activity was analyzed by microbiological and histological techniques and was compared with that of fluconazole (FCZ). The results showed essential oil was active in vitro against all tested strains, with MICs ranging from 0.312 μL/mL (for C. tropicalis, C. parapsilosis, and C. krusei) to 0.625 μL/mL (for 6 isolated C. albicans strains). Essential oil (2% v/v) was highly efficacious in accelerating C. albicans 09-1555 clearance from experimentally infected mice vagina by prophylaxis and therapeutic treatments. In both therapeutic efficacy and prophylaxis studies, the histological findings confirmed the microbiological results. The experimental results revealed that the tested essential oil is effective against vulvovaginal candidiasis in immunosuppressed mice

Efficacy of dill oil on the activities of the mitochondrial ATPase of <i>A. flavus</i> cells.

<p>Significant differences (<i>p</i><0.05) between means are indicated by letters above the histogram bars. Values are means (n = 3) ± standard deviations.</p

Inhibition of ergosterol biosynthesis in <i>A. flavus</i> by dill oil.

<p>UV spectrophotometric sterol profiles of <i>A. flavus</i> cells treated with dill oil and in comparison with those of an untreated control.</p

Efficacy of dill oil on the endogenous ROS in <i>A. flavus</i> cells detected by flow cytometry.

<p>(A) Dill oil-induced increase of endogenous ROS in <i>A. flavus</i> cells. (B) Effect of antioxidant Cys on dill oil-induced ROS production in <i>A. flavus</i> cells.</p

Efficacy of different concentrations of dill oil on the glucose-dependent acidification of medium in <i>A. flavus</i>.

<p>Values are means (n = 3) ± standard deviations.</p

Efficacy of dill oil on the lesion of plasma membrane of <i>A. flavus</i> cells.

<p>(A) Sequence of density plots exhibiting <i>A. flavus</i> cell size (forward scatter, FS-Lin) analyzed by flow cytometry at PMT4 channel (620 nm), and the respective percentages of PI-stained cells (right quadrant) for a series of samples treated with increasing concentrations of dill oil. (a) autofluorescence of non-treated cells; (b) fluorescence of non-treated cells stained with 1 µg/ml PI for 30 min; (c–f) cells treated with dill oil at 0.25 µl/ml (c), 0.5 µl/ml (d), 1.0 µl/ml (e), 2.0 µl/ml (f). (B) Effect of dill oil on percentage of PI-stained <i>A. flavus</i> cells analyzed by flow cytometry for 12 h and compared with an untreated control. Significant differences (<i>p</i><0.05) between means are indicated by letters above the histogram bars. Values are means (n = 3) ± standard deviations.</p

Efficacy of dill oil on MMP of <i>A. flavus</i> cells.

<p>The measurements were performed by confocal laser scanning microscope (top panels) and flow cytometry (bottom panels) stained with Rh123. (A, a) Control, (B, b) Dill oil at 0.25 µl/ml, (C, c) Dill oil at 0.5 µl/ml, (D, d) Dill oil at 1.0 µl/ml, (E, e) Dill oil at 2.0 µl/ml.</p