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Maharishi Arvind Institute of Pharmacy, Jaipur, Rajasthan, India.
Opportunistic fungal infections caused by Candida species represent a major clinical concern due to increasing antifungal resistance to conventional synthetic drugs. This study evaluates and compares the in vitro antifungal activity of ethanolic extracts of Nigella sativa Linn. (seeds) and Catharanthus roseus Linn. (leaves) against four clinically significant Candida strains (Candida albicans, Candida glabrata, Candida tropicalis, and Candida krusei). Antimicrobial potency was determined using disk diffusion (Zone of Inhibition, mm) and broth microdilution assays (Minimum Inhibitory Concentration, MIC, ?"g/mL" ). N. sativa demonstrated significantly stronger antifungal activity across all tested strains compared to C. roseus (p<0.05). The largest zone of inhibition and lowest MIC for N. sativa were observed against C. albicans (15.2±1.4?"mm" ; "MIC"=62.5±5.3??"g/mL" ), whereas C. roseus exhibited a 12.8±1.6?"mm" zone of inhibition and an MIC of 125.0±7.2??"g/mL" . These findings indicate that N. sativa possesses higher concentrations of potent membrane-disrupting bioactive compounds—primarily thymoquinone—making it a promising candidate for plant-based antifungal drug development
Incidence rates of invasive and superficial candidiasis have risen significantly, particularly among immunocompromised individuals such as cancer chemotherapy patients, organ transplant recipients, and individuals living with HIV/AIDS. While Candida albicans remains the predominant pathogen, non-albicans species such as C. glabrata, C. tropicalis, and C. krusei are increasingly implicated in refractory bloodstream and mucosal infections. Management of these infections is complicated by drug-induced toxicities (e.g., amphotericin B-induced nephrotoxicity) and the rapid emergence of resistant strains to azoles and echinocandins.
Medicinal plants provide a rich alternative source of multi-targeted bioactive metabolites. Nigella sativa Linn. seeds contain thymoquinone, carvacrol, and essential oils with documented antimicrobial properties. Catharanthus roseus Linn. contains cytotoxic and antimicrobial indole alkaloids such as vincristine, vinblastine, and ajmalicine. This study compares the relative antifungal efficacy of standardized ethanolic extracts of N. sativa and C. roseus against four Candida strains in vitro.
2. MATERIALS AND METHODS
2.1 Plant Material Sourcing and Extract Preparation
Authentic samples of Nigella sativa seeds and Catharanthus roseus leaves were procured from verified suppliers and botanically authenticated. Dried plant materials were finely pulverized using a mechanical grinder to ensure uniform particle size.
Extractions were performed via solvent extraction using absolute ethanol:
2.2 Microorganisms and Growth Conditions
Test yeast strains—Candida albicans, Candida glabrata, Candida tropicalis, and Candida krusei—were obtained from reference culture collections (ATCC). Cultures were maintained on Sabouraud Dextrose Agar (SDA) slants at 4^? "C" . Liquid inocula were prepared in Sabouraud Dextrose Broth (SDB) and adjusted spectrophotometrically to a standardized cell density of 1×10^6 "CFU/mL" .
2.3 Disk Diffusion Assay
2.4 Minimum Inhibitory Concentration (MIC) Determination
MIC values were determined using the broth microdilution method in 96-well microtiter plates in accordance with CLSI guidelines:
2.5 Statistical Analysis
All experimental procedures were performed in triplicate (n=3). Data are expressed as "Mean"±"Standard Deviation (SD)" . Statistical differences between extract treatments were evaluated using Student's paired t-test with a significance threshold set at p<0.05.
3. RESULTS
3.1 Zone of Inhibition (Qualitative Antifungal Potency)
The disk diffusion assay demonstrated measurable antifungal activity for both plant extracts across all tested Candida strains. N. sativa extract consistently produced larger inhibition zones compared to C. roseus.
Zone of Inhibition Data Summary
|
Extract Source |
C. albicans (mm) |
C. glabrata (mm) |
C. tropicalis (mm) |
C. krusei (mm) |
|
Nigella sativa |
15.2 ± 1.4 |
13.5 ± 1.2 |
14.8 ± 1.7 |
12.7 ± 1.5 |
|
Catharanthus roseus |
12.8 ± 1.6 |
11.4 ± 1.3 |
13.0 ± 1.4 |
10.9 ± 1.6 |
|
Fluconazole Control |
17.5 ± 2.4 |
14.6 ± 1.6 |
16.9 ± 1.3 |
13.7 ± 1.2 |
N. sativa demonstrated maximum activity against C. albicans (15.2±1.4 "mm" ), while C. roseus exhibited its lowest activity against fluconazole-resistant C. krusei (10.9±1.6 "mm" ).
3.2 Minimum Inhibitory Concentration (MIC)
Quantification via broth microdilution confirmed that N. sativa exhibits significantly greater antifungal potency (lower MIC values) than C. roseus across all strains (p<0.05).
Minimum Inhibitory Concentration (MIC) Data Summary
|
Extract Source |
C. albicans (μg/mL) |
C. glabrata (μg/mL) |
C. tropicalis (μg/mL) |
C. krusei (μg/mL) |
|
Nigella sativa |
62.5 ± 5.3 |
100.0 ± 6.5 |
80.0 ± 4.9 |
120.0 ± 6.2 |
|
Catharanthus roseus |
125.0 ± 7.2 |
200.0 ± 8.1 |
150.0 ± 7.0 |
250.0 ± 9.3 |
|
Fluconazole Control |
61.2 ± 4.3 |
94.0 ± 6.5 |
76.0 ± 4.9 |
115.0 ± 6.2 |
DISCUSSION
These results demonstrate that ethanolic extracts of both Nigella sativa and Catharanthus roseus possess in vitro antifungal activity against opportunistic Candida species, with N. sativa exhibiting superior potency. The statistical significance (p<0.05) of the lower MICs and larger inhibition zones confirms that N. sativa extracts contain higher concentrations of highly active antifungal compounds.
4.1 Phytochemical Mechanisms of Action
Nigella sativa: The pronounced antifungal potency is largely driven by thymoquinone. Thymoquinone interacts with lipophilic components of the fungal plasma membrane, causing membrane destabilization, cytoplasmic content leakage, cellular lysis, and oxidative damage via reactive oxygen species (ROS) generation. Secondary flavonoids and essential oil components (p-cymene, carvacrol) may act synergistically to disrupt cell membrane integrity and suppress biofilm formation. Catharanthus roseus: Antifungal activity in C. roseus is primarily mediated by indole alkaloids (vincristine, vinblastine, ajmalicine). These compounds disrupt intracellular tubulin polymerization, halting nuclear division and protein synthesis. Because this intracellular mechanism does not directly lyse the cell membrane, it results in smaller inhibition zones and higher MIC values in acute in vitro assays.
4.2 Differential Strain Sensitivity
C. albicans displayed the highest susceptibility to both plant extracts, whereas non-albicans strains—particularly C. glabrata and C. krusei—showed reduced sensitivity. C. krusei is intrinsically resistant to fluconazole, and C. glabrata rapidly acquires azole resistance. The observed activity of N. sativa against these refractory strains ("MIC"=100.0-120.0 μ"g/mL" ) highlights its potential for treating azole-resistant mycoses.
CONCLUSION
Nigella sativa ethanolic extract exhibits superior in vitro antifungal activity compared to Catharanthus roseus across tested Candida species (C. albicans, C. glabrata, C. tropicalis, and C. krusei). The enhanced potency of N. sativa is attributed to thymoquinone-mediated cell membrane disruption. Future research should focus on bio-guided fractionation, in vivo pharmacokinetic evaluations, toxicological profiling, and testing against emerging multidrug-resistant strains such as Candida auris.
REFERENCES
Najrana, Richa Agarwal, Rajesh Asija, Comparative Evaluation of In Vitro Antifungal Activity of Nigella sativa Linn. and Catharanthus roseus Linn. Extracts Against Pathogenic Candida Strains, Int. J. of Pharm. Sci., 2026, Vol 4, Issue 10, 1622-1626, https://doi.org/10.5281/zenodo.23332347
10.5281/zenodo.23332347