Ex vivo nail infection as an effective preclinical method for screening of new topical antifungals.

Onychomycosis are fungal nail infections comprising of about 50% of onychopathies and are commonly caused by dermatophytes. The treatment of this dermatomycosis requires a long period of time and is associated with high rates of recurrence. In view of the need to evaluate the antifungal performance of promising preclinical compounds, we developed, in this study, a practical and accessibleex vivo model for establishing a Trichophyton rubrum onychomycosis framework using porcine hooves. This model has as its main advantage the similar structural and three-dimensional characteristics that the porcine hooves have with the human nail. The proposed model allowed to evaluate the antifungal activity of a new antifungal compound and a reference drug (terbinafine), both already incorporated into a nail lacquer for topical use. Treatments with compound 3-selenocyanate-indole (Se4a) and with terbinafine incorporated into this nail lacquer completely inhibited fungal growth, corresponding to the profile of in vitro activity observed against T. rubrum. This study concludes that the ex vivo porcine hoof model is an effective alternative method for preclinical screening of drugs or new topical compounds developed to combat onychomycosis. Further studies are needed to compare the permeability of porcine hooves with human nails permeability.

Preparation, characterization and antidermatophytic activity of free- and microencapsulated cinnamon essential oil.

Essential oils (EO) are effective natural antimicrobials but are susceptible to oxidation. Microencapsulation improves EO stability, reduces toxicity, and controls release. The aim of this study was preparation, characterization and antidermatophytic activity of free and microencapsulated cinnamon essential oil (MP). MP were prepared by the spray drying method and the success of MP encapsulation was confirmed by UV-vis spectroscopy, dynamic light scattering (DLS), scanning electron microscopy (SEM), differential scanning calorimetry (DSC) and Fourier transform infrared (FT-IR) spectroscopy. The antifungal effect of EO and MP was evaluated by the broth microdilution method against Microsporum gypseum and Trichophyton mentagrophytes. The checkerboard method was used to assess synergistic interactions. Fluorescence microscopy and scanning electron microscopy were used to investigate the inhibition of hyphal growth by EO and MP. A cytotoxic assay was performed using the VERO cell line. Microencapsulated cinnamon essential oil was found to be micrometric, with a round, regular structure. The minimum inhibitory concentration of EO was found to be between 125-250µg/mL, while that of MP was 220.5-440.5µg/mL. EO was synergistic with fluconazole while microencapsulated oil was less cytotoxic than EO.

Antioxidant and antifungal activities of marrubiin, extracts and essential oil from Marrubium vulgare L. against pathogenic dermatophyte strains.

OBJECTIVE: Medicinal plants extracts and plant-derived compounds are one of the natural sources for discovering new antifungal agents, the objectives of this work were to investigate for the first time the antidermatophytic, antipathogenic activities of methanol, acetone extracts, and essential oil of Marrubium vulgare L. grown in Tunisia and its active compound marrubiin on pathogenic for animals and humans, such as some dermatophytes and pathogenic for plants, and to evaluate antioxidant activities of different extracts with consideration to their chemical compositions. MATERIAL AND METHODS: Acetone and methanol extracts were evaluated by HPLC, the essential oil was also analyzed by GC/MS. PCL assay was used to determine the antioxidant activity. RESULTS: Results showed that methanol and acetone extracts exhibited a significant antioxidant activity (261.41 and 272.90µmol TE/g respectively), while the lowest one was observed in the case of marrubiin and essential oil. The antifungal activity of different extracts, marrubiin and essential oil at two concentrations (20 and 100µg/mL) were screened against the dermatophytes fungi Microsporum gypseum, Microsporum canis, Arthroderma cajetani, Trichophyton mentagrophytes, Trichophyton tonsurans, Epidermophyton floccosum and against two fungi strains (Botrytis cinerea, Pythium ultimum). Among tested extracts, marrubiin at 100µg/mL showed about 50% inhibition for T. mentagrophytes and E. floccosum. The anti-phytopathogenic activity was also carried out, only marrubiin had in activity against B. cinerea at the highest dose (32.40%), while methanol extract of M.vulgare and marrubiin are able to increase the mycelial growth of P. ultimum at the highest concentration (45.15 and 40.30% respectively). CONCLUSION: In our study, we conclude that M.vulgare and marrubiin can be used as natural antioxidants and antifungal agent for treatment of skin dermatophyte infections.

Biomass-Derived Dialdehyde Cellulose Cross-linked Chitosan-Based Nanocomposite Hydrogel with Phytosynthesized Zinc Oxide Nanoparticles for Enhanced Curcumin Delivery and Bioactivity.

A sustainable biomass-based nanocomposite hydrogel was formulated, characterized, and applied for curcumin delivery. Phytosynthesized zinc oxide nanoparticles (ZnO NPs) employing musk melon (Cucumis melo) seed extract was embedded in the hydrogel matrices and cross-linked using Dialdehyde cellulose prepared from sugarcane (Saccharum officinarum) bagasse (SCB). Nanoparticle incorporation enhanced the hydrogel's swelling degree to 4048% at pH 4.0. Also, an improved tensile strength of 14.1 ± 0.32 MPa was exhibited by the nanocomposite hydrogel compared to 9.79 ± 0.76 MPa for the pure chitosan cellulose hydrogel. A curcumin loading efficiency of 89.68% with around 30% increased loading was exhibited for the nanocomposite hydrogel. A Fickian diffusion-controlled curcumin release mechanism with maximum release at pH 7.4 was obtained. The synergistic effect on the antimicrobial activity was exhibited against Staphylococcus aureus and Trichophyton rubrum. The in vitro cytotoxicity studies employing L929 cells and A431 cells demonstrated good biocompatibility and enhanced anticancer activity of the curcumin-loaded green nanocomposite hydrogel compared to pure curcumin.

In vitro Antifungal Activity of Japanese Folk Herb Extracts against Trichophyton rubrum.

Folk herbs are vital to the Japanese health care system, with some being used to treat fungal skin infections. We tested folk herbs used in traditional Japanese medicine for in vitro antifungal activity against Trichophyton rubrum, the most common pathogen in patients with superficial dermatitis. Acetone, ethanol, methanol, water, and acetic acid extracts from 15 species of Japanese folk herbs (Bi-wa, Aka-me-gashiwa, Ta-mushi-ba, Yomogi, Shi-so, Ku-ko, So-ba, Yama-momo, Kuro-mo-ji, Ichi-ji-ku, Kaki-no-ki, Kuwa-no-ki, Kusa-gi, Chimaki-zasa, and I-bukijya-kou-sou) were evaluated for fungal growth inhibition, as measured by absorbance. Crude acetic acid extracts of Aka-me-gashiwa, Ta-mushi-ba, Yomogi, Ku-ko, Kaki-no-ki, and I-bukijya- kou-sou exhibited antifungal activity comparable to that of amphotericin B. Although the active compounds and underlying antifungal mechanisms are not known, the data contribute to the development of new and safe antifungal drugs, highlight new extraction methods for herbs, and support the traditional use of Japanese folk herbs against fungal infections.

Antimicrobial and cytotoxic effects of the Copaifera reticulata oleoresin and its main diterpene acids.

ETHNOPHARMACOLOGICAL RELEVANCE: The oleoresin of Brazilian Copaifera reticulata is a traditional remedy used for the treatment of skin and urinary tract infections, respiratory diseases, rheumatism, ulcer and tumours; thus, playing an important role in the primary health care of the indigenous population. AIM: As most previous pharmacological tests used the crude oleoresin and only a few studies so far dealt with enriched fractions or pure chemically defined compounds, the aim of this study was to systematically evaluate the antimicrobial and cytotoxic properties of the Copaifera reticulata oleoresin and to assign traditional uses to specific secondary metabolites. MATERIALS AND METHODS: The oleoresin, as well as its neutral and acidic fractions were tested for their activity against six cancer cell lines, two clinically relevant bacterial strains, and two dermatophytes. Both fractions were analysed by GC-MS and UHPLC-ELSD, respectively. The antibacterial acidic phase was further fractionated by preparative chromatography to purify and characterize the compounds responsible for the observed pharmacological effect. RESULTS: Whereas no cytotoxic activity was detected, the crude oleoresin and its acidic fraction showed antibacterial activity against gram-positive bacteria Enterococcus faecium (IC50 values 4.2 and 4.8 µg/mL, respectively) and methicillin-resistant Staphylococcus aureus (MRSA, IC50 values 5.3 and 7.2 µg/mL, respectively). Purification of the acidic fraction of the C. reticulata oleoresin yielded two dicarboxylic diterpene acids and the four main diterpene acids, comprising three different diterpene scaffolds. Interestingly, the activity was not restricted to a particular diterpene-type but rather depended on the compounds' lipophilicity, with the most active constituent showing IC50 values of 1.6 (E. faecium) and 2.5 µg/mL (MRSA), respectively. Furthermore, ent-polyalthic acid, the major diterpenoid, was significantly active against dermatophytes with IC50 values of 6.8 µg/mL (Trichophyton rubrum) and 4.3 µg/mL against (T. mentagrophytes). CONCLUSION: The present study proved the antimicrobial effects of the C. reticulata oleoresin and its diterpenoid constituents, confirming its wide use in folk medicine for the treatment of skin and urinary tract infections. The inhibitory activity of copaiba diterpenoids against dermatophytic fungi as well as the gram-positive bacteria E. faecium and MRSA is being reported for the first time, providing potential lead structures for the treatment of these clinically relevant bacterial strains.

Antimicrobial activity of Elytropappus rhinocerotis (Asteraceae) against micro-organisms associated with foot odour and skin ailments.

ETHNOPHARMACOLOGICAL RELEVANCE: The twigs of Elytropappus rhinocerotis are widely used in the Cape region of South Africa to treat foot odour, perspiration and also itchy, chilblained and burning feet. However, no antimicrobial studies have hitherto been published on this popular Cape herbal medicine, which is also used for a wide range of ailments. AIMS OF THE STUDY: To determine the antimicrobial activity of the extracts, essential oil and two major labdane diterpenes isolated from E. rhinocerotis against micro-organisms associated with foot odour and other conditions associated with skin infections. MATERIALS AND METHODS: Leafy stems were harvested from three individual plants at three separate geographical localities, giving a total of nine plant samples. The samples were air-dried, powdered and extracted with a 1:1 mixture of methanol and dichloromethane, and also with sterile distilled water. A portion of each sample was also hydrodistilled to obtain nine samples of essential oil. Isolation of the major labdane diterpenes was performed using silica and ethyl acetate in hexane (3:7 v/v) as the mobile phase. Minimum inhibitory concentrations (MIC) were determined for nine crude extracts, as well as three essential oil samples and two labdane diterpenes obtained from a bulk sample. The test organisms used in this study were from Deutsche Sammlung von Mikrooganismen (DSM) and American type culture collection (ATCC) strains and included five bacterial species (Brevibacterium agri ATCC 51663, B. epidermidis DSM 20660, B. linens DSM 20425, Staphylococcus aureus ATCC 25923, S. epidermidis ATCC 12228) and one fungal species (Trichophyton mentagrophytes ATCC 9533) associated with the skin. RESULTS: The presence of essential oil is reported for the first time. Organic extracts and essential oil samples showed moderate to noteworthy activity against selected test organisms. For the Brevibacteria, the lowest MIC values for phenolic extracts were several times lower than the corresponding values for the positive control zinc (shown in brackets): 0.0031 mg/mL against Brevibacterium agri (0.33 mg/mL), 0.17 mg/mL against B. epidermidis (3.91 mg/mL) - both for sample 2 of Vanwyksdorp; 0.13 mg/mL against B. linens (2.28 mg/mL) - for sample 3 of Vanwyksdorp. Two isolated labdane diterpenoids (one of which is here first reported) were also antimicrobially tested and showed moderate activity but had high abundance in the extracts. Two major monoterpenes and four sesquiterpenes in the essential oil were identified as 1,8-cineole (4.6-12.3%), terpinen-4-ol (9.2-24.3%), germacrene A (3.9-15.6%), (-)-spathulenol (1.7-37.8%), viridiflorol (0.3-100%), and silphiperfol-6-en-5-one (4.5-26.8%). CONCLUSION: The antimicrobial results particularly for the essential oils and against the Brevibacteria support the traditional topical use of Elytropappus rhinocerotis twigs to treat foot perspiration, foot odour and other related skin conditions.

Analysis of chemical composition and in vitro antidermatophyte activity of ethanol extracts of Dryopteris fragrans (L.) Schott.

ETHNOPHARMACOLOGICAL RELEVANCE: Dryopteris fragrans (L.) Schott is a deciduous perennial herb, which has been used traditionally for treatment of ringworm infections and others skin diseases in the north of China. AIM OF THE STUDY: To characterize the chemical composition, evaluate the antifungal activity and explore possible mechanisms about action of ethanol extracts of D. fragrans. MATERIALS AND METHODS: The chemical components in the ethanol extracts of D. fragrans were detected by high-performance liquid chromatography coupled with electrospray ionization and quadruple time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS/MS). The minimal inhibitory concentrations (MIC) and minimal fungicidal concentrations (MFC) of the ethanol extracts of D. fragrans were determined by the Clinical and Laboratory Standards Institute (CLSI) M38-A2 method against 62 isolates of dermatophytes. The kinetics of fungal kill, synergy testing by checkerboard dilution and quantitation of sterol by ultra-performance liquid chromatography (UPLC) on Trichophyton rubrum and Trichophyton mentagrophytes were also investigated. RESULTS: Fourteen derivatives of phloroglucinol were identified in the ethanol extracts of D. fragrans. The MIC of the ethanol extracts of D. fragrans ranged from 0.059 to 3.780 mg/mL while MFC ranged from 0.118 to 3.780 mg/mL. The ethanol extracts of D. fragrans exerted fungicidal activity after 12 h of incubation against Trichophyton rubrum while it required 36 h of incubation against Trichophyton mentagrophytes at concentrations of 8 × MIC. In synergy testing, the interaction between miconazole (MCZ) and terbinafine (TBF) with the ethanol extracts of D. fragrans proved to be indifferent by testing fractional inhibitory concentration (FIC) values. Sterol in samples of fungal cells treated with the ethanol extracts of D. fragrans was significantly reduced. CONCLUSIONS: The ethanol extracts of D. fragrans had antifungal and fungicidal activity against dermatophytes and was likely a strain-dependent fungicidal agent. Interaction between drugs was indifferent on tested isolates. The inhibition of ergosterol biosynthesis was one of the antifungal mechanisms of the ethanol extracts of D. fragrans. These results showed that the ethanol extracts of D. fragrans could be explored for promising antifungal drugs. Dozens of phloroglucinol derivatives may contribute to high antifungal activity of the ethanol extracts of D. fragrans.

Determining the optimal parameters of 420-nm intense pulsed light on Trichophyton rubrum growth in vitro.

The effect of and the optimal parameters for intense pulsed light (IPL) with a 420-nm filter on an isolate of the fungus Trichophyton rubrum (T. rubrum) were examined in vitro. Colonies of T. rubrum were irradiated by using 420-nm IPL with various pulse numbers and energies. Colony areas were photographed and compared with those of untreated colonies to assess growth inhibition. Statistically significant inhibition of T. rubrum growth was detected in colonies treated with 12 pulses of greater than or equal to 12 J/cm2. The optimal parameters of 420-nm IPL were 12 pulses of 12 J/cm2. However, more in vitro and in vivo studies are necessary to investigate and explore this mechanism to determine whether IPL would have a potential use in the treatment of fungal infections of the skin.

Activity of Salvia dolomitica and Salvia somalensis Essential Oils against Bacteria, Molds and Yeasts.

Essential oils (EOs) from Salvia dolomitica and Salvia somalensis, widely employed in the cosmetic and perfume industry, were analyzed for composition and tested against bacterial and fungal pathogens isolated from clinical and environmental specimens. The analyses were carried out against Staphylococcus aureus, Staphylococcus pseudointermedius, Pseudomonas aeruginosa, Escherichia coli, Streptococcus canis, Streptococcus pyogenes, Klebsiella pneumoniae, Proteus mirabilis, Microsporum canis, Microsporum gypseum, Trichophyton mentagrophytes, Aspergillus niger, Aspergillus flavus, Candida albicans, Candida krusei, Mucor sp. and Trichothecium roseum. Both EOs showed similar percentages of total monoterpenes and sesquiterpene hydrocarbons. The main constituents were 1,8-cineole and ß-caryophyllene in S. dolomitica and bornyl acetate and camphor in S. somalensis. The selected EOs have no relevant antifungal or antibacterial activities if compared to conventional drugs.

Susceptibility of Trichopyton mentagrophytes to Visible Light Wavelengths.

OBJECTIVE: To determine whether a blue light (405 nm) could inhibit the growth of Trichopyton mentagrophytes without using a photosensitizing material as part of the treatment protocol. DESIGN: Basic physiologic randomized trial using laboratory specimens (T mentagrophytes). INTERVENTIONS/METHODS: Plated on a growth medium, T mentagrophytes were exposed to 3 to 5 administrations of blue light at 20 J/cm over 28 hours. Following 7 days of incubation, colony-forming units were counted and compared with nonirradiated controls. RESULTS: The study found 3, 4, and 5 administrations of blue light produced significant inhibition of T mentagrophytes (P < .05); 4 and 5 applications produced the greatest inhibition of growth (84.7% and 93.6% kill rates, respectively). CONCLUSIONS: The application of 405-nm light at a dose of 20 J/cm is an effective in vitro inhibitor of T mentagrophytes. To give results similar to those seen when a photosensitizing material is included, 3 to 5 applications of this wavelength and dose condition delivered over 28 hours is likely needed.

Investigation of the antifungal potential of linalool against clinical isolates of fluconazole resistant Trichophyton rubrum.

The aim of this study was to investigate the activity of the monoterpene linalool against clinical isolates of Trichophyton rubrum. Initially, a sensitivity assay for commercial antifungals with solid disks in diffusion medium was performed. Minimum inhibitory concentration (MIC) of linalool and ketoconazole (positive control) were determined by microdilution in RPMI 1640 medium (CLSI M38-A2). We then evaluated the action of linalool and ketoconazole at different concentrations (1/2MIC, MIC and 2×MIC) on mycelial growth (radial mycelial growth), conidia production and conidia germination using a hemacytometer. The effects on cell membrane (release of intracellular material) were also investigated. Finally, changes in fungal morphology as induced by the test drugs were analyzed. Based on the sensitivity tests, the fungal strains showed resistance to 5-fluorocytosine and fluconazole. The linalool MIC values ranged from 256µg/mL to 512µg/mL, whereas ketoconazole showed values of 4µg/mL to 8µg/mL. For the LM 305 strain, the test drugs showed the following MIC values: linalool 256µg/mL and ketoconazole 8µg/mL. The mycelial growth of T. rubrum LM 305 was inhibited by linalool (2×MIC) and ketoconazole (1/2MIC, MIC, 2×MIC), in 7 days of treatment (P<0.05). The test-drugs also inhibited conidial germination and conidiogenesis (P<0.05). Linalool also caused leakage of intracellular material (P<0.05). Finally, we verified the effectiveness of linalool and ketoconazole to induce micro-morphological changes, forming abnormal, wide, short and crooked hyphae. Based on these results, we conclude that linalool presents as an antifungal agent with anti-Trichophyton rubrum potential, an important dermatophytosis agent.

Antifungal activity of acetone extracts from Punica granatum L., Quercus suber L. and Vicia faba L.

Human and animal mycoses become more frequent and more resistant to traditional treatments. In this work, we tested the in vitro antifungal activity of acetonic extracts of Punica granatum L., Quercus suber L. and Vicia faba L. against seven pathogen fungi and the in vivo antifungal activity against Candida albicans and Trichophyton mentagrophytes. The phytochemical screening was also carried out and showed that the extracts contained mainly proanthocyanidins. Other polyphenols were also present but in low quantity. The acetone extract of V. faba L. gave a good in vitro inhibition of yeasts and was the most active for treating candidiasis in mice. It decreased the percentage of mortality with only 20µg. But the in vivo antifungal activity of this extract on T. mentagrophytes was low. It only showed a small diminution of crusting and erythema after the administration of 100µg. On the contrary, the acetone extracts of P. granatum L. had a poor activity against yeasts and a better one against moulds. It gave the best in vivo antifungal activity against T. mentagrophytes by healing animals with 40µg. The extract of P. granatum L. gave also an interesting in vivo antifungal activity against T. mentagrophytes with an active dose of 80µg.

Onychomycosis in diabetic patients in Fako Division of Cameroon: prevalence, causative agents, associated factors and antifungal sensitivity patterns.

BACKGROUND: Onychomycosis is an infection of the nail unit by a fungus. This is a very common infection amongst diabetics. Its occurrence among diabetics in Fako division is unknown. In this study we provide information on the characteristics of onychomycosis in diabetics in Fako division, Cameroon. METHODS: A cross-sectional descriptive and analytical hospital-based study was conducted in two diabetic clinics in the Buea and Limbe regional hospitals. We recruited 152 consenting diabetics into the study. Demographic, behavioural, and clinical data of patients were obtained through the use of structured questionnaires. Toenail, finger nail, skin scrapings and nail clippings were collected from participants, KOH mounts were prepared and observed under the microscope and cultured on Sabouraud Dextrose Agar supplemented with chloramphenicol to isolate causative fungi. Identification of isolates was done to species level using the cello tape flag method and slide culture. The presence of a dermatophyte by either microscopy or culture or both methods was considered positive for onychomycosis. Antifungal susceptibility testing was carried out using selected antifungals by the Kirby-Bauer disk diffusion method on Sabouraud Dextrose Agar. RESULTS: Clinical onychomycosis was found in 77 of the 152 diabetics tested giving a prevalence of 50.7% (95% CI 42.4-58.9) in diabetics in Fako. No socio-demographic or clinical factor studied was significantly associated with onychomycosis. Trichophyton rubrum was the most common isolate (62%). Other isolates included Trichophyton metagraphyte (22%) and Trichophyton tonsurans (16%). Dermatophytes were sensitive to miconazole (66%), amphotericin B (19%) and ketoconazole (14%). CONCLUSION: Onychomycosis is common in diabetics in Fako signifying the need for regular screening by either microscopy or culture. Infected nails could be treated with miconazole.

In Vitro Antifungal Activity of Kampo Medicine Water Extracts against Trichophyton rubrum.

Kampo medicines consist of a variety of crude animal, plant, and mineral extracts that have long been used to relieve different symptoms, and are relatively safe. However, their mechanisms of actions have not been well investigated. We screened 61 commercially available Kampo medicines to determine if they contain constituents with antifungal activity against Trichophyton rubrum. The antifungal effect of the Kampo medicines was determined by measuring the mean absorbance of treated fungal culture media. Lower absorbance values suggested a higher inhibition of the growth rate of T. rubrum by the Kampo medicines. We found that seven of the evaluated formulations exhibited a comparable antifungal activity to that of fluconazole at 14 mg/mL. The seven active Kampo medicines were Saiko-keishi-kankyou-to, Saiko-ka-ryukotsu-borei-to, Saiko-keishi-to, Keishi-ka-ryukotsu-borei-to, Dai-saiko-to, Bohu-tsu-sho-san, and Otsu-ji-to. The seven Kampo medicines with antifungal activity contain 30 different crude extracts, and Ou-gon (Scutellaria root) is a supplement contained in six of the seven formulations. Therefore, Ou-gon was considered to play a major role in their antifungal effect. The antifungal assay of the Ou-gon water extract showed that it significantly inhibited the growth of T. rubrum at a concentration of 20 mg/mL. Future studies will focus on the isolation and identification of the antifungal components of the crude extracts of Ou-gon, which may be potentially useful, new, and safe antifungal drugs.

Discovery of antifungal constituents from the Miao medicinal plant Isodon flavidus.

ETHNOPHARMACOLOGICAL RELEVANCE: Leigong Mountain is an area in the Southwest of China where there is a high incidence rate of athlete's foot, but the Miao people, a Chinese minority who reside in this mountainous area have suffered less from this disease due to their use of the herbal medicine Isodon flavidus (Hand.-Mazz.) H. Hara. AIM OF THE STUDY: The present study is to identify the active chemical constituents responsible for antifungal effects of the folk medicine plant. MATERIALS AND METHODS: The natural compounds were separated from the methanol extract of the twigs and leaves of I. flavidus by phytochemical study using chromatographic methods, and their chemical structures were determined by analysis of the spectroscopic data including 1D and 2D NMR spectra. The absolute configuration of fladin A (1) was further confirmed by X-ray crystallographic analysis. The compounds were evaluated for their antifungal activity against the athlete's foot fungus Trichophyton rubrum. They were further evaluated for their antimicrobial and anti-biofilm activity against the dental pathogens Streptococcus mutans, Porphyromonas gingivalis and Candida albicans. RESULTS: Phytochemical and biological studies of I. flavidus led to the discovery of two antifungal compounds, fladin A (1) and lophanic acid (2). Fladin A (1) is a novel diterpene with an unprecedented cyclic ether group formed between C-4 and C-9. Lophanic acid (2) displayed inhibition activity against the athlete's foot fungus Trichophyton rubrum with an MIC value of 7.8µg/mL, and fladin A (1) also showed inhibition activity against the fungus with a MIC value of 62.5µg/mL. CONCLUSIONS: Our identification of two antifungal compounds provided strong evidence for the Miao people to use I. flavidus as a medicinal plant for treatment of athlete's foot disease. The very different chemical structures of the active compounds from those in the market presents them as potential antifungal lead compounds for follow-up study.

Comparative Sensitivity of Trichophyton and Aspergillus Conidia to Inactivation by Violet-Blue Light Exposure.

OBJECTIVE: To investigate the use of 405 nm light for inhibiting the growth of selected species of dermatophytic and saprophytic fungi. BACKGROUND DATA: The increasing incidence and resilience of dermatophytic fungal infections is a major issue, and alternative treatment methods are being sought. METHODS: The sensitivity of the dermatophytic fungi Trichophyton rubrum and Trichophyton mentagrophytes to 405 nm violet-blue light exposure was investigated, and the results compared with those obtained with the saprophytic fungus Aspergillus niger. Microconidia of T. rubrum and T. mentagrophytes and conidia of A. niger were seeded onto Sabauroud dextrose agar plates and irradiated with 405 nm light from an indium-gallium-nitride 99-DIE light-emitting diode (LED) array and the extent of inhibition was measured. RESULTS: Germination of the microconidia of the Trichophyton species was completely inhibited using an irradiance of 35 mW/cm(2) for 4 h (dose of 504 J/cm(2)). A. niger conidia showed greater resistance, and colonial growth developed after light exposure. In liquid suspension tests, 405 nm light dose levels of 360, 720, and 1440 J/cm(2) resulted in complete inactivation of T. rubrum microconidia, whereas A. niger showed greater resistance, and at the highest dose level applied (1440 J/cm(2)) although A niger hyphae were completely inactivated, only a 3-log10 reduction of a 5-log10 conidial suspension was achieved. CONCLUSIONS: The study results demonstrate the relatively high sensitivity of Trichophyton microconidia to 405 nm violet-blue light, and this is may be of potential interest regarding the control and treatment of dermatophyte infections.

CREATION OF OINTMENT COMPOSITIONS CONTAINING PHENOL COMPOUNDS FOR MEDICAL TREATMENT.

The research was aimed at studying the possibility of obtaining drug ointments with a potential anti-mycotic activity by using thick extracts obtained from the leaves of fustic, hazel, nut and bark of the oak. There were prepared the ointment compositions on different bases. As a methodology for studying the properties of the obtained ointments, there have been used the studies of colloidal stability and resorption of tanning substances in agar. The obtained results allow for making conclusion that the selected ointments are colloidally stable, and the values of movement of zones of biologically active substances in the agar body are large enough. These data allow for forecasting both the possibility of creating the similar ointment systems and their rather high properties. Based on the obtained data, there have been selected the ointment composition variants for further studies.

Antifungal activity of 40 TCMs used individually and in combination for treatment of superficial fungal infections.

ETHNOPHARMACOLOGICAL RELEVANCE: A series of 40 important Traditional Chinese Medicines (TCMs), which were reported effective in treating superficial fungal infections of the skin in Chinese clinical trial publications and Chinese Herbal Classics, were chosen for the investigation of the individual and combination antifungal properties against 8 superficial fungal strains in vitro. MATERIALS AND METHODS: Plant preparations were followed the theory of TCM by using sterile water boiled with plant material at 100°C to produce water decoction of the tested sample. The minimum inhibitory concentration (MIC) of each plant for each fungus was determined. For the compatibility investigation, both invariable (same amounts of each tested TCM) and variable (different amounts of each tested TCM) combinations were evaluated. RESULTS: All the tested TCMs demonstrated varying degrees of antifungal activities against one or more of the tested superficial fungi, and 16 of which were effective on all of the fungi. Strong antifungal activities were exhibited by water decoction of 7 TCMs with MIC at about 100µg/ml, and among these effective antifungal extracts, 4 TCMs including Melaphis chinensis, Polygonum cuspidatum, Punica granatum and Schisandra chinensis showed the significantly inhibitory activities against all of the fungi with MICs among 50µg/ml. Most of the invariable combinations of the above-mentioned 4 TCMs showed synergic effects against 4 of the least susceptible fungi strains, especially the invariable combination of Punica granatum, Melaphis chinensis and Schisandra chinensis, with the MIC at 23.4µg/ml. However, their further variable combinations investigation demonstrated that only the combination of 7.5g Punica granatum with 10g Melaphis chinensis and 7.5g Schisandra chinensis showed synergic effect with the MIC at19.5µg/ml. CONCLUSIONS: The present study aimed the discovery of therapeutically useful agents for treatment of superficial fungal infections. Findings suggested that the combination of 3 TCMs including Punica granatum, Melaphis chinensis and Schisandra chinensis showed potential antifungal activity and thus appeared to be promising agents in preventing superficial fungal skin infectious in a natural way through herbal resources. The synergic effects of invariable and variable combinations of the tested TCMs threw a light on our further animal model and clinical practice as well as the bio-guided isolation and identification of the antifungal compounds.

The anti-dermatophyte activity of Allium hirtifolium Boiss aqueous extract.

In an attempt at demonstrating the efficacy of Allium hirtifolium aqueous extract in control of skin fungal infections as traditional use, we evaluated the anti-dermatophyte activities of A. hirtifolium aqueous extract from bulbs and of ketoconazole against Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, M. gypseum, Trichophyton schoenleinii and Trichophyton verrucosum var. album by food poisoning technique, disc diffusion and micro broth dilution assays. The anti-fungal activity of A. hirtifolium was excellent when it was compared with ketoconazole. The anti-fungal evaluation by food poisoning method showed that A. hirtifolium extract inhibited the growth of dermatophytes dose-dependently. The inhibition zone diameter (IZ) of A. hirtifolium extract (15 µg/disc) was in the range of 28.8 ± 0.31 to 67.7 ± 1.5mm, while ketoconazole (15 µg/disc) had the IZ lower than 13mm. The MIC and MFC values of A. hirtifolium extract were in the range of 0.2-1.7 and 0.4-0.7 µg/mL; respectively. Therefore, A. hirtifolium extract showed a strong anti-fungal activity against human and animal dermatophytes.