Can Phlorotannins Purified Extracts Constitute a Novel Pharmacological Alternative for Microbial Infections with Associated Inflammatory Conditions?

Bacterial and fungal infections and the emerging multidrug resistance are driving interest in fighting these microorganisms with natural products, which have generally been considered complementary to pharmacological therapies. Phlorotannins are polyphenols restricted to brown seaweeds, recognized for their biological capacity. This study represents the first research on the antibacterial, antifungal, anti-inflammatory and antioxidant activity of phlorotannins purified extracts, which were obtained from ten dominant brown seaweeds of the occidental Portuguese coast.Phlorotannins content was determined by the specific dimethoxybenzaldehyde (DMBA) method and a yield between 75 and 969 mg/Kg phloroglucinol units (dry matter) was obtained. Fucus spiralis ranked first, followed by three Cystoseira species. The anti-inflammatory potential of the purified extracts was assessed via inhibitory effect on nitric oxide (NO) production by lipopolysaccharide-stimulated RAW 264.7 macrophage cells, Cystoseira tamariscifolia being the one showing promising activity for the treatment of inflammation. NO scavenging ability was also addressed in cell free systems, F. spiralis being the species with highest capacity. The antimicrobial potential of the extracts was checked against five Gram-positive and four Gram-negative bacteria and three fungi strains, that commonly colonize skin and mucosa and are responsible for food contamination. The different extracts were more effective against Gram-positive bacteria, Staphylococcus epidermidis being the most susceptible species. Concerning antifungal activity, Trichophyton rubrum was the most sensitive species.Although the molecular mechanisms underlying these properties remain poorly understood, the results obtained turn phlorotannins purified extracts a novel and potent pharmacological alternative for the treatment of a wide range of microbial infections, which usually also present an inflammatory component. In addition to the biological properties demonstrated herein, phlorotannins extracts may also be preferred, in order to avoid side effects and allergic reactions commonly associated with synthetic drugs.     

UPLC-MS profiling of low molecular weight phlorotannin polymers in Ascophyllum nodosum,Pelvetia canaliculata and Fucus spiralis

Phlorotannins are a group of complex polymers, found in particular brown macroalgae, composed solely of the monomer phloroglucinol (1,3,5-trihydroxybenzene). Their structural complexity arises from the number of possible linkage positions between each monomer unit. This study aimed to profile the phlorotannin metabolite composition and the complexity of isomerisation present in brown macroalgae Ascophyllum nodosum, Pelvetia canaliculata and Fucus spiralis using UPLC-MS utilising a tandem quadrupole mass spectrometer. Phlorotannin-enriched fractions from water and aqueous ethanol extracts were analysed by UPLC-MS performed in multiple reaction monitoring mode to detect molecular ions consistent with the molecular weights of phlorotannins. Ascophyllum nodosum and P. canaliculata appeared to contain predominantly larger phlorotannins (degree of polymerisation (DP) of 6–13 monomers) compared to F. spiralis (DP of 4–6 monomers). This is the first report observing the complex chromatographic separation and metabolomic profiling of low molecular weight phlorotannins consisting of more than ten monomers. Extracted ion chromatograms, for each of the MRM transitions, for each species were analysed to profile the level of isomerisation for specific molecular weights of phlorotannins between 3 and 16 monomers. The level of phlorotannin isomerisation within the extracts of the individual macroalgal species differed to some degree, resulting in substantially different numbers of phlorotannin isomers for particular molecular weights. A similar UPLC-MS/MS separation procedure, as outlined in this study, may be used in the future as a means of screening the metabolite profile of macroalgal extracts, therefore, allowing extract consistency to be monitored for standardisation purposes.     

Antifungal Properties of Cranberry Juice

Cranberry juice exerts a significant in vitro antifungal effect on eight representative species of dermatophytes, whereas it has no apparent effect on Candida albicans. The antifungal effect is fungistatic. Benzoic acid or other small molecular weight components, or both, were responsible for the fungistatic action. Studies with C. albicans on the effect of pH alone and the effect of pH on the ionization of benzoic acid indicate that cranberry juice would exert an even more significant antifungal action if the pH were left at its native value of 2.8; not adjusted to 5.6. This would probably be due to pH and a larger amount of free benzoic acid. Further investigation suggested that benzoic acid loses some of its antifungal properties in cranberry juice at pH 5.6. This investigation suggests that the dermatophytes may have a higher sensitivity to benzoic acid or other small molecular weight components of cranberry juice, or to both.     

The Phenolic Contents and Antimicrobial Activities of Some Marine Algae from the Mediterranean Sea (Algeria)

In this study, three marine algae collected from western coast of algerian mediteranean sea (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) were tested using the agar-well diffusion method for their production of antibacterial and antifungal agents on various organisms that cause diseases of humans and plants (Eschirichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Salmonella sp, Candida albicans, and Penicillium sp.). The total phenol content and antimicrobial activity were determined using different crude seaweeds extracts (methanol, diethylether, and chloroform). The results show that the chloroform extracts of (Ulva lactuca and Corallina elongata) had the highest activity against E. coli and Salmonella sp. The methanol extract obtained from (Ulva lactuca, Dictyota dichotoma, and Corallina elongata) showed antifungal activity for Candida albicans. The results of the study revealed that the seaweeds from Algeria appear to have immense potential as a source of antibacterial and antifungal compounds; they can be used in treating diseases caused by these organisms.     

Effect of phlorotannin-rich extracts of Ascophyllum nodosum and Himanthalia elongata (Phaeophyceae) on cellular oxidative markers in human HepG2 cells

Phlorotannins have received much attention due to their ecophysiological importance and potential applications in the biotechnology and food industries. Antioxidant activity studies in seaweeds have mainly focused on in vitro assays; however, there is a paucity of data regarding the effect of brown algal phlorotannins on living cultured cells. The aim of the present study was to investigate both direct and protective effects of phlorotannin-rich extracts on cell viability and the cellular oxidative status of cultured liver cells HepG2 against oxidative stress induced by tert-butyl hydroperoxide (t-BOOH). Extracts of the Phaeophyceae Ascophyllum nodosum (Fucaceae) and Himanthalia elongata (Himanthaliaceae) were submitted to gastrointestinal digestion prior to incubation for 20 h in a HepG2 culture at physiological concentrations (0.5–50 μg mL^?1). Various markers of cellular oxidative stress were then assessed, such as the generation of reactive oxygen species (ROS), antioxidant defences (concentration of reduced glutathione and activities of glutathione peroxidase, reductase and glutathione-S-transferase) and the levels of malondialdehyde as a marker for lipid peroxidation. The direct effect on cellular markers was assessed immediately after the incubation period, whereas for the protective effect, the incubation period was followed by a 3-h treatment with t-BOOH. The results indicated no effect on cell viability, and both extracts showed reduced levels of ROS and increased antioxidant defences in the direct treatment. Moreover, the extracts showed a significant protective effect against chemically induced oxidative stress in HepG2 cells by reducing ROS generation and enhancing antioxidant defences, hence supporting the utility of including brown algal extracts in functional food products.     

Solanum melongena: A potential source of antifungal agent

The antifungal activity of Solanum melongena leaf, extracted with petroleum ether, chloroform, methanol and water was evaluated against three human pathogenic dermatophytes namely Trichophyton mentagrophytes, T. rubrum and T. tonsurans and two opportunistic fungi Candida albicans and Trichosporon beigelii. Maximum yield of plant components was 4.32 g, extracted in water and minimum 1.07 g in petroleum ether from 150 g of dry plant material. Except water extract, all the extracts possessed significant antifungal property. All the test pathogens showed highest sensitivity towards chloroform extract, exhibiting maximum inhibition zone diameter of 50.0 mm in T. mentagrophytes and minimum 30.0 mm in C. albicans at 2 × 10⁵ μg/ml concentration. Chloroform extract at lower concentration 2.5 × 10⁴ μg/ml was inhibitory for all the test pathogens, exhibiting inhibition zone diameter 21.0 mm against T. tonsurans and 15.0 mm against C. albicans and T. beigelii. The activity of the different solvent extracts against the test pathogens in terms of inhibition zone diameter in decreasing order was as followsChloroform extract > Petroleum ether extract > Methanol extract for T. mentagrophytes, T. rubrum and T. tonsurans.Chloroform extract > Methanol extract > Petroleum ether extract for C. albicans and T. beigelii.     

Effectiveness of five antidandruff cosmetic formulations against planktonic cells and biofilms of dermatophytes

This study aimed to investigate the in vitro antifungal effectiveness of five different formulations against dandruff and ringworm dermatophytes. Candida albicans was also included in our assays. Fungal susceptibility tests were performed with planktonic cells and biofilms of reference strains. Microbiological and physicochemical quality parameters were assessed for all formulations. Our data indicated that the formulations were effective against the dermatophytes strains, and to our knowledge, the effectiveness of cosmetic formulations against fungal biofilms is shown for the first time. The formulations were considered effective against the explored dermatophytes and were considered safe given the adequate microbiological and physicochemical characteristics shown in the proposed assays.     

The Influence of Chemical Composition of Commercial Lemon Essential Oils on the Growth of Candida Strains

Candida yeasts are saprophytes naturally present in the environment and forming colonies on human mucous membranes and skin. They are opportunistic fungi that cause severe and even fatal infections in immunocompromised individuals. Several essential oils, including eucalyptus, pine, cinnamon and lemon, have been shown to be effective against Candida strains. This study addresses the chemical composition of some commercial lemon essential oils and their antifungal potential against selected Candida yeast strains. Antifungal potential and minimum inhibitory concentrations were determined for six commercial lemon essential oils against five Candida yeast strains (Candida albicans 31, Candida tropicalis 32, Candida glabrata 33, Candida glabrata 35 and Candida glabrata 38). On the basis of the GCMS analysis, it was found that the tested lemon essential oils had different chemical compositions, but mostly, they contained almost exclusively terpenes and oxygenated terpenes. The tests show that antifungal potential of lemon essential oils against Candida yeast strains was related to the high content of monoterpenoids and the type of Candida strains. From six tested commercial oils, only four (ETJA, Vera-Nord, Avicenna-Oil and Aromatic Art) shows antifungal potential against three Candida species (C. albicans, C. tropicalis and C. glabrata). Vera-Nord and Avicenna-Oil show the best activity and effectively inhibit the growth of the C. albicans strain across the full range of the concentrations used. Our study characterises lemon essential oils, which could be used as very effective natural remedies against candidiasis caused by C. albicans.     

Evaluation of the antifungal photodynamic activity of Thymophylla pentachaeta extracts against Candida albicans and its virulence factors

BackgroundCandida albicans is one of the most common causative of opportunistic infections. Treatment of candidiasis is challenging considering the few antifungal drugs available and the increase in resistance. Antimicrobial photodynamic therapy (aPDT) is a recently developed therapeutic option that combines a non-toxic photosensitizer (PS) and light to kill the microbial pathogens. Targeting virulence, defined as the ability of a pathogen to cause overt disease, represents another attractive target for the development of novel antifungal agents. Thymophylla pentachaeta (DC.) Small var. belenidium (DC.) is an endemic plant from Argentina in which the presence of thiophenes, biologically active compounds whose antifungal activity is enhanced by irradiation with Ultraviolet A (UVA), have been already described.PurposeThe purpose of this study was to evaluate the photodynamic antifungal activity of hexane (Hex), dichloromethane (DCM), ethyl acetate (EtOAc) and methanol (MeOH) extracts from T. pentachaeta var. belenidium and their inhibitory effects on C. albicans virulence factors as well as biofilm formation and eradication.Study Design/MethodsAntifungal photodynamic activity of Hex, DCM, EtOAc and MeOH extracts from different parts of the plant were assessed with the microbroth dilution, bioautography and the time-kill assays, under light and darkness conditions. The capacities of the most active extracts of inhibiting Candida virulence factors (adherence to epithelial cells, germ tube and pseudomycelium formation and hydrolytic enzyme secretion) were assessed. In addition, the activity against biofilm formation and eradication has been investigated by reaction with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) that quantifies living cells in these structures.ResultsHex and DCM extracts from T. pentachaeta roots exhibited high photodynamic antifungal activity against C. albicans [Minimal fungicide concentrations (MFCs)= 7.8 µg/ml] under UVA light irradiation. Chemical analysis of active extracts (Hex and DCM from roots) revealed the presence of photoactive thiophenes. Both extracts generate reactive oxygen species through type I and II mechanisms. These extracts, at sub-inhibitory concentrations, under light conditions decreased the adherence of C. albicans to Buccal Epithelial Cells (BEC), inhibited germ tube formation and reduced esterase production. Finally, they demonstrated activity against preformed biofilms submitted to irradiation (MFCs= 3.91 µg/ml and 15.63 µg/ml for Hex and DCM extracts, respectively).ConclusionTaking together, results demonstrated the strong photodynamic effects of T. pentachaeta root extracts under UVA irradiation, making them valuable alternatives to the already established antifungal drugs against C. albicans.     

Priming of Nicotiana benthamiana antioxidant defences using brown seaweed extracts

Extracts from the brown seaweeds Cystoseira myriophylloides, Laminaria digitata and Fucus spiralis were evaluated as plant defence inducers against the wild fire disease of Nicotiana benthamiana. Seeds’ imbibition in aqueous seaweed extracts (SE) results in plants with reduced disease severity. In addition, bacterial populations were significantly reduced in these plants when compared to those germinated in sterile distilled water. They primed N. benthamiana for H₂O₂ accumulation and for enhanced activity of several antioxidant enzymes including catalase, ascorbate peroxidase, guaiacol peroxidase and polyphenol oxidase. These results revealed that soaking of seeds in SE before sowing allows N. benthamiana to reduce pathogen attack and can be easily applied in practice.     

Disruption of the intestinal mucosal barrier in Candida albicans infections

Candida albicans is a common microorganism in the intestine. However, invasive C. albicans infection has emerged as a life-threatening disease in recent years. The mortality rate of invasive candidiasis is high in critically ill hosts. C. albicans can switch from the yeast to the hyphal morphology, and take advantage of the impaired intestinal mucosal barrier and insufficient immunity of the host to facilitate its colonization and penetration. Despite the availability of potent new antifungal drugs in recent years, the treatment of severe candidiasis, especially candidaemia, has not been substantially improved. In this review, the virulence factors of C. albicans, as well as the antagonistic role of the intestinal mucosal barrier will be discussed to illuminate the mechanisms of C. albicans enterogenic infections.     

Nutritional value of selected macroalgae

Macroalgae are traditionally used in human and animal nutrition. Their protein and fiber content have been widely studied and differ according to the species, their geographic origin and their seasonal conditions. In addition to their value for human nutrition, seaweeds have multiple therapeutically applications (e.g., weight control, hypocholesterolemic, antioxidant and antitumor activities, others) and, in general, contribute and promote human health. In the archipelago of the Azores, the consumption of seaweeds is widespread and accepted as a common practice in some islands. This work is aimed at providing information on the protein and fiber content of the locally consumed species, to promote this regional food product that can be potentially profitable from the biotechnology and commercial perspective, and also benefit public health, particularly, taking into account the low level of marine pollution in the Azores archipelago. Protein and fiber content of eight seaweeds (Porphyra sp., Osmundea pinnatifida, Pterocladiella capillacea, Sphaerococcus coronopifolius, and Gelidium microdon, Rhodophyta; Cystoseira abies-marina and Fucus spiralis, Phaeophyta; Ulva compressa, Chlorophyta) were determined using the Kjeldahl method and the Weende method, respectively. The protein content ranged from 6.81 to 26.62 of dry weight for C. abies-marina and U. compressa, respectively. Fiber content was generally higher as compared with that in seaweeds from other origins and ranged from 33.82 to 63.88 for O. pinnatifida and F. spiralis, respectively.     

Anticandidial and anticryptococcal activity of <Emphasis Type="Italic">Euphorbia fusiformis</Emphasis>, a rare medicinal plant

The present work demonstrates the screening of extracts of the rare medicinal herb Euphorbia fusiformis for antifungal activity. The main aim was to investigate its antifungal properties against Candida albicans and Cryptococcus neoformans, the causative agents of human candidiasis and cryptococcosis, respectively. Aqueous and organic solvent extracts from the leaves and rootstock of the plant were tested against the fungi by the well-in-agar method. Almost all the organic solvent extracts exhibited an inhibitory effect against C. albicans and to some extent on C. neoformans, except for the aqueous extracts, which had no effect. The combined formulations of the extracts also had better activity against C. albicans than C. neoformans. This study thus concludes by demonstrating the antifungal properties of E. fusiformis and also the potential research in identifying the active principles, which may have future therapeutic value.     

The antibacterial agent,moxifloxacin inhibits virulence factors of <Emphasis Type="Italic">Candida albicans</Emphasis> through multitargeting

Fluoroquinolines are broad spectrum fourth generation antibiotics. Some of the Fluoroquinolines exhibit antifungal activity. We are reporting the potential mechanism of action of a fluoroquinoline antibiotic, moxifloxacin on the growth, morphogenesis and biofilm formation of the human pathogen Candida albicans. Moxifloxacin was found to be Candidacidal in nature. Moxifloxacin seems to inhibit the yeast to Hyphal morphogenesis by affecting signaling pathways. It arrested the cell cycle of C. albicans at S phase. Docking of moxifloxacin with predicted structure of C. albicans DNA Topoisomerase II suggests that moxifloxacin may bind and inhibit the activity of DNA Topoisomerase II in C. albicans. Moxifloxacin could be used as a dual purpose antibiotic for treating mixed infections caused by bacteria as well as C. albicans. In addition chances of developing moxifloxacin resistance in C. albicans are less considering the fact that moxifloxacin may target multiple steps in yeast to hyphal transition in C. albicans.     

Infection Risk by Dermatophytes During Storage and After Domestic Laundry and Their Temperature-Dependent Inactivation

In the developed countries infections of the feet (tinea pedis, athlete’s foot) and nails (onychomycosis) with the anthropophile fungus Trichophyton rubrum are most common. We examined the propagation of dermatophytes before and during domestic laundering. About 10% of the infectious material was transferred from contaminated textiles to sterile textiles during storage in a clothes basket simulation indicating a high infection risk during storage. This was evaluated with two quantification techniques: cultivation with subsequent colony counting and tracing of radioactively labelled propagating units. Both approaches reliably revealed similar results with the latter method reducing experimental time to few minutes compared to 2 weeks with the traditional method. The tracer technique allowed favourably to directly reflect the textile-bound infectious material at the moment of skin contact. To address the infection risk during domestic laundry, bioindicators with T. rubrum or the yeast Candida albicans were introduced into common domestic washing procedures with different temperature courses. While C. albicans did not survive any of the tests, T. rubrum could be recovered after washing at 30°C, indicating the risk potential of dermatophyte infections at home. Up to 16% of the initial fungus load was detected in the rinsing water. Washing at 60°C however, eliminated both pathogens, T. rubrum and C. albicans.     

Phytochemical Analysis and Evaluation of the Antioxidant,Anti-Inflammatory,and Antinociceptive Potential of Phlorotannin-Rich Fractions from Three Mediterranean Brown Seaweeds

Phlorotannins, phenolic compounds produced exclusively by seaweeds, have been reported to possess various pharmacological properties. However, there have been few works on these compounds from Mediterranean seaweeds. In this study, we investigated the phytochemical analysis and pharmacological potential of phlorotannin-rich fractions from three brown seaweeds collected along the Tunisia coast: Cystoseira sedoides (PHT-SED), Cladostephus spongeosis (PHT-CLAD), and Padina pavonica (PHT-PAD). Phytochemical determinations showed considerable differences in total phenolic content (TPC) and phlorotannin content (PHT). The highest TPC level (26.45 mg PGE/g dry material (Dm)) and PHT level (873.14 μg PGE/g Dm) were observed in C. sedoides. The antioxidant properties of these three fractions assessed by three different methods indicated that C. sedoides displayed the highest total antioxidant activity among the three species (71.30 mg GAE/g Dm), as well as the free radical scavenging activity with the lowest IC₅₀ value in both DPPH (27.7 μg/mL) and ABTS (19.1 μg/mL) assays. Furthermore, the pharmacological screening of the anti-inflammatory potential of these fractions using in vivo models, in comparison to reference drugs, established a remarkable activity of PHT-SED at the dose of 100 mg/kg; the inhibition percentages of ear edema in mice model and paw edema in rats model were of 82.55 and 81.08%, respectively. The content of malondialdehyde (MDA) in liver tissues has been quantified, and PHT-SED was found to remarkably increase the lipid peroxidation in rat liver tissues. In addition, in two pain mice models, PHT-SED displayed a profound antinociceptive activity at 100 mg/kg and has proved a better analgesic activity when used in combination with the opioid drug, tramadol.     

Antifungal properties of some pyrazolyl-alkyl-sulfides

Four 5-amino-4-alkylthio-pyrazoles were synthesized and their antifungal activity was evaluated in vitro in Trichophyton mentagrophytes, Microsporum cookei and Candida albicans. The compounds slightly influenced the growth kinetics of the yeast, but at concentrations ranging from 20 to 40 g/ml completely prevented the mycelial growth of the two dermatophytes cultivated on Sabouraud's agar medium. An electron microscopic study, undertaken by using the most active compound, showed that in C. albicans mitochondria were the only cell targets affected whereas in the dermatophytes cell wall, plasmalemma and the main cytoplasmic organelles were damaged in various degrees. Since the most remarkable alterations were connected with membrane abnormalities, the cytological changes observed were tentatively interpreted as a consequence of the compound intrusion into the lipid bilayer of the membranes, since the drug is lipophilic in nature.Investigations supported by a grant from Consiglio Nazionale delle Ricerche of Italy (Contract No. 79.01871.04)     

Anti-Candida property of a lignan glycoside derived from Styrax japonica S. et Z. via membrane-active mechanisms

Styraxjaponoside C was investigated with respect to its antifungal activity and mechanisms of action. Devoid of hemolytic activity, Styraxjaponoside C demonstrated an antifungal effect against the human pathogenic yeast Candida albicans in an energy-independent manner. To characterize the mechanisms of the antifungal activity of Styraxjaponoside C, fluorescence analysis with membrane probe 1,6-diphenyl-1,3,5-hexatriene, and flow cytometric analysis on C. albicans were conducted. The results showed that Styraxjaponosdie C induced cytoplasmic membrane perturbation. The current study suggested that Styraxjaponoside C was active against C. albicans with membrane-active mechanisms.     

Influencia del grupo ecológico sobre la sensibilidad in vitro de los hongos dermatofitos a los antifúngicos

BackgroundDermatophytes can be divided into geophilic (soil), zoophilic (animals) and anthropophilic (humans) strains, depending on the source of the keratin that they use for nutritional purposes.AimsThe in vitro susceptibility of clinical isolates of dermatophyte fungi has been studied in the 3 ecological groups with several antifungal agents for the topical management of dermatophytoses in order to determine their relationship with the ecological group.MethodsA standardised dilution micromethod in a liquid medium was used for the determination of the in vitro antifungal activity of 9 topical antifungal drugs: amorolfine (AMR), bifonazole (BFZ), clotrimazole, econazole, ketoconazole, miconazole, oxiconazole, terbinafine (TRB) and tioconazole. The in vitro activity was obtained against 124 clinical isolates of dermatophyte moulds from the anthropophilic, zoophilic and geophilic ecological groups.ResultsThe in vitro antifungal activity was different depending on the ecological group, although a species-dependent profile was also observed.ConclusionsAzole derivatives showed a similar antifungal profile, being more active against anthropophilic dermatophytes > zoophilic > geophilic. Activity of TRB and AMR was different from that of azole derivatives (zoophilic > anthropophilic > geophilic). A higher in vitro antifungal activity against the 3 ecological groups was observed with TRB and AMR, whilst BFZ was the less active drug.