Antimicrobial Activity of Arthrospira (Former Spirulina) and Dunaliella Related to Recognized Antimicrobial Bioactive Compounds.

With the increasing rate of the antimicrobial resistance phenomenon, natural products gain our attention as potential drug candidates. Apart from being used as nutraceuticals and for biotechnological purposes, microalgae and phytoplankton have well-recognized antimicrobial compounds and proved anti-infectious potential. In this review, we comprehensively outline the antimicrobial activity of one genus of cyanobacteria (Arthrospira, formerly Spirulina) and of eukaryotic microalgae (Dunaliella). Both, especially Arthrospira, are mostly used as nutraceuticals and as a source of antioxidants for health supplements, cancer therapy and cosmetics. Their diverse bioactive compounds provide other bioactivities and potential for various medical applications. Their antibacterial and antifungal activity vary in a broad range and are strain specific. There are strains of Arthrospira platensis with very potent activity and minimum inhibitory concentrations (MICs) as low as 2-15 µg/mL against bacterial fish pathogens including Bacillus and Vibrio spp. Arthrospira sp. has demonstrated an inhibition zone (IZ) of 50 mm against Staphylococcus aureus. Remarkable is the substantial amount of in vivo studies of Arthrospira showing it to be very promising for preventing vibriosis in shrimp and Helicobacter pylori infection and for wound healing. The innovative laser irradiation of the chlorophyll it releases can cause photodynamic destruction of bacteria. Dunaliella salina has exhibited MIC values lower than 300 µg/mL and an IZ value of 25.4 mm on different bacteria, while Dunaliella tertiolecta has demonstrated MIC values of 25 and 50 µg/mL against some Staphylococcus spp. These values fulfill the criteria for significant antimicrobial activity and sometimes are comparable or exceed the activity of the control antibiotics. The bioactive compounds which are responsible for that action are fatty acids including PUFAs, polysaccharides, glycosides, peptides, neophytadiene, etc. Cyanobacteria, such as Arthrospira, also particularly have antimicrobial flavonoids, terpenes, alkaloids, saponins, quinones and some unique-to-them compounds, such as phycobiliproteins, polyhydroxybutyrate, the peptide microcystin, etc. These metabolites can be optimized by using stress factors in a two-step process of fermentation in closed photobioreactors (PBRs).

Antimicrobial and Antioxidant Potential of Scenedesmus obliquus Microalgae in the Context of Integral Biorefinery Concept.

Small-scale photobioreactors (PBRs) in the inoculum stage were designed with internal (red or green) and external white LED light as an initial step of a larger-scale installation aimed at fulfilling the integral biorefinery concept for maximum utilization of microalgal biomass in a multifunctional laboratory. The specific growth rate of Scenedesmus obliquus (Turpin) Kützing biomass for given cultural conditions was analyzed by using MAPLE software. For the determination of total polyphenols, flavonoids, chlorophyll "a" and "b", carotenoids and lipids, UHPLC-HRMS, ISO-20776/1, ISO-10993-5 and CUPRAC tests were carried out. Under red light growing, a higher content of polyphenols was found, while the green light favoured the flavonoid accumulation in the biomass. Chlorophylls, carotenoids and lipids were in the same order of magnitude in both samples. The dichloromethane extracts obtained from the biomass of each PBR synergistically potentiated at low concentrations (0.01-0.05 mg/mL) the antibacterial activity of penicillin, fluoroquinolones or oregano essential oil against the selected food-borne pathogens (Staphylococcus aureus, Escherichia coli and Salmonella typhimurium) without showing any in vitro cytotoxicity. Both extracts exhibited good cupric ion-reducing antioxidant capacity at concentrations above 0.042-0.08 mg/mL. The UHPLC-HRMS analysis revealed that both extracts contained long chain fatty acids and carotenoids thus explaining their antibacterial and antioxidant potential. The applied engineering approach showed a great potential to modify microalgae metabolism for the synthesis of target compounds by S. obliquus with capacity for the development of health-promoting nutraceuticals for poultry farming.

Junipers of Various Origins as Potential Sources of the Anticancer Drug Precursor Podophyllotoxin.

Juniper representatives are natural sources of plenty of bioactive metabolites and have been used since ancient times as folk remedies against tapeworms, warts, cancer, etc. The antiproliferative activities of junipers are attributed to podophyllotoxin (PPT), which is a precursor for the synthesis of efficient anticancer drugs. However, the natural sources of PPT, Sinopodophyllum hexandrum (Royle) T. S. Ying and Podophyllum peltatum L., are already endangered species because of their intensive industrial exploitation. Therefore, identification of other sources of PPT is necessary. This study is a broad comparative investigation of junipers, for which original sources have been accessed from different continents of the world. The present research is aimed at the identification of species, producing PPT and other lignans at concentrations that are sufficient for the high antiproliferative activity of the corresponding extracts. Cytotoxic juniper leaf extracts demonstrated a broad spectrum of activity on a panel of cancer cell lines. The antiproliferative properties of junipers were attributed to the combined activity of great diversity of lignans (podophyllotoxin, deoxypodophyllotoxin, ß-peltatin, yatein, matairesinol, anhydropodorhizol, etc.), detected by UHPLC-HRMS and LC-ESI-MS/MS in the corresponding extracts. Several species of the genus Juniperus L. were outlined as perspective sources of drug precursors with potential pharmaceutical applications.

Polycarbonate-Based Copolymer Micelles as Biodegradable Carriers of Anticancer Podophyllotoxin or Juniper Extracts.

Podophyllotoxin (PPT) is used in the industrial production of efficient anticancer, antiviral and other drugs. Sinopodophyllum hexandrum or Podophyllum peltatum are natural sources of PPT, but at present they are considered as endangered species. Their PPT content is variable, depending on the growing conditions. Searching for new sources of PPT, some representatives of the genus Juniperus were found to exhibit efficient PPT biosynthesis. However, PPT is highly toxic and poorly soluble in water compound, which limits its clinical applications. In this connection, amphiphilic polymer micelles are considered to be suitable PPT carriers, aimed at increase in water solubility and decrease in toxicity. The present research deals with the evaluation of MPEG-polycarbonate block copolymer micelles loaded with PPT or juniper extracts. The active component-loaded polymer nanocarriers were characterized by dynamic and electrophoretic light scattering, as well as by transmission electron microscopy. The active component loading efficiency and loading capacity were also determined. Highly efficient antiproliferative activity of the loaded micelles was determined in a panel of cancer cell lines. The obtained amphiphilic nanocarriers, loaded with PPT-containing bioactive components, have application in future in vivo preclinical trials of their pharmacokinetics and pharmacodynamics as potential therapeutical agents in the prospective nanomedicine.

Pathogenic Potential of Opportunistic Gram-Negative Bacteria Isolated from the Cloacal Microbiota of Free-Living Reptile Hosts Originating from Bulgaria.

Reptiles are known to be asymptomatic carriers of various zoonotic pathogens. A number of Gram-negative opportunistic commensals are causative agents of bacterial infections in immunocompromised or stressed hosts and are disseminated by reptiles, whose epidemiological role should not be neglected. Since most studies have focused on exotic species, in captivity or as pet animals, the role of wild populations as a potential source of pathogens still remains understudied. In the present study, we isolated a variety of Gram-negative bacteria from the cloacal microbiota of free-living lizard and tortoise hosts (Reptilia: Sauria and Testudines) from the Bulgarian herpetofauna. We evaluated their pathogenic potential according to their antibiotic susceptibility patterns, biofilm-forming capacity, and extracellular production of some enzymes considered to play roles as virulence factors. To our knowledge, the phenotypic manifestation of virulence factors/enzymatic activity and biofilm formation in wild reptile microbiota has not yet been widely investigated. All isolates were found to be capable of forming biofilms to some extent and 29.6% of them could be categorized as strong producers. Two strains proved to be excellent producers. The majority of the isolated strains showed extracellular production of at least one exoenzyme. The most pronounced pathogenicity could be attributed to the newly isolated Pseudomonas aeruginosa strain due to its multiresistance, excellent biofilm formation, and expression of exoenzymes.

Insight into the influence of natural deep eutectic solvents on the extraction of phenolic compounds from poplar type propolis: Composition and in vitro biological activity.

Natural deep eutectic solvents (NADESs) have been considered promising to replace traditional volatile and toxic organic solvents for the extraction of biologically active substances from natural sources. This work applied an efficient and ethanol-exclusion strategy for extraction of phenolic compounds from poplar type propolis using five known NADESs (lactic acid:1,2-propanediol 1:1, lactic acid:fructose 5:1, choline chloride:1,2-propanediol 1:3, choline chloride:1,2-propanediol:water 1:1:1 and betaine:malic acid:water 1:1:6). The selected NADESs' extractability was evaluated by measuring the concentrations of total phenolics and total flavones and flavonols in the propolis extracts obtained, which qualitative chemical composition was further determined in detail by gas chromatography-mass spectrometry (GC-MS) analysis. It demonstrated that the chemical profiles of NADES and 70% ethanolic propolis extracts are similar. To expand the knowledge about the role of the applied solvents in the poplar propolis extraction process, the in vitro antimicrobial, cytotoxic and genotoxic activity of both NADESs and liquid NADES extracts were evaluated. The results revealed that the use of the selected NADESs as an extraction media for phenolic compounds from poplar propolis not only delivered a good extraction yield in some cases, but generally led to the preservation of propolis extracts' biological activity and even to the enhancement of their antimicrobial effect in comparison with the hydroethanolic one. Besides, the tested NADESs except for lactic acid:fructose and betaine:malic acid:water exerted low to negligible toxicity against normal cells treated and apart from lactic acid:fructose the remaining solvents demonstrated concentration-dependent moderate to subtle genotoxicity. There is a probability that not the supramolecular structure of the NADESs, but their components, played a key role for the observed biological effects. The present study has demonstrated an alternative approach for extracting the biologically active complex from poplar type propolis using NADESs, which could be useful for further pharmaceutical and cosmeceutical applications.

Outstanding Antibacterial Activity of Hypericum rochelii-Comparison of the Antimicrobial Effects of Extracts and Fractions from Four Hypericum Species Growing in Bulgaria with a Focus on Prenylated Phloroglucinols.

Microbial infections are by no means a health problem from a past era due to the increasing antimicrobial resistance of infectious strains. Medicine is in constant need of new drugs and, recently, plant products have had a deserved renaissance and garnered scientific recognition. The aim of this work was to assess the antimicrobial activity of ten active ingredients from four Hypericum species growing in Bulgaria, as well as to obtain preliminary data on the phytochemical composition of the most promising samples. Extracts and fractions from H. rochelii Griseb. ex Schenk, H. hirsutum L., H. barbatum Jacq. and H. rumeliacum Boiss. obtained with conventional or supercritical CO2 extraction were tested on a panel of pathogenic microorganisms using broth microdilution, agar plates, dehydrogenase activity and biofilm assays. The panel of samples showed from weak to extraordinary antibacterial effects. Three of them (from H. rochelii and H. hirsutum) had minimum inhibitory concentrations as low as 0.625-78 mg/L and minimum bactericidal concentrations of 19.5-625 mg/L against Staphylococcus aureus and other Gram-positive bacteria. These values placed these samples among the best antibacterial extracts from the Hypericum genus. Some of the agents also demonstrated very high antibiofilm activity against methicillin-resistant S. aureus. Ultra-high-performance liquid chromatography-high-resolution mass spectrometry revealed the three most potent samples as rich sources of biologically active phloroglucinols. They were shown to be good drug or nutraceutical candidates, presumably without some of the side effects of conventional antibiotics.

Antimicrobial Resistance and Biofilm Formation of Escherichia coli Isolated from Pig Farms and Surroundings in Bulgaria.

Escherichia coli (E. coli) is a ubiquitous microorganism with pathogenic and saprophytic clones. The objective of this study was to evaluate the presence, virulence, antibiotic resistance and biofilm formation of E. coli in three industrial farms in Bulgaria, as well as their adjacent sites related to the utilization of manure (feces, wastewater in a separator, lagoons, means of transport, and soils). The isolation of single bacterial cultures was performed via standard procedures with modifications, and E. coli isolates were identified via matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and polymerase chain reaction (PCR). The disk diffusion method was used to assess antimicrobial resistance, and PCR was used to detect genes for antibiotic resistance (GAR) (qnr, aac(3), ampC, blaSHV/blaTEM and erm) and virulence genes (stx, stx2all, LT, STa, F4 and eae). The protocol of Stepanovic was utilized to measure the biofilm formation of the isolates. A total of 84 isolates from different samples (n = 53) were identified as E. coli. Almost all demonstrated antimicrobial resistance, and most of them demonstrated resistance to multiple antibiotics from different classes. No virulence genes coding the Shiga toxin or enterotoxins or those associated with enteropathogenicity were detected. No GAR from those tested for quinolones, aminoglycosides and macrolides were found. However, all isolates that were resistant to a penicillin-class antibiotic (56) had ß-lactamase-producing plasmid genes. All of them had ampC, and 34 of them had blaTEM. A total of 14 isolates formed strongly adherent biofilms. These results in a country where the use of antibiotics for growth promotion and prophylaxis in farms is highly restricted corroborate that the global implemented policy on antibiotics in human medicine and in animal husbandry needs revision.

Cytotoxic and Antibacterial Prenylated Acylphloroglucinols from Hypericum olympicum L.

Two new bicyclo[3.3.1]nonane type bicyclic polyprenylated acylphloroglucinol derivatives (BPAPs), olympiforin A and B as well as three known prenylated phloroglucinols, were isolated from the aerial parts of Hypericum olympicum L. The structures of the isolated compounds were established by means of spectral techniques (HRESIMS and 1D and 2D NMR). All compounds were tested on a panel of human tumor (MDA-MB-231, EJ, K-562, HL-60 and HL-60/DOX) and non- tumorigenic (HEK-293 and EA.hy926) cell lines using the MTT assay. All tested compounds exerted significant in vitro cytotoxicity with IC50 values ranging from 1.2 to 24.9 µM and from 0.9 to 34 µM on tumor and non-cancerous cell lines, respectively. Most of the compounds had good selectivity and were more cytotoxic to the tumor cell lines than to the normal ones. A degradation of the precursor caspase 9 for some of the compounds was observed; therefore, the intrinsic pathway of apoptosis is the most likely mechanism of cytotoxic activity. The BPAPs were examined for antibacterial and antibiofilm activity through the broth microdilution method and the protocol of Stepanovic. They showed a moderate effect against Enterococcus faecalis and Streptococcus pyogenes but a very profound activity against Staphylococcus aureus with minimum inhibitory concentrations (MIC) in the range of 0.78-2 mg/L. Olympiforin B also had a great effect against methicillin-resistant S. aureus (MRSA) with an MIC value of 1 mg/L and a very significant antibiofilm activity on that strain with a minimum biofilm inhibition concentration (MBIC) value of 0.5 mg/L. The structures of the isolated compounds were in silico evaluated using ADME and drug likeness tests.

Prevalence, Genetic Homogeneity, and Antibiotic Resistance of Pathogenic Yersinia enterocolitica Strains Isolated from Slaughtered Pigs in Bulgaria.

Yersiniosis is the third most commonly reported foodborne zoonosis in the European Union. Here, we evaluated the prevalence of pathogenic Yersinia enterocolitica among healthy pigs (as a major reservoir) in a slaughterhouse in Bulgaria. A total of 790 tonsils and feces from 601 pigs were examined. Isolation and pathogenicity characterization was carried out by the ISO 10273:2003 protocol and Polymerase Chain Reaction (PCR), detecting the 16S rRNA gene, attachment and invasion locus (ail), Yersinia heat-stable enterotoxin (ystA), and Yersinia adhesion (yadA) genes. Genetic diversity was assessed by pulsed-field gel electrophoresis (PFGE), and antimicrobial resistance by the standard disk diffusion method. Of all the pigs tested, 6.7% were positive for Y. enterocolitica. All isolates belonged to Y. enterocolitica bioserotype 4/O:3. ail, and ystA genes were detected in all positive strains (n = 43), while the plasmid Yersinia virulence plasmid (pYV) was detected in 41. High homogeneity was observed among the strains, with all strains susceptible to ceftriaxone, amikacin and ciprofloxacin, and resistant to ampicillin. In conclusion, a low prevalence of Y. enterocolitica 4/O:3 was found in healthy pigs slaughtered in Bulgaria, not underestimating possible contamination of pork as a potential risk to consumer health.

Photodynamic Inactivation of Bovine Coronavirus with the Photosensitizer Toluidine Blue O.

Coronaviruses (CoVs) belong to the group of enveloped positive-sense single-strand RNA viruses and are causative agents of respiratory, gastro-intestinal, and central nervous systems diseases in many host species, i.e., birds, mammals, and humans. Beta-CoVs revealed a great potential to cross the barrier between species by causing three epidemics/pandemics among humans in the 21st century. Considering the urgent need for powerful antiviral agents for decontamination, prevention, and treatment of BCoV infections, we turned our attention to the possibility of photodynamic inactivation with photosensitizers in combination with light irradiation. In the present study, we evaluated, for the first time, the antiviral activity of toluidine blue O (TBO) against Beta-coronavirus 1 (BCoV) in comparison to methylene blue (MB). First, we determined the in vitro cytotoxicity of MB and TBO on the Madin-Darby bovine kidney (MDBK) cell line with ISO10993-5/Annex C. Thereafter, BCoV was propagated in MDBK cells, and the virus titer was measured with digital droplet PCR, TCID50 assay and plaque assay. The antiviral activity of non-toxic concentrations of TBO was estimated using the direct inactivation approach. All effects were calculated in MAPLE 15® mathematical software by developing programs for non-linear modeling and response surface analysis. The median inhibitory concentration (IC50) of TBO after 72 h of incubation in MDBK cells was 0.85 µM. The antiviral activity of TBO after the direct inactivation of BCoV (MOI = 1) was significantly stronger than that of MB. The median effective concentration (EC50) of TBO was 0.005 µM. The cytopathic effect decreased in a concentration-dependent manner, from 0.0025 to 0.01 µM, and disappeared fully at concentrations between 0.02 and 0.3 µM of TBO. The number of virus particles also decreased, depending on the concentration applied, as proven by ddPCR analysis. In conclusion, TBO exhibits significant potential for direct inactivation of BCoV in vitro, with a very high selectivity index, and should be subjected to further investigation, aiming at its application in veterinary and/or human medical practice.

In Vitro Study of the Biological Potential of Wastewater Obtained after the Distillation of Four Bulgarian Oil-Bearing Roses.

The wastewater after rose oil distillation is usually discharged into the drainage systems and it represents a serious environmental problem. While being rich in polyphenols, which have beneficial biological activity and application in the pharmaceutical industry, limited research has been carried out about the biological activity of the specific wastewaters per se. Wastewaters after distillation of the four Bulgarian oil-bearing roses Rosa damascena Mill., R. alba L., R. centifolia L., and R. gallica L. exerted significant antioxidant activity and good antiherpes simplex virus type-1 (HSV-1) activity while maintaining a good toxicological safety profile (low cytotoxic effect) towards normal cell lines. More precisely, the non-tumorigenic cells were a human (HEK-293 embryonic kidney cells) and a mouse cell line (CCL-1 fibroblasts, which are recommended as a standard for cytotoxicity evaluation in Annex C of ISO 10993-5). The concentrations that achieved antioxidant and radical scavenging effects (0.04-0.92% v/v) were much lower than most of the maximum tolerated concentrations for the tissue culture cells (0.2-3.4% v/v). The wastewaters had a weak antiproliferative effect against Staphylococcus aureus. None of the wastewaters had activity against Gram-negative bacteria or a bactericidal or antifungal effect. We can conclude that these four species, which are the most preferred species worldwide for producing high-quality rose oil, have the potential to be developed as promising antioxidant and antiherpesvirus nutraceuticals.

Micellar Curcumin Substantially Increases the Antineoplastic Activity of the Alkylphosphocholine Erufosine against TWIST1 Positive Cutaneous T Cell Lymphoma Cell Lines.

Cutaneous T-cell lymphoma (CTCL) is a rare form of cancer with local as well as systemic manifestations. Concomitant bacterial infections increase morbidity and mortality rates due to impaired skin barrier and immune deficiency. In the current study, we demonstrated that the in vitro anti-lymphoma potential of erufosine is diminished by TWIST1 expression and micellar curcumin substantially increases its antineoplastic activity. Pharmacokinetic analysis showed that the micellar curcumin (MCRM) used in our study was characterized by low zeta potential, slow release of curcumin, and fast cell membrane penetration. The combination ratio 1:4 [erufosine:MCRM] achieved strong synergism by inhibiting cell proliferation and clonogenicity. The combined antiproliferative effects were calculated using the symbolic mathematical software MAPLE 15. The synergistic combination strongly decreased the expression of TWIST1 and protein kinase B/Akt as proven by western blotting. Significant reductions in NF-κB activation, induction of apoptosis, and altered glutathione levels were demonstrated by corresponding assays. In addition, the synergistic combination enhanced the anti-staphylococcal activity and prevented biofilm formation, as shown by crystal violet staining. Taken together, the above results show that the development of nanotechnological treatment modalities for CTCL, based on rational drug combinations exhibiting parallel antineoplastic and antibacterial effects, may prove efficacious.

Redox-Modulating Capacity and Antineoplastic Activity of Wastewater Obtained from the Distillation of the Essential Oils of Four Bulgarian Oil-Bearing Roses.

The wastewater from the distillation of rose oils is discharged directly into the soil because it has a limited potential for future applications. The aim of the present study was to determine in vitro the chromatographic profile, redox-modulating capacity, and antineoplastic activity of wastewater obtained by distillation of essential oils from the Bulgarian Rosa alba L., Rosa damascena Mill., Rosa gallica L., and Rosa centifolia L. We applied UHPLC-HRMS for chromatographic analysis of rose wastewaters, studied their metal-chelating and Fe(III)-reducing ability, and performed MTT assay for the evaluation of cytotoxic potential against three tumorigenic (HEPG2-hepatocellular adenocarcinoma, A-375-malignant melanoma, A-431-non-melanoma epidermoid squamous skin carcinoma) and one non-tumorigenic human cell lines (HaCaT-immortalized keratinocytes). The median inhibitory concentrations (IC50) were calculated with nonlinear modeling using the MAPLE® platform. The potential of the wastewaters to induce apoptosis was also examined. Mono-, di-, and acylated glycosides of quercetin and kaempferol, ellagic acid and its derivatives as main chemical components, and gallic acid and its derivatives-such as catechin and epicatechin-were identified. The redox-modulating capacity of the samples (TPTZ test) showed that all four wastewaters exhibited the properties of excellent heavy metal cleaners, but did not exert very strong cytotoxic effects. The lowest IC50 rate was provided in wastewater from R. centifolia (34-35 µg/mL of gallic acid equivalents after a 72 h period for all cell lines). At 24 and 48 hours, the most resistant cell line was HEPG2, followed by HaCaT. After 72 h of exposure, the IC50 values were similar for tumor and normal cells. Still, R. damascena had a selectivity index over 2.0 regarding A-431 non-melanoma skin cancer cells, showing a good toxicological safety profile in addition to moderate activity-IC50 of 35 µg/mL polyphenols. The obtained results related to wastewaters acquired after the distillation of essential oils from the Bulgarian R. alba, R. damascena, R. gallica, and R. centifolia direct our attention to further studies for in-depth elucidation of their application as detoxifying agents under oxidative damage conditions in other experimental datasets.

Prevalence of Antibiotic-Resistant Escherichia coli Isolated from Swine Faeces and Lagoons in Bulgaria.

Antimicrobial resistance (AMR) is a worldwide health problem affecting humans, animals, and the environment within the framework of the "One Health" concept. The aim of our study was to evaluate the prevalence of pathogenic strains of the species Escherichia coli (E. coli), their AMR profile, and biofilm-forming potential. The isolated strains from three swine faeces and free lagoons (ISO 16654:2001/Amd 1:2017) were confirmed using Phoenix M50 and 16S rDNA PCR. The antibiotic sensitivity to 34 clinically applied antibiotics was determined by Phoenix M50 and the disc diffusion method, according to the protocols of the CLSI and EUCAST. We confirmed the presence of 16 E. coli isolates, of which 87.5% were multi-drug-resistant and 31.25% performed strong biofilms. The possibility for the carrying and transmission of antibiotic-resistance genes to quinolones (qnr), aminoglycosides (aac(3)), ß-lactamase-producing plasmid genes ampC, and blaSHV/blaTEM was investigated. We confirmed the carrying of blaSHV/blaTEM in one and ampC in seven isolates. The strains were negative for the virulence genes (ETEC (LT, STa, and F4), EPEC (eae), and STEC/VTEC (stx and stx2all)). The results should contribute to the development of effective measures for limitation and control on the use of antibiotics, which is a key point in the WHO action plan.

Rose Flowers-A Delicate Perfume or a Natural Healer?

Plants from the Rosacea family are rich in natural molecules with beneficial biological properties, and they are widely appreciated and used in the food industry, perfumery, and cosmetics. In this review, we are considering Rosa damascena Mill., Rosa alba L., Rosa centifolia L., and Rosa gallica L. as raw materials important for producing commercial products, analyzing and comparing the main biological activities of their essential oils, hydrolates, and extracts. A literature search was performed to find materials describing (i) botanical characteristics; (ii) the phytochemical profile; and (iii) biological properties of the essential oil sand extracts of these so called "old roses" that are cultivated in Bulgaria, Turkey, India, and the Middle East. The information used is from databases PubMed, Science Direct, and Google Scholar. Roses have beneficial healing properties due to their richness of beneficial components, the secondary metabolites as flavonoids (e.g., flavones, flavonols, anthocyanins), fragrant components (essential oils, e.g., monoterpenes, sesquiterpenes), and hydrolysable and condensed tannins. Rose essential oils and extracts with their therapeutic properties-as respiratory antiseptics, anti-inflammatories, mucolytics, expectorants, decongestants, and antioxidants-are able to act as symptomatic prophylactics and drugs, and in this way alleviate dramatic sufferings during severe diseases.

Cytotoxicity and Microbicidal Activity of Commonly Used Organic Solvents: A Comparative Study and Application to a Standardized Extract from Vaccinium macrocarpon.

The cytotoxicity and microbicidal capacity of seven organic solvents commonly applied for studying plant extracts and bioactive compounds were systematically investigated based on international standards. Four cell lines of normal (CCL-1, HaCaT) or tumor (A-375, A-431) tissue origin, seven bacterial and one fungal strain were used. The impact of the least toxic solvents in the determination of in vitro cytotoxicity was evaluated using a standardized extract from Vaccinium macrocarpon containing 54.2% v/v proanthocyanidins (CystiCran®). The solvents ethanol, methoxyethanol and polyethylene glycol were the least cytotoxic to all cell lines, with a maximum tolerated concentration (MTC) between 1 and 2% v/v. Ethanol, methanol and polyethylene glycol were mostly suitable for antimicrobial susceptibility testing, with minimum inhibitory concentrations (MICs) ≥ 25% v/v. The MTC values of the solvents dimethyl sulfoxide, dimethoxyethane and dimethylformamide varied from 0.03% to 1.09% v/v. The MICs of dimethyl sulfoxide, methoxyethanol and dimethoxyethane were in the range of 3.125-25% v/v. The cytotoxic effects of CystiCran® on eukaryotic cell lines were directly proportional to the superimposed effect of the solvents used. The results of this study can be useful for selecting the appropriate solvents for in vitro estimation of the cytotoxic and growth inhibitory effects of bioactive molecules in eukaryotic and prokaryotic cells.

Extracts of medicinal plants with natural deep eutectic solvents: enhanced antimicrobial activity and low genotoxicity.

Natural deep eutectic solvents (NADES) are a new alternative to toxic organic solvents. Their constituents are primary metabolites, non-toxic, biocompatible and sustainable. In this study four selected NADES were applied for the extraction of two medicinal plants: Sideritis scardica, and Plantago major as an alternative to water-alcohol mixtures, and the antimicrobial and genotoxic potential of the extracts were studied. The extraction efficiency was evaluated by measuring the extracted total phenolics, and total flavonoids. Best extraction results for total phenolics for the studied plants were obtained with choline chloride-glucose 5:2 plus 30% water; but surprisingly these extracts were inactive against all tested microorganisms. Extracts with citric acid-1,2-propanediol 1:4 and choline chloride-glycerol 1:2 showed good activity against S. pyogenes, E. coli, S. aureus, and C. albicans. Low genotoxicity and cytotoxicity were observed for all four NADES and the extracts with antimicrobial activity. Our results confirm the potential of NADESs for extraction of bioactive constituents of medicinal plants and further suggest that NADES can improve the effects of bioactive extracts. Further studies are needed to clarify the influence of the studied NADES on the bioactivity of dissolved substances, and the possibility to use such extracts in the pharmaceutical and food industry.

Three new prenyloxy chromanones from aerial parts of Hypericum aucheri.

Three new prenyloxy chromanone derivatives, aucherine A-C (6, 7 and 9) as well as six known prenylated phloroglucinols (1-5 and 8) were isolated from the aerial parts of Hypericum aucheri Jaub. Et Spach. The structures of the isolated compounds were established by means of spectral techniques (HRESIMS, 1D and 2D NMR). The new compounds were tested on а panel of human tumor cell line using MTT assay. All tested compounds exerted moderate cytotoxicity with IC50 values ranging from 19.6 to 57.8 µM. The influence of the new compounds on some key signaling molecules (procaspase-9 and Bcl-xL), implicated in the regulation of programmed cell death was assessed by Western blot analysis.

Cytotoxic prenylated acylphloroglucinols from Hypericum annulatum.

A phytochemical investigation of the aerial parts of Hypericum annulatum Moris led to the isolation of five new prenylated acylphloroglucinol derivatives hyperannulatins A-E (1-3, 5 and 7) in addition to the known hypercalyxone A (4) and 3-geranyl-1-(2'-methylpropanoyl)phloroglucinol (6). The structures were determined by 1D and 2D NMR and MS spectroscopic techniques. Compounds 1 and 2 have in their structure evgenyl group, a rare hydrocarbon side chain. The cytotoxicity of isolated compounds was established on a panel of tumor cell lines (HL-60, HL-60/DOX, MDA-MB, SKW-3 and K-562) and was determined using MTT based assays. The compounds 1 and 2 showed to be the most potent cytotoxic agents, whose IC50 values against the chemosensitive cell lines ranged 3.42-5.87 µM and 1.48-8.21 µM, respectively. Noteworthy, albeit all tested compounds were less potent than podophyllotoxin their IC50 values were comparable to that of the other reference drug etoposide. In some of the cell lines compounds 1 and 2 even outclassed the cytotoxicity of etoposide.