The Mucus of Actinia equina (Anthozoa,Cnidaria): An Unexplored Resource for Potential Applicative Purposes

The mucus produced by many marine organisms is a complex mixture of proteins and polysaccharides forming a weak watery gel. It is essential for vital processes including locomotion, navigation, structural support, heterotrophic feeding and defence against a multitude of environmental stresses, predators, parasites, and pathogens. In the present study we focused on mucus produced by a benthic cnidarian, the sea anemone Actinia equina (Linnaeus, 1758) for preventing burial by excess sedimentation and for protection. We investigated some of the physico-chemical properties of this matrix such as viscosity, osmolarity, electrical conductivity, protein, carbohydrate, and total lipid contents. Some biological activities such as hemolytic, cytotoxic, and antibacterial lysozyme-like activities were also studied. The A. equina mucus is mainly composed by water (96.2% ± 0.3%), whereas its dry weight is made of 24.2% ± 1.3% proteins and 7.8% ± 0.2% carbohydrates, with the smallest and largest components referable to lipids (0.9%) and inorganic matter (67.1%). The A. equina mucus matrix exhibited hemolytic activity on rabbit erythrocytes, cytotoxic activity against the tumor cell line K562 (human erythromyeloblastoid leukemia) and antibacterial lysozyme-like activity. The findings from this study improve the available information on the mucus composition in invertebrates and have implications for future investigations related to exploitation of A. equina and other sea anemones’ mucus as a source of bioactive compounds of high pharmaceutical and biotechnological interest.     

Efficacy of different steam distribution systems against five soilborne pathogens under controlled laboratory conditions

The efficacy of three steam application techniques (steam injection, iron pan and sheet steaming) was evaluated against five soilborne pathogens under controlled laboratory conditions. Injection and pan steam systems proved to be efficient and feasible alternatives to traditional sheet steaming for suppressing Fusarium oxysporum f. sp. basilici at 60% moisture field capacity in sandy-loam soil. Injecting steam was the best technique to suppress F. oxysporum f. sp. basilici, F. oxysporum f. sp. raphani, F. oxysporum f. sp. conglutinans, Rhizoctonia solani and Phytophthora capsici. The mycelia of R. solani and P. capsici were very sensitive to heat and were effectively killed by injection of steam and by the pan steam system at 80% and 40% moisture field capacity.     

Cytotoxic and antioxidant activity of 4-methylthio-3-butenyl isothiocyanate from Raphanus sativus L. (Kaiware Daikon) sprouts

There is high current interest in the chemopreventive potential of Brassica vegetables (cruciferae), particularly due to their content in glucosinolates (GL), which upon myrosinase hydrolysis release the corresponding isythiocyanates (ITC). Some ITCs, such as sulforaphane (SFN) from broccoli ( Brassica oleacea italica), have been found to possess anticancer activity through induction of apoptosis in selected cell lines, as well as indirect antioxidant activity through induction of phase II detoxifying enzymes. Japanese daikon ( Raphanus sativus L.) is possibly the vegetable with the highest per capita consumption within the Brassicaceae family. Thanks to a recently improved gram scale production process, it was possible to prepare sufficient amounts of the GL glucoraphasatin (GRH) as well as the corresponding ITC 4-methylthio-3-butenyl isothiocyanate (GRH-ITC) from its sprouts. This paper reports a study on the cytotoxic and apoptotic activities of GRH-ITC compared with the oxidized counterpart 4-methylsulfinyl-3-butenyl isothiocyanate (GRE-ITC) on three human colon carcinoma cell lines (LoVo, HCT-116, and HT-29) together with a detailed kinetic investigation of the direct antioxidant/radical scavenging ability of GRH and GRH-ITC. Both GRH-ITC and GRE-ITC reduced cell proliferation in a dose-dependent manner and induced apoptosis in the three cancer cell lines. The compounds significantly ( p < 0.05) increased Bax and decreased Bcl2 protein expression, as well as producing caspase-9 and PARP-1 cleavage after 3 days of exposure in the three cancer cell lines. GRH-ITC treatment was shown to have no toxicity with regard to normal human lymphocytes (-15 +/- 5%) in comparison with SFN (complete growth inhibition). GRH and GRH-ITC were able to quench the 2,2-diphenyl-1-picrylhydrazyl radical, with second-order rate constants of 14.0 +/- 2.8 and 43.1 +/- 9.5 M(-1) s(-1), respectively (at 298 K in methanol), whereas the corresponding value measured here for the reference antioxidant alpha-tocopherol was 425 +/- 40 M (-1) s (-1). GRH reacted with H2O2 and tert-butyl hydroperoxide in water (pH 7.4) at 37 degrees C, with rate constants of 1.9 +/- 0.3 x 10(-2) and 9.5 +/- 0.3 x 10(-4) M(-1) s (-1) (paralleling recently developed synthetic antioxidants) being quantitatively (>97%) converted to GRE. It is demonstrated that GRH-ITC has interesting antioxidant/radical scavenging properties, associated with a selective cytotoxic/apoptotic activity toward three human colon carcinoma cell lines, and very limited toxicity on normal human T-lymphocytes.     

Combined effect of curing followed by acetic acid vapour treatments improves postharvest control of Penicillium digitatum on mandarins

In recent years several alternatives to the chemical control of postharvest decay have been examined but satisfactory levels of control, with a single system, have not been achieved yet. In the present study the results of an integrated postharvest approach are reported. Early and late harvested hybrid mandarin fruit “Fremont” and “Fairchild”, inoculated with Penicillium digitatum (Pers.:Fr.) Saccardo, were cured at 36 °C for 36 h with 95% RH and then fumigated with 0, 5, 15, 25, 50, 75 and 100 μL/L of acetic acid (AAC) vapours for 15 min. Following the treatments, fruit was stored at 20 °C and 80% RH to simulate a marketing period, and after 2 weeks the decay incidence and the visual appearance were evaluated. Curing or fumigations performed alone reduced decay with respect to untreated fruit, but the best control was achieved with combined treatments. For early harvested fruit the lowest decay percentage was obtained by using 75 μL/L with 8.3% and 2.1% of rots for “Fremont” and “Fairchild”, respectively, whereas for late harvested fruit the highest efficacy was observed using 50 μL/L (1.4% and 6.6%). Rind damage as pitting was observed only if fruit was treated with AAC alone at 100 μL/L.     

Occurrence of pipecolic acid and pipecolic acid betaine (homostachydrine) in Citrus genus plants

The presence of pipecolic acid and pipecolic acid betaine, also known as homostachydrine, is herein reported for the first time in Citrus genus plants. Homostachydrine was found in fruits, seeds, and leaves of orange, lemon, and bergamot (Citrus bergamia Risso et Poit). As homostachydrine was not commercially available, as a comparative source, extracts of alfalfa leaves ( Medicago sativa L.) were used, in which homostachydrine is present at high concentration. Then, the results where confirmed by comparison with an authentic standard synthesized and purified starting from pipecolic acid. The synthesized standard was characterized by a ESI-MS/MS study using a 3D ion-trap mass spectrometer. When subjected to MS/MS fragmentation in positive ion mode, homostachydrine, unlike its lower homologue proline betaine (also known as stachydrine), showed a pattern of numerous ionic fragments that allowed unambiguous identification of the compound. For the quantitation in the plant sources, high sensitivity and specificity were achieved by monitoring the transition (158 → 72), which is absent in the fragmentation patterns of other major osmolytes commonly used as markers for studies of abiotic stress. As for the metabolic origin of homostachydrine, the occurrence in citrus plants of pipecolic acid leads to the hypothesis that it could act as a homostachydrine precursor through direct methylation.     

Seed transmission of Plectosphaerella cucumerina, causal agent of leaf spot of Diplotaxis tenuifolia in Italy

Plectosphaerella cucumerina has been described recently as the causal agent of a leaf spot on wild rocket (Diplotaxis tenuifolia). Eight seed samples of wild rocket obtained from commercial seed lots used for sowing by farms severely affected by P. cucumerina, were assayed for the presence of the pathogen. Isolations were carried out on subsamples of seeds (400) unwashed or disinfected in 1% sodium hypochloride. The pathogenicity of the isolates of P. cucumerina obtained was tested in two trials carried out on wild rocket; four out of eight samples of rocket seeds were contaminated by P. cucumerina. Eleven isolates of P. cucumerina were obtained from 7,200 not disinfected seeds tested, while none was isolated from an equal number of disinfected seeds. All isolates were pathogenic on wild rocket. The results obtained indicate that rocket seeds are a potential source of inoculum for P. cucumerina. The possibility of isolating the pathogen from seeds, albeit from a low percentage of them, supports the hypothesis that the rapid spread of this new disease of rocket recently observed in Italy is due to the use of infected propagation material. Measures for prevention and control of the disease are discussed.     

The relevance of marine chemical ecology to plankton and ecosystem function: an emerging field

Marine chemical ecology comprises the study of the production and interaction of bioactive molecules affecting organism behavior and function. Here we focus on bioactive compounds and interactions associated with phytoplankton, particularly bloom-forming diatoms, prymnesiophytes and dinoflagellates. Planktonic bioactive metabolites are structurally and functionally diverse and some may have multiple simultaneous functions including roles in chemical defense (antipredator, allelopathic and antibacterial compounds), and/or cell-to-cell signaling (e.g., polyunsaturated aldehydes (PUAs) of diatoms). Among inducible chemical defenses in response to grazing, there is high species-specific variability in the effects on grazers, ranging from severe physical incapacitation and/or death to no apparent physiological response, depending on predator susceptibility and detoxification capability. Most bioactive compounds are present in very low concentrations, in both the producing organism and the surrounding aqueous medium. Furthermore, bioactivity may be subject to synergistic interactions with other natural and anthropogenic environmental toxicants. Most, if not all phycotoxins are classic secondary metabolites, but many other bioactive metabolites are simple molecules derived from primary metabolism (e.g., PUAs in diatoms, dimethylsulfoniopropionate (DMSP) in prymnesiophytes). Producing cells do not seem to suffer physiological impact due to their synthesis. Functional genome sequence data and gene expression analysis will provide insights into regulatory and metabolic pathways in producer organisms, as well as identification of mechanisms of action in target organisms. Understanding chemical ecological responses to environmental triggers and chemically-mediated species interactions will help define crucial chemical and molecular processes that help maintain biodiversity and ecosystem functionality.     

Analytical discrimination of poisonous and nonpoisonous chemotypes of giant fennel (Ferula communis L.) through their biologically active and volatile fractions

Giant fennel (Ferula communis L.) from Sardinia is characterized by two chemotypes with different biological activities. One chemotype is poisonous, due to prenylcoumarins, and responsible for ferulosis, which mainly affects sheep and goats, cattle, and horses; the other chemotype is nonpoisonous and contains daucane esters. The two chemotypes cannot be distinguished botanically. High-performance liquid chromatography-diode array-ultraviolet detection-mass spectrometry (HPLC-DAD-UV-MS) analysis of the composition of the fractions containing the biologically active metabolites and of the volatile fractions, by gas chromatography-mass spectrometry (GC-MS), of both essential oil and headspace sampled by headspace solid-phase microextraction (HS-SPME) are here shown to be effective in discriminating the poisonous and nonpoisonous chemotypes. HS-SPME with CAR/PDMS/DVB in combination with GC-MS has also been found to be a successful, fully automated one-step method for rapid and unequivocal discrimination of the two chemotypes, using aristolene and allohedycaryol as markers of the poisonous and nonpoisonous chemotypes, respectively.