Ameliorative effect of polyamines on the high temperature inhibition of in vitro pollen germination in tomato (Lycopersicon esculentum Mill.)

Effect of polyamines on in vitro pollen germination at high temperatures in tomato (Lycopersicon esculentum Mill.) was investigated. Pollen germination and tube growth were significantly inhibited at 33°C and 35°C compared to those at 25°C. This inhibition was reversed by the addition of spermidine or spermine in the germination medium. Spermidine at 0.5 mM was slightly more effective than spermine at 0.05 mM. Spermidine at 0.05 and 0.5 mM and spermine at 0.05 mM slightly increased pollen germination rates at 25°C. Spermidine at 5 mM and spermine at 0.5 M were inhibitory to pollen germination, regardless of incubation temperatures. Spermidine also promoted germination of pollen grains incubated at 38°C for 1 and 3 h and then at 25°C for the rest of the 20 h incubation period. The effect was higher at 0.5 mM than at 0.05 mM. Treatment of spermidine to intact flowers 1 day before anthesis was also effective in ameliorating the high temperature inhibition of in vitro pollen germination on the polyamine-free medium. Here, the optimum concentration was 5 mM. These results demonstrate that polyamines can counteract the inhibitory effects of high temperature on pollen germination. They also suggest that the endogenous level of polyamines in germinating pollen grains is an important factor for the pollen germinability at high temperature.     

Influence of aluminum on the membranes of mycorrhizal fungi

The influence of Al³⁺ on the membrane fluidity of the mycorrhizal fungus Lactarius piperatus (L.ex.Fr.) S. F. Gray was studied. When mycelia grew on media supplemented with 1 or 10 mM aluminum for 21 days, a significant increase in fluidity was detected, compared with the corresponding controls. The comparisons of the EPR spectra calculated by a model, with the experimental ones showed that the spectra are superimpositions of two components with different membrane ordering. The action of aluminum induced a relative enlargement of the less immobilized portion in the membrane. No significant alterations of fluidity were measured after 30-minute treatment, with Al³⁺.     

Solanidine hydrolytic extraction and separation from the potato (Solanum tuberosum L.) vines by using solid-liquid-liquid systems

Solanidine is a steroidal aglycon of potato (Solanum tuberosum L.) glycoalkaloids and a very important precursor for the synthesis of hormones and some pharmacologically active compounds. Glycoalkaloids are hydrolyzed by mineral acid, yielding solanidine. This paper deals with the kinetics of solanidine hydrolytic extraction in different solid-liquid-liquid systems. The dried and milled potato (S. tuberosum L.) vines were used as a source of glycoalkaloids and as the solid phase. The solutions of hydrochloric acid in 2 and 10% (w/v) aqueous acetic acid, in 50% (volume) aqueous methanol, and in 50% (volume) aqueous ethanol were first liquid phase, and the medium for glycoalkaloid extraction from potato vines and their hydrolysis to solanidine. The chloroform, trichloroethylene, or carbon tetrachloride were the second, organic, liquid phase and the medium for solanidine extraction. This procedure combines three different processes: extraction of glycoalkaloids from potato vines, their hydrolysis to solanidine, and the extraction of solanidine, in a single step. The term hydrolytic extraction of solanidine was used for these processes. The purpose of the paper was to choose an optimal solid-liquid-liquid system for solanidine extraction and to define the procedure for its isolation from the organic liquid phase. The best degree of solanidine hydrolytic extraction (DHE) of more than 98% was achieved when 10% (w/v) hydrochloric acid in 50% (volume) methanol were the first liquid phase and chloroform was the second liquid phase, after 90 min. The yield of solanidine (q(S)) under these conditions is calculated to be 0.24 g/100 g of potato vines. Approximately 78% of the maximal possible yield of solanidine was isolated from chlorofom liquid phase. The IR and MS spectra of isolated solanidine were recorded.     

High-performance liquid chromatographic determination of biogenic amines in poultry carcasses

Biogenic amines, produced by bacterial decarboxylation of amino acids, have been associated with toxicological symptoms in broilers fed various poultry byproducts. A reversed-phase high-performance liquid chromatographic method is described for the quantitation of eight biogenic amines (tryptamine, phenylethylamine, putrescine, cadaverine, histamine, tyramine, spermidine, and spermine) in chicken carcasses. Amines were extracted with perchloric acid, derivatized with dansyl chloride, separated using gradient elution (methanol and water), and detected by fluorescence. Benzylamine was used as the internal standard. Linearity, repeatability, and recovery of the method were evaluated. The method was linear for all of the amines studied at concentrations ranging from 0.05 to 25 microg/mL. Average recoveries ranged from 92.6% to 96.8% for all amines except for histamine, which was 74.6%.     

Role of polyamines in peach fruit development and storage

To investigate the role of polyamines in pre- and post-harvest fruit development of 'Akatsuki' peach (Prunus persica (L.) Batsch.) we measured polyamine concentrations, activities of polyamine biosynthetic enzymes and expression of genes encoding these enzymes. Concentrations of the free polyamines, putrescine (Put), spermidine (Spd) and spermine (Spm) in pre-harvest fruit peaked 16 days after full bloom (DAF) and then progressively decreased until harvest with the exception of Put, which showed a second peak at 94 DAF, just before the onset of ethylene production. In post-harvest fruit, minor changes in concentrations of Spd and Spm were observed, whereas Put concentration peaked on the harvest day, followed by an abrupt decrease and a subsequent 2-fold increase, which was opposite to the fluctuating pattern of ethylene production. Activities of arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) peaked during the first stage of fruit development and then decreased until 80 DAF, after which the activities were below detection limits, suggesting that Put is synthesized during the early stage of fruit development. Activity of S-adenosylmethionine decarboxylase (SAMDC) decreased progressively until the end of S2. Expression levels of five putative polyamine biosynthetic genes, ADC, ODC, SAMDC, spermidine synthase (SPDS) and spermine synthase (SPMS), in pre-harvest and post-harvest fruit did not coincide precisely with the observed changes in enzymatic activities and polyamine concentrations. The possible role of polyamines during peach fruit development and the relationship between polyamines and ethylene biosynthesis are discussed.     

Antioxidant activity of Filipendula hexapetala flowers

The antioxidant activity of the methanol extract of Filipendula hexapetala flowers was assessed by the assay for ferric-reducing antioxidant power (FRAP), the assay for DPPH free radical scavenging ability (DPPH) and the assay for the influence of lipid peroxidation in liposomes, induced by Fe(2+)/ascorbate system and measured by the TBA test (LP). The activity of the investigated extract in all test-systems was found to be significant. The principal constituent responsible for the observed effects was isolated and identified as spiraeoside.     

Cyclodepsipeptides: Isolation from Endophytic Fungi of Sarcophyton ehrenbergi and Verification of Their Larvicidal Activity via In-Vitro and In-Silico Studies

Culex pipiens mosquitoes are vectors to many viruses and can transmit diseases such as filariasis and avian malaria. The present study evaluated the larvicidal activity of marine-derived endophytic fungi Aspergillus nomius and Aspergillus flavus from the soft coral Sarcophyton ehrenbergi along with two known cyclodepsipeptide compounds, scopularide A (1) and B (2), isolated from A. flavus extract, against third-instar larvae of C. pipiens, using distilled water as a negative control and toosenedanin as a positive control. The structures of the isolated compounds were confirmed by various spectroscopic analyses. The lethal concentrations (LC₅₀ and LC₉₀) were calculated by probit analysis. Scopularide A was the most potent after 96 h treatment, with LC₅₀ and LC₉₀ values of 58.96 and 994.31 ppm, respectively, and with 82.66% mortality at a concentration of 300 ppm. To unravel the biochemical mechanism of the tested extracts and compounds, their effects against protease, chitinase, phenoloxidases and lipase enzymes from the whole-body tissue of C. pipiens were evaluated after 72 h treatment at LC₅₀ dose. Superior activity was observed for A. flavus extract against all tested enzymes. A molecular docking study was conducted for scopularide A and B on the four tested enzymes, to further verify the observed activity. Results revealed good binding affinities for both compounds as compared to the docked ligands, mainly via a number of hydrogen bonds. This was the first study to report the isolation of endophytic fungi A. flavus and A. nomius from the marine soft coral S. ehrenbergi. The endophytic fungal extract of A. flavus was found to be a promising source for a natural larvicidal agent against C. pipiens populations.     

Establishment of feather follicle stem cells as potential vehicles for delivering exogenous genes in birds

The present study was performed to develop a culture system for feather keratinocyte stem cells to enable the genetic manipulation of endangered avian species. The feather follicle cells were isolated from growing feathers of adult White Leghorn chicken. Leukemia inhibitory factor (LIF) was used to maintain the characterization of the keratinocyte colony-forming cells (KCFCs). The EGFPN1 plasmid DNA retroviral vector was used to deliver Green Fluorescent Protein (GFP) gene, which was introduced to the KCFCs by lipofection. After removal of the fibroblast-like cells, the feather KCFCs attached to the substrate within 24 h of seeding. The cells continued to proliferate for at least 30 days in the presence of LIF. The cell-adhesion molecules such as integrin beta1 and CD49c were immunocytochemically positive in the cells. The KCFCs differentiated into barbular cells and pennaceous feather vane in the LIF-free medium. The GFP gene-transfected KCFCs stably expressed GFP. The present results indicate that the KCFCs derived from feather follicles are closely related to multipotent stem cells. In addition, gene manipulation of such stem cells may be useful for the production of chimera in avian species.