Assessment of Lemna gibba L. (duckweed) as a potential ecological indicator for contaminated aquatic ecosystem by boron mine effluent

Duckweeds, as a group, are important early warning indicators for the assessment of contaminated ecosystems due to their propensity to accumulate pollutants. In the present study, we investigated the potential use of Lemna gibba L. (Lemnaceae) as an ecological indicator for boron (B) mine effluent containing B concentration above 10 mg l⁻¹. For this purpose, L. gibba fronds were grown for 7 days in simulated water contaminated with B mine effluent. The important note is that this study was carried out in Kırka (Eskişehir, Turkey) B reserve area, which is the largest borax reserve in all over the world, under natural climatic conditions in the field. The results demonstrated that accumulations of B by L. gibba gradually increased based on the initial B concentrations (10, 25, 50, 100, and 150 mg l⁻¹) of the mine effluent. B concentration in the dry weight of the plant reached 639 mg kg⁻¹ when the minimum initial dosage (10 mg l⁻¹) was applied and 2711 mg kg⁻¹ when the maximum initial dosage (150 mg l⁻¹) was applied during the study. However, significant reductions in their relative growth rates occurred in 50, 100 and 150 mg l⁻¹ initial B concentrations. Results suggest that 25 mg l⁻¹ B concentration in water seemed to be a sensitive endpoint for L. gibba that could be used as a critical bioindicator level of B contaminated water. Following our data, we also constructed a simple growth model under the climatic conditions in this region of Turkey, but in instructive as a worldwide model. L. gibba is, therefore, suggested to be able to use as both an indicator and a phytoremediation tool because of its high accumulation capacity for B contaminated water.     

Thalassia testudinum response to the interactive stressors hypersalinity,sulfide and hypoxia

A large-scale mesocosm (sixteen 500 L tanks) experiment was conducted to investigate the effects of hypersalinity (45–65 psu), porewater sulfide (2–6 mM) and nighttime water column hypoxia (5–3 mg L⁻¹) on the tropical seagrass Thalassia testudinum Banks ex König. We examined stressor effects on growth, shoot survival, tissue sulfur (S⁰, TS, δ³⁴S) and leaf quantum efficiencies, as well as, porewater sulfides (∑TS_pw) and mesocosm water column O₂ dynamics. Sulfide was injected into intact seagrass cores of T. testudinum exposing below-ground tissues to 2, 4, and 6 mM S²⁻, but rapid oxidation resulted in ∑TS_pw < 1.5 mM. Hypersalinity at 65 psu lowered sulfide oxidation and significantly affected plant growth rates and quantum efficiencies (F_v/F_m < 0.70). The most depleted rhizome δ³⁴S signatures were also observed at 65 psu, suggesting increased sulfide exposure. Hypoxia did not influence ∑TS_pw and plant growth, but strengthened the hypersalinity response and decreased rhizome S⁰, indicating less efficient oxidation of ∑TS_pw. Following nighttime hypoxia treatments, ecosystem level metabolism responded to salinity treatments. When O₂ levels were reduced to 5 and 4 mg L⁻¹, daytime O₂ levels recovered to approximately 6 mg L⁻¹; however, this recovery was more limited when O₂ levels were lowered to 3 mg L⁻¹. Subsequent to O₂ reductions to 3 mg O₂ L⁻¹, nighttime O₂ levels rose in the 35 and 45 psu tanks, stayed the same in the 55 psu tanks, and declined in the 65 psu tanks. Thus, hypersalinity at 65 psu affects T. testudinum's oxidizing capacity and places subtle demands on the positive O₂ balance at an ecosystem level. This O₂ demand may influence T. testudinum die-off events, particularly after periods of high temperature and salinity. We hypothesize that the interaction between hypersalinity and sulfide toxicity in T. testudinum is their synergistic effect on the critical O₂ balance of the plant.     

Microbial processes and bacterial populations associated to anaerobic treatment of sulfate-rich wastewater

A pilot-scale (1.2 m³) anaerobic sequencing batch biofilm reactor (ASBBR) containing mineral coal for biomass attachment was fed with sulfate-rich wastewater at increasing sulfate concentrations. Ethanol was used as the main organic source. Tested COD/sulfate ratios were of 1.8 and 1.5 for sulfate loading rates of 0.65–1.90 kgSO₄²⁻/cycle (48 h-cycle) or of 1.0 in the trial with 3.0 gSO₄²⁻ l⁻¹. Sulfate removal efficiencies observed in all trials were as high as 99%. Molecular inventories indicated a shift on the microbial composition and a decrease on species diversity with the increase of sulfate concentration. Beta-proteobacteria species affiliated with Aminomonas spp. and Thermanaerovibrio spp. predominated at 1.0 gSO₄²⁻ l⁻¹. At higher sulfate concentrations the predominant bacterial group was Delta-proteobacteria mainly Desulfovibrio spp. and Desulfomicrobium spp. at 2.0 gSO₄²⁻ l⁻¹, whereas Desulfurella spp. and Coprothermobacter spp. predominated at 3.0 gSO₄²⁻ l⁻¹. These organisms have been commonly associated with sulfate reduction producing acetate, sulfide and sulfur. Methanogenic archaea (Methanosaeta spp.) was found at 1.0 and 2.0 gSO₄²⁻ l⁻¹. Additionally, a simplified mathematical model was used to infer on metabolic pathways of the biomass involved in sulfate reduction.     

Freeze tolerance and accumulation of cryoprotectants in the enchytraeid Enchytraeus albidus (Oligochaeta) from Greenland and Europe

The freeze tolerance and accumulation of cryoprotectants was investigated in three geographically different populations of the enchytraeid Enchytraeus albidus (Oligochaeta). E. albidus is widely distributed from the high Arctic to temperate Western Europe. Our results show that E. albidus is freeze tolerant, with freeze tolerance varying extensively between Greenlandic and European populations. Two populations from sub Arctic (Nuuk) and high Arctic Greenland (Zackenberg) survived freezing at −15 °C, whereas only 30% of a German population survived this temperature. When frozen, E. albidus responded by catabolising glycogen to glucose, which likely acted as a cryoprotectant. The average glucose concentrations were similar in the three populations when worms were frozen at −2 °C, approximately 50 μg glucose mg⁻¹ tissue dry weight (DW). At −14 °C the glucose concentrations increased to between 110 and 170 μg mg⁻¹ DW in worms from Greenland. The average glycogen content of worms from Zackenberg and Nuuk were about 300 μg mg⁻¹ DW, but only 230 μg mg⁻¹ DW in worms from Germany showing that not all glycogen was catabolised during the experiment. Nuclear magnetic resonance spectrometry (NMR) was used to screen for other putative cryoprotectants. Proline, glutamine and alanine were up regulated in frozen worms at −2 °C but only in relatively small concentrations suggesting that they were of little significance for freeze survival. The present study confirms earlier reports that freeze tolerant enchytraeids, like other freeze tolerant oligochaete earthworms, accumulate high concentrations of glucose as a primary cryoprotectant.     

Protocorm culture of Dendrobium candidum in balloon type bubble bioreactors

Protocorm cultures of Dendrobium candidum were established in balloon type bubble bioreactors using Murashige and Skoog (MS) medium with 0.5 mg l⁻¹ α-naphthaleneacetic acid (NAA), 2.5% (w/v) sucrose, 5:25 mM NH₄:NO₃ and 1% (v/v) banana homogenate for the production of biomass and bioactive compounds. In 3 l bioreactor containing 2 l medium, a maximum protocorm biomass (21.0 g l⁻¹ dry biomass) and also optimum quantities of total polysaccharides (389.3 mg g⁻¹ DW), coumarins (18.0 mg g⁻¹ DW), polyphenolics (11.9 mg g⁻¹ DW), and flavonoids (4.5 mg g⁻¹ DW) were achieved after 7 weeks of culture. Based on these studies, 5 and 10 l bioreactor cultures were established to harvest 80 g and 160 g dry biomass. In 10 l bioreactors, the protocorms grown were accumulated with optimal levels of polysaccharides (424.1 mg g⁻¹ DW), coumarins (15.8 mg g⁻¹ DW), polyphenols (9.03 mg g⁻¹ DW) and flavonoids (4.7 mg g⁻¹ DW). The bioreactor technology developed here will be useful for the production of important bioactive compounds from D. candidum.     

CO2 fixation for succinic acid production by engineered Escherichia coli co-expressing pyruvate carboxylase and nicotinic acid phosphoribosyltransferase

In wild-type Escherichia coli, 1 mol of CO₂ was fixated in 1 mol of succinic acid generation anaerobically. The key reaction in this sequence, catalyzed by phosphoenolpyruvate carboxylase (PPC), is carboxylation of phosphoenolpyruvate to oxaloacetate. Although inactivation of pyruvate formate-lyase and lactate dehydrogenase is found to enhance the PPC pathway for succinic acid production, it results in excessive pyruvic acid accumulation and limits regeneration of NAD⁺ from NADH formed in glycolysis. In other organisms, oxaloacetate is synthesized by carboxylation of pyruvic acid by pyruvate carboxylase (PYC) during glucose metabolism, and in E. coli, nicotinic acid phosphoribosyltransferase (NAPRTase) is a rate-limiting enzyme of the NAD(H) synthesis system. To achieve the NADH/NAD⁺ ratio decrease as well as carbon flux redistribution, co-expression of NAPRTase and PYC in a pflB, ldhA, and ppc deletion strain resulted in a significant increase in cell mass and succinic acid production under anaerobic conditions. After 72 h, 14.5 g L⁻¹ of glucose was consumed to generate 12.08 g L⁻¹ of succinic acid. Furthermore, under optimized condition of CO₂ supply, the succinic acid productivity and the CO₂ fixation rate reached 223.88 mg L⁻¹ h⁻¹ and 83.48 mg L⁻¹ h⁻¹, respectively.     

A high pressure liquid chromatography-based assay for glutathione-S-transferase class distinction assay

In order to expedite the process of classification of the members of the family of glutathione-S-transferases (GSTs) high performance liquid chromatography with photodiode array detection (HPLC-PDA) was used as a means for measuring enzymatic activity. The GST chosen for the development of the HPLC-PDA technique was from equine liver (E-GST). The characterizing substrates, ethacrynic acid (EA) and bromosulfophthalein (BSP), along with previously gathered characterization data allowed for the distinction of α, μ or π-class enzymes. In an initial characterization of the previously unclassified E-GST it was determined that the enzyme was of the π-class with specific activities of 0.062, ± 0.0015 μmol min^− 1 mg^− 1 and 0.0019, ± 0.00064 μmol min^− 1 mg^− 1 for EA and BSP, respectively. Finally, the activity of the E-GST with the EA and BSP substrates, was measured by HPLC-PDA, and was found to be 0.027, ± 0.003 μmol min^− 1 mg^− 1 and 0.002, ± 0.0005 μmol min^− 1 mg^− 1, respectively. While the HPLC-PDA data do not mirror the spectrophotometric results quantitatively the overall response by the E-GST was the same. In general, the E-GSTs were shown to belong to the π-class when characterized by HPLC-PDA due to an EA specific activity greater than 0.01 μmol min^− 1 mg^− 1 and a negligible BSP activity (≤ 0.002 μmol min^− 1 mg^− 1).     

Treatment of dye-rich wastewater by an immobilized thermophilic cyanobacterial strain: Phormidium sp.

The removal of Remazol Blue and Reactive Black B by the immobilized thermophilic cyanobacterial strain Phormidium sp. was investigated under thermophilic conditions in a batch system, in order to determine the optimal conditions required for the highest dye removal. In the experiments, performed at pH 8.5, with different initial dye concentrations between 9.1 mg l⁻¹ and 82.1 mg l⁻¹ and at 45 °C, calcium alginate immobilized Phormidium sp. showed high dye decolorization, with maximum uptake yields ranging from 50% to 88% at all dye concentrations tested. When the effects of high dye concentrations on dye removal were investigated, the highest uptake yield in the beads was 50.3% for 82.1 mg l⁻¹ Remazol Blue and 60.0% for 79.5 mg l⁻¹ Reactive Black B. The highest color removal was detected at 45 °C and 50 °C incubation temperatures for all dye concentrations. As the temperature decreased, the removal yield of immobilized Phormidium sp. also decreased. At about 75 mg l⁻¹ initial dye concentrations, the highest specific dye uptake measured was 41.29–41.17 mg g⁻¹ for Remazol Blue and 47.69–43.82 mg g⁻¹ for Reactive Black B at 45 °C and 50 °C incubation temperatures, respectively, after 8 days incubation.     

Chromate reduction by resting cells of Achromobacter sp. Ch-1 under aerobic conditions

Chromate reduction was carried out by resting cells of Achromobacter sp. Ch-1 with lactate as electron donor under aerobic conditions. The reduction activity of the samples supplemented with lactate was two times as those without lactate. The reduction rate was influenced by initial pH and lactate concentration. Under the optimal conditions, pH 9.0 and 4000 mg l⁻¹ lactate supplement, reduction rate was 5.45 mg l⁻¹ min⁻¹. The reduction rate decreased with increasing of Cr(VI) concentrations and increased with cell densities proportionally. The maximum reduction limit of Ch-1 cells was obtained at 2107 mg l⁻¹ of Cr(VI).     

Operation of partial nitrification to nitrite of landfill leachate and its performance with respect to different oxygen conditions

The coupled system of partial nitrification and anaerobic ammonium oxidation (Anammox) is efficient in nitrogen removal from wastewater. In this study, the effect of different oxygen concentrations on partial nitrification performance with a sequencing batch reactor (SBR) was investigated. Results indicate that, partial nitrification of landfill leachate could be successfully achieved under the 1.0–2.0 mg L⁻¹ dissolved oxygen (DO) condition after 118 d long-term operation, and that the effluent is suitable for an Anammox reactor. Further decreasing or increasing the DO concentration, however, would lead to a decay of nitrification performance. Additionally, the MLSS concentration in the reactor increased with increasing DO concentration. Respirometric assays suggest that low DO conditions (<2 mg L⁻¹) favor the ammonia-oxidizing bacteria (AOB) and significantly inhibit nitrite oxidizing bacteria (NOB) and aerobic heterotrophic bacteria (AHB); whereas high DO conditions (>3 mg L⁻¹) allow AHB to dominate and significantly inhibit AOB. Therefore, the optimal condition for partial nitrification of landfill leachate is 1.0–2.0 mg L⁻¹ DO concentration.     

Dual purpose system that treats anaerobic effluents from pig waste and produce Neochloris oleoabundans as lipid rich biomass

Dual purpose systems that treat wastewater and produce lipid rich microalgae biomass have been indicated as an option with great potential for production of biodiesel at a competitive cost. The aim of the present work was to develop a dual purpose system for the treatment of the anaerobic effluents from pig waste utilizing Neochloris oleoabundans and to evaluate its growth, lipid content and lipid profile of the harvested biomass and the removal of nutrients from the media. Cultures of N. oleoabundans were established in 4 L flat plate photobioreactors using diluted effluents from two different types of anaerobic filters, one packed with ceramic material (D1) and another one packed with volcanic gravel (D2). Maximum biomass concentration in D1 was 0.63 g L⁻¹ which was significantly higher than the one found in D2 (0.55 g L⁻¹). Cultures were very efficient at nutrient removal: 98% for NNH₄⁺ and 98% for PO₄³⁻. Regarding total lipid content, diluted eflluents from D2 promoted a biomass containing 27.4% (dry weight) and D1 a biomass containing 22.4% (dry weight). Maximum lipid productivity was also higher in D2 compared to D1 (6.27 ± 0.62 mg L⁻¹ d⁻¹ vs. 5.12 ± 0.12 mg L⁻¹ d⁻¹). Concerning the FAMEs profile in diluted effluents, the most abundant one was C18:1, followed by C18:2 and C16:0. The profile in D2 contained less C18:3 (linolenic acid) than the one in D1 (4.37% vs. 5.55%). In conclusion, this is the first report demonstrating that cultures of N. oleoabundans treating anaerobic effluents from pig waste are very efficient at nutrient removal and a biomass rich in lipids can be recovered. The maximum total lipid content and the most convenient FAMEs profile were obtained using effluents from a digester packed with volcanic gravel.     

Mathematical analysis of trickling filter response to pentachlorophenol shock load

The response of a laboratory trickling filter to a step increase in pentachlorophenol (PCP) feed concentration was analyzed using continuous stirred tank (CSTR) and plug flow reactor (PFR) models. The CSTR model provided a slightly better fit to experimental data than the plug flow model when specific growth rate, μ, and PCP-degrading biomass concentration before the shock load, X₀, were variable parameters but was clearly superior when the mean residence time, τ, was added as a third parameter. The three-parameter CSTR model accurately represented six of seven concentration response curves corresponding to step increases in PCP feed concentration of 12–165 mg l⁻¹ and 20–150 mg l⁻¹. The continuing improvement in system response to repetitive 20–150 mg l⁻¹ shock loads was reflected by a monotonic increase in the optimal estimates of initial rate of biomass production.     

Decolorization of azo dye by immobilized Pseudomonas luteola entrapped in alginate–silicate sol–gel beads

Pseudomonas luteola was immobilized by entrapment in alginate–silicate sol–gel beads for decolorization of the azo dye, Reactive Red 22. The influences of biomass loading and operating conditions on specific decolorization rate and dye removal efficiency were studied in details. The immobilized cells were found to be less sensitive to changes in agitation rates (dissolved oxygen levels) and pH values. Michaelis–Menten kinetics could be used to describe the decolorization kinetics with the kinetic parameters being 36.5 mg g⁻¹ h⁻¹, 300.1 mg l⁻¹ and 18.2 mg g⁻¹ h⁻¹, 449.8 mg l⁻¹ for free and immobilized cells, respectively. After five repeated batch cycles, the decolorization rate of the free cells decreased by nearly 54%, while immobilized cells still retained 82% of their original activity. The immobilized cells exhibited better thermal stability during storage and reaction when compared with free cells. From SEM observation, a dense silicate gel layer was found to surround the macroporous alginate–silicate core, which resulted in much improved mechanical stability over that of alginate beads when tested under shaking conditions. Alginate–silicate matrices appeared to be the best matrix for immobilization of P. luteola in decolorization of Reactive Red 22 when compared with previous results using synthetic or natural polymer matrices.     

Promotion of Salvia miltiorrhiza hairy root growth and tanshinone production by polysaccharide–protein fractions of plant growth-promoting rhizobacterium Bacillus cereus

This study was to examine the effects of polysaccharides from a plant growth-promoting rhizobacterium (PGPR) Bacillus cereus on the growth and tanshinone production of Salvia miltiorrhiza hairy roots. A polysaccharide fraction designated BPS was isolated from the hot water extract of B. cereus cells by ethanol precipitation. BPS applied to the root culture at 100–400 mg l⁻¹ a few days before the stationary growth phase stimulated the tanshinone accumulation of roots by about 7-fold (1.59 mg g⁻¹ versus 0.19 mg g⁻¹) and also notably promoted the root growth (15% increase in biomass). BPS was a polysaccharide–protein complex containing about 27% protein, which is essential for root growth promotion. BPS was separated by ultrafiltration into two molecular weight (MW) fractions, of which the high MW fraction (∼35.8 kDa) with higher protein content (∼31%) promoted the root growth while the lower MW fraction with lower protein content (∼17%) suppressed the growth. The results suggest that the polysaccharide portion of BPS was responsible for stimulating the tanshinone accumulation while the protein portion was responsible for promoting the hairy root growth. Polysaccharides from PGPR are potential sources of active elicitors and growth-promoting agents for plant roots in culture.     

Decolorization of Reactive Red 195 by a mixed culture in an alternating anaerobic–aerobic Sequencing Batch Reactor

The mono-azo dye Reactive Red 195 (RR 195) is a widely used color compound in the textile industry. As many other colors, it is persistent and difficult to be removed from water with conventional processes. The present study investigates biological decolorization of RR 195 under alternate anaerobic–aerobic conditions in a laboratory scale Sequencing Batch Reactor (SBR) containing a mixed culture and fed with a biodegradable carbon source. Different values of the Sludge Retention Time (SRT), Hydraulic Retention Time (HRT), influent color and organic carbon loadings were adopted during the experimental activity and their effects on color and Chemical Oxygen Demand (COD) removal efficiencies and process kinetics determined. The optimal operating conditions were found to be: 800 mg l⁻¹ influent COD, 50 d SRT and a 24 h-cycle. Under these conditions, the maximum color efficiency of 97% was achieved for a 40 mg l⁻¹ RR 195 in the feed. Some inhibition was present at influent color loadings above 40 mg l⁻¹, which was confirmed by the application of the Haldane model.     

Oxidation of hydrogen sulphide in sour gas by Chlorobium limicola

The aim of this work was to assess the potential for bacterial oxidation of hydrogen sulphide as a purification method of sour gas. Using a continuous culture of Chlorobium limicola, high efficiencies of oxidation of both soluble and gaseous sulphide were achieved, with efficiencies for the latter exceeding 95%. Sulphide added as aqueous sodium sulphide was converted to sulphur and sulphate with almost total removal of the initial 100 mg S l⁻¹ within 24 h. Gaseous sulphide was oxidized at an efficiency of 95% (approximately 3 mmol S h⁻¹ (unit biomass Abs)⁻¹) over 1 h runs at a gas flow rate of 60 ml min⁻¹. With a sulphur recovery system to prevent sulphur accumulation, an efficiency of 70% was maintained. Biological removal of sulphide represents a potentially important biotechnological process, with high potential for viable scale up.     

Influence on dithiolopyrrolone antibiotic production by organic acids in Saccharothrix algeriensis NRRL B-24137

The influence of organic acids on growth and dithiolopyrrolone antibiotic production by Saccharothrix algeriensis NRRL B-24137 was studied. The production of dithiolopyrrolones depends upon the nature and concentration of the organic acids in the culture medium. Study of the nature of organic acids showed that the most effective organic acids for thiolutin specific production were maleic, 4-hydroxybenzoic, benzentetracarboxylic, pantothenic, pivalic and pyruvic acids (which yielded almost five-fold over the starting medium) and pimelic acid (more than three-fold). 4-Bromobenzoic acid showed the best production of senecioyl-pyrrothine (59 mg g⁻¹ DCW). Tiglic acid showed the best production of tigloyl-pyrrothine (22 mg g⁻¹ DCW). The highest yield of isobutyryl-pyrrothine (7.6 mg g⁻¹ DCW) was observed in the presence of crotonic acid. Sorbic acid yielded the best production of butanoyl-pyrrothine (26 mg g⁻¹ DCW). Methacrylic, butyric, pyruvic and 4-bromobenzoic acids also exhibited the best production of butanoyl-pyrrothine (27–11-fold).Study of organic acid concentration showed that among the selected organic acids, pimelic acid yielded the highest specific production of thiolutin (91 mg g⁻¹ DCW) at 7.5 mM; and senecioyl-pyrrothine (11 mg g⁻¹ DCW), tigloyl-pyrrothine (9 mg g⁻¹ DCW) and butanoyl-pyrrothine (3.5 mg g⁻¹ DCW) at 5 mM. Pyruvic acid at 1.25 mM enhanced the production of senecioyl-pyrrothine (4.3 mg g⁻¹ DCW). The maximum production of tigloyl-pyrrothine (18.6 mg g⁻¹ DCW) was observed in the presence of tiglic acid at 2.5 mM. Maximum production of isobutyryl-pyrrothine was observed in the presence of 7.5 mM tiglic acid. In addition, methacrylic acid (at 5 mM) and butyric acid (at 2.5 mM) enhanced the production of butanoyl-pyrrothine (26 and 20 times, respectively).The above results can be employed in the optimisation of the culture medium for the production of dithiolopyrrolone in higher quantities.     

Multiple regeneration pathways in Spathoglottis plicata Blume — A study in vitro

Asymbiotic germination of immature seeds (embryos), and mature seeds and micropropagation of Spathoglottis plicata were described. Effects of three nutrition media namely, Murashige & Skoog (MS); Phytamax (PM); and Phyto-Technology orchid seed sowing medium (P₇₂₃), two carbon sources such as glucose and sucrose at 2–3% (w/v), two plant growth regulators such as 6-benzylaminopurine (BAP; 0.5–3.0 mg l^− 1) and α-naphthalene acetic acid (NAA; 0.5–2.0 mg l^− 1) and peptone (2.0 g l^− 1) were examined on seed germination, early protocorm development and micropropagation. The maximum germination of mature seeds (95%) was recorded in PM medium supplemented with 2% (w/v) sucrose + 2.0 g l^− 1 peptone. For germination of embryos P₇₂₃ medium supplemented with 1.0 mg l^− 1 BAP proved best. Multiple shoot buds or protocorm-like bodies (PLBs) were produced from stem segments of in vitro raised seedlings. Both direct organogenesis and embryogenesis were observed and the morphogenetic response was initiated by different concentrations and combinations of PGRs. The optimum PGR combination for maximal PLB regeneration was 1.0 mg l^− 1 NAA + 2.5 mg l^− 1 BAP, while 1.0 mg l^− 1 NAA + 1.0 mg l^− 1 BAP for shoot bud development. Strong and stout root system was induced in half strength PM medium supplemented with 0.5 mg l^− 1 IAA. The well-rooted plantlets were transferred to pots containing a potting mixture composed of saw dust, coconut coir, humus, and coal pieces at 1:1:1:2 (w/w) with 80% survival in outside environment and flowered after two years of transfer.     

In vivo antimicrobial evaluation of an alanine-rich peptide derived from Pleuronectes americanus

In several organisms, the first barrier against microbial infections consists of antimicrobial peptides (AMPs) which are molecules that act as components of the innate immune system. Recent studies have demonstrated that AMPs can perform various functions in different tissues or physiological conditions. In this view, this study was carried out in order to evaluate the multifunctional activity in vivo of an alanine-rich peptide, known as Pa-MAP, derived from the polar fish Pleuronectes americanus. Pa-MAP was evaluated in intraperitoneally infected mice with a sub-lethal concentration of Escherichia coli at standard concentrations of 1 and 5 mg kg⁻¹. At both concentrations, Pa-MAPs exhibited an ability to prevent E. coli infection and increase mice survival, similar to the result observed in mice treated with ampicillin at 2 mg kg⁻¹. In addition, mice were monitored for weight loss. The results showed that mice treated with Pa-MAPs at 1 mg kg⁻¹ gained 0.8% of body weight during the 72 h of experiment. The same was observed with Pa-MAP at 5 mg kg⁻¹, which had a gain of 0.5% in body weight during the treatment. Mice treated with ampicillin at 2 mg kg⁻¹ show a significant weight loss of 5.6% of body weight. The untreated group exhibited a 5.5% loss of body weight. The immunomodulatory effects were also evaluated by the quantification of IL-10, IL-12, TNF-α, IFN-γ and nitric oxide cytokines in serum, but no immunomodulatory activity was observed. Data presented here suggest that Pa-MAP should be used as a novel antibiotic against infection control.     

Evaluation of the association of mercury(II) with some dicysteinyl tripeptides

The present study was undertaken to gain insight into the associations of mercury(II) with dicysteinyl tripeptides in buffered media at pH 7.4. We investigated the effects of increasing the distance between cysteinyl residues on mercury(II) associations and complex formations. The peptide–mercury(II) formation constants and their associated thermodynamic parameters in 3-(N-morpholino)propanesulfonic acid (MOPS) buffered solutions were evaluated by isothermal titration calorimetry. Complexes formed in different relative ratios of mercury(II) to cysteinyl peptides in ammonium formate buffered solutions were characterized by LTQ Orbitrap mass spectrometry. The results from these studies show that n-alkyl dicysteinyl peptides (CP 1–4), and an aryl dicysteinyl peptide (CP 5) can serve as effective “double anchors” to accommodate the coordination sites of mercury(II) to form predominantly one-to-one Hg(peptide) complexes. The aryl dicysteinyl peptide (CP 5) also forms the two-to-two Hg₂(peptide)₂ complex. In the presence of excess peptide, Hg(peptide)₂ complexes are also detected. Notably, increasing the distance between the ligating groups or “anchor points” in CP 1–5 does not significantly affect their affinity for mercury(II). However, the enthalpy change (ΔH) values (ΔH₁ ∼ −91 kJ mol⁻¹ and ΔH₂ ∼ −66 kJ mol⁻¹) for complex formation between CP 4 and 5 with mercury(II) are about one and a half times larger than the related values for CP 1, 2 and 3 (ΔH₁ ∼ −66 kJ mol⁻¹ and ΔH₂ ∼ 46 kJ mol⁻¹). The corresponding entropy change (ΔS) values (ΔS₁ ∼ −129 J K⁻¹ mol⁻¹ and ΔS₂ ∼ −116 J K⁻¹ mol⁻¹) of the structurally larger dicysteinyl peptides CP 4 and 5 are less entropically favorable than for CP 1, 2 and 3 (ΔS₁ ∼ −48 J K⁻¹ mol⁻¹ and ΔS₂ ∼ −44 J K⁻¹ mol⁻¹). Generally, these associations result in a decrease in entropy, indicating that these peptide–mercury complexes potentially form highly ordered structures. The results from this study show that dicysteinyl tripeptides are effective in binding mercury(II) and they are promising motifs for the design of multi-cysteinyl peptides for binding more than one mercury(II) ion per peptide.