Optimization of cultural conditions of Arthrobacter sp. Sphe3 for growth-associated chromate(VI) reduction in free and immobilized cell systems.

The current study aimed to characterize Arthrobacter sp. Sphe3 ability to reduce Cr(VI) in suspended cell cultures as well as in immobilized form using Ca-alginate beads. Adaptation studies in the presence of 5 mg L(-1) Cr(VI) showed a significant increase in specific growth rate from 0.25 to 0.3 h(-1) and bioremoval percentage from 64% to 94% (p<0.05), whereas Arthrobacter sp. Sphe3 could tolerate up to 50 mg L(-1) Cr(VI). Optimization of culture conditions resulted in complete reduction of 45 mg L(-1) Cr(VI) at 30 °C, pH 8 and 10 g L(-1) of glucose. High glucose concentrations helped at reducing (80±2.4)% of initial 100 mg L(-1) Cr(VI), whereas the bacterial strain could tolerate 850 mg L(-1) Cr(VI). Cr(III) formation was first evidenced by the appearance of a green insoluble precipitate in the medium. Cell biomass was successfully immobilized in Ca-alginate beads that were evaluated for their stability. Cell release was sharply decreased when 4% Na-alginate was used under non-shaking conditions. Biotransformation efficiency was enhanced when 25-50 mg cells mL(-1) Na-alginate from the exponential growth phase were collected and co-encapsulated with either 1% glucose and 0.5% (NH4)2SO4, or 1% LB medium. Immobilized biocatalyst could be reused up to 6 continuous cycles in the presence of 10 mg L(-1) Cr(VI), but its performance was lowered at higher metal concentrations comparing with free cells that significantly maintained their reducing ability up to 300 mg L(-1) Cr(VI).

Equilibrium, thermodynamic and kinetic studies on biosorption of Mn(II) from aqueous solution by Pseudomonas sp., Staphylococcus xylosus and Blakeslea trispora cells.

Biosorption of Mn(II) from aqueous solutions using Pseudomonas sp., Staphylococcus xylosus and Blakeslea trispora cells was investigated under various experimental conditions of pH, biomass concentration, contact time and temperature. The optimum pH value was determined to 6.0 and the optimum biomass concentration to 1.0 g L(-1) for all types of cells. Mn(II) biosorption was found to fit better to the Langmuir model for Pseudomonas sp. and B. trispora and to Freundlich model for S. xylosus. Langmuir model gave maximum Mn(II) uptake capacity 109 mg g(-1) for Pseudomonas sp. and much lower, 59 mg g(-1) and 40 mg g(-1) for S. xylosus and B. trispora, respectively. Pseudo-second-order kinetic model was also found to be in good agreement with the experimental results. Thermodynamic parameters of the adsorption confirmed the endothermic nature of sorption process with positive heat of enthalpy, accompanied by a positive value of entropy change. Interestingly, desorption experiments by treating biomass with 0.1 M HNO(3) solution resulted to more than 88% recovery of the adsorbed Mn(II) from Pseudomonas sp. and almost 95% and 99% from S. xylosus and B. trispora cells respectively, thus indicating that Mn(II) can be easily and quantitatively recovered from biomass.

Co-metabolism of 2,4-dichlorophenol and 4-Cl-m-cresol in the presence of glucose as an easily assimilated carbon source by Staphylococcus xylosus.

Comparison of the ability of Staphylococcus xylosus to degrade 2,4-dichlorophenol and 4-Cl-m-cresol in separate cultures is reported. Bacterial adaptation and the continuous presence of glucose, as a conventional carbon source, were found to stimulate the degrading efficiency of S. xylosus. 4-Cl-m-cresol exhibited higher substrate-induced toxicity with K(ig) value at 0.25 mM, comparing to 2,4-dichlorophenol (K(ig) value at 0.90 mM) at initial concentration ranging from 0.1 to 0.5 mM. Degradation rate of 4-Cl-m-cresol was found to decrease only, revealing lower value of inhibition degradation constant (K(i) at 0.019 mM) comparing to that of 2,4-dichlorophenol (K(i) at 0.41 mM). Both glucose and each one of the chloro-aromatic compounds tested were simultaneously consumed and an increase of chloride ions in the medium appeared, during the exponential phase of growth. The chloride ions increase was nearly stoichiometric in the presence of 2,4-dichlorophenol and one of its several intermediate products identified was 2-Cl-maleylacetic acid. In the case of 4-Cl-m-cresol, only one metabolic product was found and identified as 3-methyl-4-oxo-pentanoic acid.

Effect of biomass pre-treatment and solvent extraction on beta-carotene and lycopene recovery from Blakeslea trispora cells.

The production of carotenoids from Blakeslea trispora cells in a synthetic medium has been reported, with the main products being beta-carotene, lycopene, and gamma-carotene. The effect of biomass pretreatment and solvent extraction on their selective recovery is reported here. Eight solvents of class II and III of the International Conference of Harmonization: ethanol, methanol, acetone, 2-propanol, pentane, hexane, ethyl acetate, and ethyl ether, and HPLC analysis were used for the evaluation of their selectivities towards the three main carotenoids with regard to different biomass pre-treatment. The average C(max) values (maximum concentration of caronoids in a specific solvent) were estimated to 16 mg/L with the five out of eight solvents investigated, whereas methanol, pentane, and hexane gave lower values of 10, 11, and 9 mg/L, respectively. The highest carotenoid yield was obtained in the case of wet biomass, where 44-56% is recovered with one solvent and three extractions and the rest is recovered only after subsequent treatment with acetone; thus, four extractions of 2.5 h are needed. Two extractions of 54 min are enough to recover carotenoids from dehydrated biomass, with the disadvantage of a high degree of degradation. Our results showed that, for maximum carotenoid recovery, ethyl ether, 2-propanol, and ethanol could be successfully used with biomass without prior treatment, whereas fractions enriched in beta-carotene or lycopene can be obtained by extraction with the proper solvent, thus avoiding degradation due to time-consuming processes.

Kinetics of 2,4-dichlorophenol and 4-Cl-m-cresol degradation by Pseudomonas sp. cultures in the presence of glucose.

Comparison of the ability of Pseudomonas sp. to degrade 2,4-dichlorophenol and 4-Cl-m-cresol in separate cultures in the presence of glucose, as a conventional carbon source, is reported. The specific growth rates at 0.1 mM 2,4-dichlorophenol and 4-Cl-m-cresol were estimated to be 0.181 and 0.154 h(-1), respectively, showing that Pseudomonas sp. is mainly inhibited by 4-Cl-m-cresol. The percentage of consumption ranges between 65% and 11% for 2,4-dichlorophenol and between 37% and 8% for 4-Cl-m-cresol, respectively, depending on its initial concentration. The dechlorination of the two compounds was investigated in the growth media and it was found that chloride liberation in the case of 2,4-dichlorophenol took place during the exponential phase of growth, followed by pH decrease from 6.1 to 5.8 at 0.1 mM. In contrast, in the case of 4-Cl-m-cresol chloride ion release was observed to a lesser extent, indicating the different metabolic pathway of 4-Cl-m-cresol. 2,4-Dichlorophenol and 4-Cl-m-cresol degradation followed a first-order kinetics model, whereas glucose consumption fitted well a zero-order kinetics model.

Comparative study of Cd(II) and Cr(VI) biosorption on Staphylococcus xylosus and Pseudomonas sp. in single and binary mixtures.

Biosorption of Cd(II) and Cr(VI) ions in single solutions using Staphylococcus xylosus and Pseudomonas sp., and their selectivity in binary mixtures was investigated. Langmuir and Freundlich models were applied to describe metal biosorption and the influence of pH, biomass concentration and contact time was determined. Maximum uptake capacity of cadmium was estimated to 250 and 278 mg g(-1), whereas that of chromium to 143 and 95 mg g(-1) for S. xylosus and Pseudomonas sp., respectively. In binary mixtures with Cd(II) ions as the dominant species, there is a profound selectivity for cadmium biosorption, reaching 96% and 89% for Pseudomonas sp. and S. xylosus, respectively, at 10 mg l(-1) Cd(II) and 5 mg l(-1) Cr(VI). Interesting, when chromium (VI) ions are the dominant species, there is selectivity towards chromium around 92% with S. xylosus only.

Conjugation of resveratrol with RGD and KGD derivatives.

The reaction between Arg-Gly-Asp (RGD) and Lys-Gly-Asp (KGD) derivatives with 3,4',5-trihydroxy-trans-stilbene (resveratrol) was investigated. Knowing that resveratrol, RGD as well as KGD analogues inhibit human platelet aggregation in vitro, it was tempting for us to examine whether their coupling products present enhanced biological activity. Here, we report on the synthesis and identification of these coupling products. The N-protected peptides were synthesized by solid phase technique, using the 2-chlorotrityl-chloride resin, by the method of carbodiimides. Coupling reactions with resveratrol took place in solution using N,N-dicyclohexylcarbodiimide (DCC) as coupling reagent and 4-dimethylaminopyridine (DMAP) as catalyst. The reaction products were purified by reversed phase HPLC and identified by ESI-MS. The mono-esterified resveratrol derivative was the main (or only) reaction product, whereas the di- and the tri-ester (to a less extent) formation was noticed in some cases.

Synthesis and cytogenetic effects of aminoquinone derivatives with a di- and a tripeptide.

Quinones are of significant interest due to their important role in specific cellular functions. Quinoproteins are a big class of oxyreductive agents occurring in bacteria and other organisms. In this investigation derivatives of 2-amino-1,4-benzoquinone, 2-amino-1,4-naphthoquinone and 2-amino-5,8-dihydroxy-1,4-naphthoquinone with a di- and a tripeptide were prepared for first time. The effect of the synthesized compounds on sister chomatid exchange (SCE) rates and human lymphocyte proliferation kinetics on a molar basis was studied. Among these coupled products the most effective in inducing SCEs and depressing proliferation rate indices is the coupling product of 2-amino-1,4-naphthoquinone with the tripeptide GHK (10). Next in order of magnitude in inducing cytogenetic effects is 2-amino-1,4-naphthoquinone (2) and its coupling products with glycine and serine (4 and 5), while the rest displayed marginal activity.

Spectral analysis of a series of partially protected and deprotected tetrapeptides, analogues of AS-I phytotoxin.

A series of six tetrapeptides, analogues of AS-I phytotoxin, pathogenic to sunflower, have been synthesized either in solution and/or by solid phase methods and have been tested for phytotoxic activity in various plants and cytotoxic activity in three cancer cell lines. These peptides were identified as model compounds by fast atom bombardment (FAB), plasma desorption (PD), electrospray ionization (ESI) mass spectrometry and by 1H, 1H-1H, 13C and 1H-13C NMR. The data presented show that in protected tetrapeptides the molecular ion was easily identified whereas some difficulties appeared with the fully deprotected peptides. NMR spectra are given.

Cu(III)-polypeptide complexes exhibiting SOD-like activity.

The SOD-like activity of Cu(III)-complexes with polypeptides poly-L-lysine and poly-L-glutamic acid respectively was investigated. The Cu(II)-polypeptide complexes were first oxidized by K2IrCl6 to give the corresponding Cu(III)-compounds. The oxidation of Cu(II) and the corresponding Cu(II)/Cu(III) potential was evaluated by cyclic voltammetry (c.v.), UV-Vis and EPR spectroscopic (r.t.) experiments. Spin trapping EPR spectra were also conducted to confirm the formation of the superoxide radical. The SOD-like activity of each Cu(III)-complex was proved using the nitro blue tetrazolium (NBT) method slightly modified.

Xanthan gum production by Xanthomonas campestris w.t. fermentation from chestnut extract.

Xanthomonas campestris w.t. was used for production of xanthan gum in fermentations with chestnut flour for the first time. Fermentations were carried out with either chestnut flour or its soluble sugars (33.5%) and starch (53.6%), respectively, at 28 degrees C and 200 rpm at initial pH 7.0 in flasks. The effect of agitation rate (at 200, 400, and 600 rpm) on xanthan gum production was also studied in a 2-L batch reactor. It was found that xanthan production reaches a maximum value of 3.3 g/100 mL at 600 rpm and 28 degrees C at 45 h.

Antiproliferative activity of synthetic tetrapeptides, analogs of AS-I phytotoxin, towards cancer cell lines.

The in vitro chemosensitivity of three cancer cell lines [HT29 (colon), HeLa (cervical) and T47D (breast)] to eight synthetic tetrapeptides, analogs of AS-I toxin, with phytotoxic effect on a series of plants was studied. Mouse fibroblast L929 cell line was also tested for chemosensitivity to these peptides. All cell lines were especially sensitive to Cys-Val-Gly-Glu tetrapeptide with IC50 values of 0.18, 0.3 and 0.63 mM for HT29, HeLa and T47D cells, respectively, whereas the IC50 value for the L929 cells was higher than 1 mM. Antiproliferative activity was also observed with peptides Tyr-Val-Gly-Glu and His-Val-Gly-Glu with IC50 values higher than those obtained for Cys-Val-Gly-Glu. For the rest of the peptides tested the IC50 values were found close to or higher than 3 mM.

Hyperalkaline and thermostable phosphatase in Thermus thermophilus.

The phosphatases existing in the extreme thermophilic bacterium Thermus thermophilus have been studied. Utilizing ion exchange, hydrophobic, pseudoaffinity, and affinity chromatography, a number of distinct phosphatase activities were identified. At least four phosphatases, with optimum pH ranging between 5.0 and 11.5, were assayed with p-nitrophenylphosphate, and two with optimum pH between 7.0 and 11.0, with 32P-casein as substrate. The authors have focused on the hyperalkaline phosphatase and have tried to purify and characterize it. This hyperalkaline phosphatase reaches a maximal level at the stationary phase of the growth, and is co-purified with alkaline phosphatase with optimum pH of 10.2. The enzymes present a relative mol wt of 65 and 58 kDa, respectively, as judged by SDS-PAGE and Sephadex G-150 column, and possess similar properties, indicating that they are isoforms. These enzymes barely function in the presence of tartrate, and are inhibited by EDTA, pyrophosphate, and molybdate. Among the metals tested, Hg2+ appeared as the strongest inhibitor of the hyperalkaline phosphatase. The two enzymes are thermostable and, upon treatment at 90 degrees C for 10 min, 75% of their activity remains. The physiological role and function of these phosphatases need further investigation.

A low molecular weight peptide from Allium porum with inhibitory activity on platelet aggregation in vitro.

A peptide factor was isolated from Allium porum with strong inhibitory activity on platelet aggregation induced by collagen or ADP. The peptide was isolated by a series of chromatographic techniques and purified to homogeneity by HPLC on RP C8 column. Its molecular weight, as it was estimated by plasma desorption mass spectrometry, is 1052 Da. Amino acid analysis showed the presence of Phe, Asp, Ser, Thr, Arg and Pro.

Effect of X-Asp (or Asn)-Phe-Nh2 (where X; H, Met) peptides on human platelet aggregation in vitro.

A series of di- and tripeptides containing aspartic or asparagine as N-terminal or intermediate amino acid were synthesized and tested for their effect on human platelet aggregation in vitro. It was found that only Met-Asp(or Asn)-Phe-NH2 inhibited platelet aggregation induced by collagen, ADP or adrenaline. Asn-Phe-NH2 and to a small extent Asp-Phe-NH2 presented strong aggregatory activity at low concentrations (at 0.5 mM or lower than this). All the other peptides tested, did not show any effect on platelet aggregation even at the concentration of 10 mM.

Synthesis of S-farnesyl-L-cysteine methylester and purification by HPLC.

S-trans, trans-farnesyl-L-cysteine methylester, a post translational modified amino acid, was synthesized from farnesyl bromide and L-cysteine methylester hydrochloride salt in the presence of triethylamine. Its purification as well as separation from the other isomers by HPLC on RP Vydac C4 and C8 columns are reported here.

Effect of leu-(Asp, Asn, Glu, Gln) dipeptides on platelet aggregation in vitro.

A series of Leu-(Asp, Asn, Glu, Gln) dipeptides were synthesized and tested for their effect on human platelet aggregation in vitro induced by collagen, ADP, or adrenaline. It was found that only Leu-Asp-NH2 and Leu-Asn-NH2 inhibit rather strongly platelet aggregation, whereas a small inhibition was observed with Leu-Glu-NH2 and Leu-Gln-NH2, respectively.

Characterization of the platelet aggregation inducer and inhibitor isolated from Crocus sativus.

Bulbs of Crocus sativus variety Cartwrightianus were found to contain both a platelet aggregation inducer and inhibitor. The aggregating factor has a Mr of 42 kDa estimated by a Sephadex G75 column and SDS-polyacrylamide slab gel electrophoresis. It was found to lack enzymatic activity such as proteinase, esterase and acid or alkaline phosphatase. The inhibitory factor was also purified to homogeneity by different chromatographic techniques and shows a Mr of 27 kDa as it was estimated by Biogel P30 column and SDS-polyacrylamide slab gel electrophoresis. It was found to possess strong proteinase activity.