Structural modeling of glucanase–substrate complexes suggests a conserved tyrosine is involved in carbohydrate recognition in plant 1,3-1,4-β-<Emphasis Type="SmallCaps">d</Emphasis>-glucanases

Glycosyl hydrolase family 16 (GHF16) truncated Fibrobacter succinogenes (TFs) and GHF17 barley 1,3-1,4-β-d-glucanases (β-glucanases) possess different structural folds, β-jellyroll and (β/α)₈, although they both catalyze the specific hydrolysis of β-1,4 glycosidic bonds adjacent to β-1,3 linkages in mixed β-1,3 and β-1,4 β-d-glucans or lichenan. Differences in the active site region residues of TFs β-glucanase and barley β-glucanase create binding site topographies that require different substrate conformations. In contrast to barley β-glucanase, TFs β-glucanase possesses a unique and compact active site. The structural analysis results suggest that the tyrosine residue, which is conserved in all known 1,3-1,4-β-d-glucanases, is involved in the recognition of mixed β-1,3 and β-1,4 linked polysaccharide.     

Production of Cyclodextrins by CGTase from Bacillus clausii Using Different Starches as Substrates

Cyclodextrins (CDs) are cyclic oligasaccharides composed by d-glucose monomers joined by α-1,4-d glicosidic linkages. The main types of CDs are α-, β- and γ-CDs consisting of cycles of six, seven, and eight glucose monomers, respectively. Their ability to form inclusion complexes is the most important characteristic, allowing their wide industrial application. The physical property of the CD-complexed compound can be altered to improve stability, volatility, solubility, or bio-availability. The cyclomaltodextrin glucanotransferase (CGTase, EC 2.4.1.19) is an enzyme capable of converting starch into CD molecules. In this work, the CGTase produced by Bacillus clausii strain E16 was used to produce CD from maltodextrin and different starches (commercial soluble starch, corn, cassava, sweet potato, and waxy corn starches) as substrates. It was observed that the substrate sources influence the kind of CD obtained and that this CGTase displays a β-CGTase action, presenting a better conversion of soluble starch at 1.0%, of which 80% was converted in CDs. The ratio of total CD produced was 0:0.89:0.11 for α/β/γ. It was also observed that root and tuber starches were more accessible to CGTase action than seed starch under the studied conditions.     

Polysaccharide hydrolases from leaves of Boscia senegalensis: properties of endo-(1-->3)-beta-D-glucanase

The leaves of Boscia senegalensis are traditionally used in West Africa in cereal protection against pathogens, pharmacologic applications, and food processing. Activities of α-amylase, β-amylase, exo-(1→3, 1→4)-β-d-glucanase, and endo-(1→3)-β-d-glucanase were detected in these leaves. The endo-(1→3)-β-d-glucanase (EC3.2.1.39) was purified 203-fold with 57% yield. The purified enzyme is a nonglycosylated monomeric protein with a molecular mass of 36 kDa and pI≥10.3. Its optimal activity occurred at pH 4.5 and 50°C. Kinetic analysis gave V _max, k _cat , and K _m values of 659 U/mg, 395 s⁻¹, and 0.42 mg/mL, respectively, for laminarin as substrate. The use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry and high-performance liquid chromatography revealed that the enzyme hydrolyzes not only soluble but also insoluble (1→3)-β-glucan chains in an endo fashion. This property is unusual for endo-acting (1→3)-β-d-glucanase from plants. The involvement of the enzyme in plant defense against pathogenic microorganisms such as fungi is discussed.     

Β-d-Xylosidase from Selenomonas ruminantium: Catalyzed Reactions with Natural and Artificial Substrates

Catalytically efficient β-d-xylosidase from Selenomonas ruminantium (SXA) exhibits pK _as 5 and 7 (assigned to catalytic base, D14, and catalytic acid, E186) for k _cat/K _m with substrates 1,4-β-d-xylobiose (X2) and 1,4-β-d-xylotriose (X3). Catalytically inactive, dianionic SXA (D14⁻E186⁻) has threefold lower affinity than catalytically active, monoanionic SXA (D14⁻E186^H) for X2 and X3, whereas D14⁻E186⁻ has twofold higher affinity than D14⁻E186^H for 4-nitrophenyl-β-d-xylopyranoside (4NPX), and D14⁻E186⁻ has no affinity for 4-nitrophenyl-α-l-arabinofuranoside. Anomeric isomers, α-d-xylose and β-d-xylose, have similar affinity for SXA. 4-Nitrophenol competitively inhibits SXA-catalyzed hydrolysis of 4NPX. SXA steady-state kinetic parameters account for complete progress curves of SXA-catalyzed hydrolysis reactions. The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned.     

New polyhydroxysteroids and steroid glycosides from the far east starfishCeramaster patagonicus

Two new polyhydroxysteroids and five new glycosides were isolated from the starfishCeramaster patagonicus and their structures were elucidated: 5α-cholestane-3β,6α,15β,16β,26-pentol, (22E)-5α-cholest-22-ene-3β,6α,8,15α,24-pentol, (22E)-28-O-[O-(2-O-methyl-β-d-xylopyranosyl)-(1→2)-β-d-galactofuranosyl]-24-hydroxymethyl-5α-cholest-22-ene-3β,4β, 6α,8,15β,16β,28-heptol (ceramasteroside C₁), (22E)-28-O-[O-(2,4-di-O-methyl-β-d-xylopyranosyl)-(1→2)-β-d-galactofuranosyl]-24-hydroxymethyl-5α-cholest-22-ene-3β, 6α,8,15β,16β,28-hexol (ceramasteroside C₂), (22E)-28-O-[O-methyl-β-d-xylopyranosyl)-(1→2)-β-d-galactofuranosyl]-24-hydroxymethyl-5α-cholest-22-ene-3β,6α,8,15β,16β 28-hexol (eramasteroside C₃), (22E)-28-O-[O-(2-O-methyl-β-d-xylopyranosyl)-(1→2)-β-d-galactofuranosyl]-24-methyl-5α-cholest-22-ene-3β,4β,6α,8, 15β, 26-hexol (ceramasteroside C₄), and (22E)-28-O-[O-(2-O-methyl-β-d-xylopyranosyl)-(1→2)-β-d-xylopyranosyl]-5α-cholest-22-ene-3β,6α,8,15β,24-pentol (ceramasteroside C₅)). Three known polyhydroxysteroids (24-methylene-5α-cholestane-3β,6α,8,15β,16β,26-hexol, 5α-cholestane-3β,6α,8,15β,16β,26-hexol, and 5α-cholestane-3β,6β,15α,16β,26-pentol) were also isolated. Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 1, pp. 190–195, January, 1997.     

Characterization of a Thermostable Family 1 Glycosyl Hydrolase Enzyme from <Emphasis Type="Italic">Putranjiva roxburghii</Emphasis> Seeds

A 66-kDa thermostable family 1 Glycosyl Hydrolase (GH1) enzyme with β-glucosidase and β-galactosidase activities was purified to homogeneity from the seeds of Putranjiva roxburghii belonging to Euphorbiaceae family. N-terminal and partial internal amino acid sequences showed significant resemblance to plant GH1 enzymes. Kinetic studies showed that enzyme hydrolyzed p-nitrophenyl β-d-glucopyranoside (pNP-Glc) with higher efficiency (K _cat/K _m = 2.27 × 10⁴ M⁻¹ s⁻¹) as compared to p-nitrophenyl β-d-galactopyranoside (pNP-Gal; K _cat/K _m = 1.15 × 10⁴ M⁻¹ s⁻¹). The optimum pH for β-galactosidase activity was 4.8 and 4.4 in citrate phosphate and acetate buffers respectively, while for β-glucosidase it was 4.6 in both buffers. The activation energy was found to be 10.6 kcal/mol in the temperature range 30–65 °C. The enzyme showed maximum activity at 65 °C with half life of ~40 min and first-order rate constant of 0.0172 min⁻¹. Far-UV CD spectra of enzyme exhibited α, β pattern at room temperature at pH 8.0. This thermostable enzyme with dual specificity and higher catalytic efficiency can be utilized for different commercial applications.     

Evaluation of host-guest complex formation between a benzimidazolic derivative and cyclodextrins by UV-VIS spectrophotometry and differential scanning calorimetry

Interactions between a benzimidazolic derivative, omeprazole (OME), beta-cyclodextrin (βCD) and a chemically modified βCD, methyl-beta-cyclodextrin (MβCD) were investigated in aqueous solution by UV-VIS spectroscopy and in solid state by differential scanning calorimetry (DSC). Phase solubility studies were used to evaluate the complexation in aqueous solution. The two solubility diagrams obtained were A_L type, indicating the formation of a drug-cyclodextrin complex with 1:1 stoichiometry. The complex of OME with MβCD showed a higher stability constant (K _S) than those with βCD. Some evidences of inclusion complexation in solid state were obtained from DSC. Only in thermal curves of OME-βCD lyophilized product and in OME-MβCD spray-dried and lyophilized systems the melting point of the drug disappeared completely suggesting the possible formation of an inclusion complex.     

Effects of Gene Orientation and Use of Multiple Promoters on the Expression of XYL1 and XYL2 in Saccharomyces cerevisiae

The gene encoding a glycoside hydrolase family 39 xylosidase (BH1068) from the alkaliphile Bacillus halodurans strain C-125 was cloned with a C-terminal His-tag, and the recombinant gene product termed BH1068(His)₆ was expressed in Escherichia coli. Of the artificial substrates tested, BH1068(His)₆ hydrolyzed nitrophenyl derivatives of β-d-xylopyranose, α-l-arabinofuranose, and α-l-arabinopyranose. Deviation from Michaelis−Menten kinetics at higher substrate concentrations indicative of transglycosylation was observed, and k _cat and K _m values were measured at both low and high substrate concentrations to illuminate the relative propensities to proceed along this alternate reaction pathway. The pH maximum was 6.5, and under the conditions tested, maximal activity was at 47°C, and thermal instability occurred above 45°C. BH1068(His)₆ was inactive on arabinan, hydrolyzed xylooligosaccharides, and released only xylose from oat, wheat, rye, beech, and birch arabinoxylan, and thus, can be classified as a xylosidase with respect to natural substrate specificity. The enzyme was not inhibited by up to 200 mM xylose. The oligomerization state was tetrameric under the size-exclusion chromatography conditions employed.     

Xylanolytic enzyme production by anAspergillus niger isolate

Production of xylanolytic enzymes by anAspergillus niger CCMI 850 isolate was investigated in batch cultures. The effect of the composition of a fermentation medium that did not include chemical inducers, on β-xylanase, β-xylosidase, α-l-arabinofuranosidase, and total cellulase activity was studied. With 4% xylan as the carbon source, about 65 U/mL of β-xylanase was obtained, whereas the total cellulase activity was undetectable, under the specified conditions. This β-xylanase activity represents the highest reported for a wild-type strain ofA. niger. The effect of pH and temperature on the activity of β-xylanase was studied. Partial characterization of the β-xylanase showed that with insoluble birchwood as substrate theK _m andV _max were 0.3 mM and 19 μmol/min, respectively. Aspects of using the crude β-xylanase preparation for applications in the pulp and paper industry were discussed.     

Synthesis of blockwise alkylated tetrasaccharide–organic quantum dot complexes and their utilization for live cell labeling with low cytotoxicity

Bioimaging is a key to understanding immune responses, cell differentiation, and development. Quantum dots (QDs) conjugated with monoclonal antibodies and other biomolecules are currently utilized for flow cytometry and immunohistochemistry, but monoclonal antibody–QD complexes are of limited use when cell surface markers are not available. In this study, we synthesized novel amphiphilic blockwise alkylated tetrasaccharides and developed a simple method for labeling a wide variety of live cells with organic QDs encapsulated with these carbohydrates. The novel amphiphilic blockwise alkylated tetrasaccharides were as follows: methyl β-d-glucopyranosyl-(1 → 4)-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-methyl-d-glucopyranoside (1), methyl β-d-galactopyranosyl-(1 → 4)-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-methyl-d-glucopyranoside (2), ethyl β-d-glucopyranosyl-(1 → 4)-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-ethyl-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-ethyl-d-glucopyranoside, (3), and ethyl β-d-galactopyranosyl-(1 → 4)-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-ethyl-β-d-glucopyranosyl-(1 → 4)-2,3,6-tri-O-ethyl-d-glucopyranoside (4). The newly synthesized blockwise alkylated tetrasaccharides spontaneously assembled into micelle-like particles, in which the hydrophobic moiety of the blockwise alkylated tetrasaccharides played an important role. They were less toxic to human cells than octyl β-d-glucopyranoside, a commonly used amphiphilic glucoside. Flow cytometry and confocal laser scanning microscopy revealed that the blockwise alkylated tetrasaccharide–organic QD complexes were stably attached to live cells. The affinity of compounds 1 and 2 to the live cell surface was slightly higher than that of compounds 3 and 4. Because the preparation of these carbohydrate–QD complexes is simple and does not require sophisticated equipment, and because the complexes can be autonomously attached to a wide spectrum of cell lines, they can be used as cell labeling reagents in biomedical studies.     

Polar steroidal compounds from the Far-Eastern starfish <Emphasis Type="Italic">Lethasterias fusca</Emphasis>

Nine steroidal compounds including three new steroidal glycosides, viz., sodium (24S)-3,24-di-O-(β-D-xylopyranosyl)-5α-cholestane-3β,6β,8,15α,24-pentol 15-sulfate (fuscaside A), (24S)-3,24-di-O-(β-D-xylopyranosyl)-5α-cholestane-3β,6β,8,15α,24-pentol (fuscaside B), and (22E,24R)-24-O-(β-D-xylopyranosyl)-5α-cholest-22-ene-3β,6α,8,15β,24-pentol (desulfated minutoside A); three previously known glycosides, viz., distolasterosides D₁ and D₂ and pycno-podioside A; two previously known polyhydroxysteroids, viz., 5α-cholestane-3β,6α,8,15β,16β,26-hexaol and 5α-cholestan-3β,4β,6α,7⇇8,15β,16β,26-octol; and the known sodium 24,25-dihydro-marthasterone 3-sulfate were isolated from the Far-Eastern starfish Lethasterias fusca. The structures of these compounds were elucidated by NMR spectroscopy and mass spectrometry. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 1, pp. 196–200, January, 2008.     

Hydrolysis and Transglycosylation Activity of a Thermostable Recombinant β-Glycosidase from <Emphasis Type="Italic">Sulfolobus acidocaldarius</Emphasis>

We expressed a putative β-galactosidase from Sulfolobus acidocaldarius in Escherichia coli and purified the recombinant enzyme using heat treatment and Hi-Trap ion-exchange chromatography. The resultant protein gave a single 57-kDa band by SDS-PAGE and had a specific activity of 58 U/mg. The native enzyme existed as a dimer with a molecular mass of 114 kDa by gel filtration. The maximum activity of this enzyme was observed at pH 5.5 and 90 oC. The half-lives of the enzyme at 70, 80, and 90 oC were 494, 60, and 0.2 h, respectively. The hydrolytic activity with p-nitrophenyl(pNP) substrates followed the order p-nitrophenyl-β-d-fucopyranoside > pNP-β-d-glucopyranoside > pNP-β-d-galactopyranoside > pNP-β-d-mannopyranoside > pNP-β-d-xylopyranoside, but not toward aryl-α-glycosides or pNP-β-l-arabinofuranoside. Thus, the enzyme was actually a β-glycosidase. The β-glycosidase exhibited transglycosylation activity with pNP-β-d-galactopyranoside, pNP-β-d-glucopyranoside, and pNP-β-d-fucopyranoside in decreasing order of activity, in the reverse order of its hydrolytic activity. The hydrolytic activity was higher toward cellobiose than toward lactose, but the transglycosylation activity was lower with cellobiose than with lactose.     

Structure-function relationships of a catalytically efficient β-<Emphasis Type="SmallCaps">D</Emphasis>-xylosidase

β-d-Xylosidase from Selenomonas ruminantium is revealed as the best catalyst known (k _cat, k _cat/K _m) for promoting hydrolysis of 1,4-β-d-xylooligosaccharides. ¹H nuclear magnetic resonance experiments indicate the family 43 glycoside hydrolase acts through an inversion mechanism on substrates 4-nitrophenyl-β-d-xylopyranoside (4NPX) and 1,4-β-d-xylobiose (X2). Progress curves of 4-nitrophenyl-β-d-xylobioside, xylotetraose and xylohexaose reactions indicate that one residue from the nonreducing end of substrate is cleaved per catalytic cycle without processivity. Values of k _cat and k _cat/K _m decrease for xylooligosaccharides longer than X2, illustrating the importance to catalysis of subsites −1 and +1 and the lack there of subsite +2. Homology models of the enzyme active site with docked substrates show that subsites bey ond−1 are blocked by protein and subsites bey ond +1 are not formed; they suggest that D14 and E186 serve catalysis as general base and general acid, respectively. Individual mutations, D14A and E186A, erode k _cat and k _cat/K _m by <10³ and to asimilar extent for substrates 4NPX and 4-nitrophenyl-α-l-arabinofuranoside (4NPA), indicating that the two substrates share the same active site. With 4NPX and 4NPA, pH governs k _cat/K _m with pK _a values of 5.0 and 7.0 assigned to D14 and E186, respectively. k _cat (4NPX) has a pK _a value of 7.0 and k _cat (4NPA) is pH independent above pH 4.0, suggesting that the catalytically inactive, “dianionic” enzyme form (D14-E187-) binds 4NPX but not 4NPA. The mention of firm names or trade products does not imply that they are end orsed or recommended by the US Department of Agriculture over other firms or similar products not mentioned.     

Inexpensive isolation of beta-D-glucopyranosidase from alpha-L-arabinofuranosidase,alpha-L-rhamnopyranosidase,and o-acetylesterase

β-d-Glucopyranosidase (βG, EC 3.2.1.21) has been isolated from some collateral activities, α-l-arabinofuranosidase (Ara, EC3.2.1.55), α-l-rhamnopyranosidase (Rha, EC 3.2.1.40), and o-acetylesterase (Est, EC 3.1.1.53), using a commercial enzyme preparation and a simple method economically sustainable for the food industry. The procedure comprises precipitation of extraneous substances by adding ethanol and CaCl₂, ultrafiltration, and adsorption, first on bentonite and then on chitosan. The results obtained were the complete isolation of βG from the above-mentioned activities, a drastic reduction in extraneous compounds, such as brown substances and polysaccharides, and a slight increase in purification.     

Immobilization of β-Galactosidase onto Magnetic Beads

A study of the cross-linking of β-galactosidase on magnetic beads is reported here. The magnetic beads were prepared from artemisia seed gum, chitosan, and magnetic fluid in the presence of a cross-linking regent (i.e., glutaraldehyde). The reactive aldehyde groups of the magnetic beads allowed the reaction of the amino groups of the enzymes. The animated magnetic beads were used for the covalent immobilization of β-galactosidase. The effect of various preparation conditions on the activity of the immobilized β-galactosidase, such as immobilizing time, amount of enzyme, and the concentration of glutaraldehyde, were investigated. The influence of pH and temperature on the activity and the stability of the enzyme, both free and immobilized, have been studied. And o-nitrophenyl-β-d-galactopyranoside (ONPG) was chosen as a substrate. The β-galactosidase immobilized on the magnetic beads resulted in an increase in enzyme stability. Optimum operational temperature for immobilized enzyme was 10 °C higher than that of free enzyme and was significantly broader.     

Inclusion Complexes of Cyclodextrins with Galangin: a Thermodynamic and Reactivity Study

The formation of the complexes of galangin (GAL) with native β-cyclodextrin (βCD), and with its substituted counterparts such as dimethyl-βCD (DMβCD) and hydroxypropyl-βCD (HPβCD), was studied by fluorescence spectra in aqueous medium. The binding association constants (K _a) of the complexes were determined at different temperatures. The formation constants obtained have the following trend upon complex formation at the three temperatures studied: HPβCD > DMβCD > βCD. The thermodynamic data for the inclusion of GAL in DMβCD and HPβCD indicated that is mainly enthalpy driven whereas for βCD it is an entropy-driven process.     

New class of carbohydrate-based nonionic surfactants: diblock co-oligomers of tri-<Emphasis Type="Italic">O</Emphasis>-methylated and unmodified cello-oligosaccharides

Mixtures of diblock co-oligomers of tri-O-methylated and unmodified cello-oligosaccharides have been found to be amphiphilic, as reported before. In order to clarify their accurate amphiphilic property, diblock co-oligomers of tri-O-methylated and unmodified cello-oligosaccharides with monodispersity, methyl β-d-glucopyranosyl-(1→4)-2,3,6–tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6–tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-d-glucopyranoside (1, pentamer), methyl β-d-glucopyranosyl-(1→4)- β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-d-glucopyranoside (2, hexamer), and methyl β-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-β-d-glucopyranosyl-(1→4)- 2,3,6-tri-O-methyl-d-glucopyranoside (3, trimer) were synthesized independently. These compounds had higher surface activities compared to the mixture of diblock co-oligomers of tri-O-methylated and unmodified cello-oligosaccharides and commercially available methylcellulose (MC) SM-4. This paper describes the methods of synthesis of these compounds, and the influence of amphiphilic character on their surface activity. A new class of carbohydrate-based nonionic surfactant without long alkyl chain was discovered.     

Combination of ionic liquid [bmim]PF<Subscript>6</Subscript> with boron trifluoride etherate as an efficient catalytic system for the glycosylation of α-tocopherol and naphthotocopherol

The use of a combination of ionic liquid [bmim]PF₆ with boron trifluoride etherate as the catalyst in the glycosylation of α-tocopherol and chromanol of vitamin K₁ (naphthotocopherol) allowed us to obtain β-glycosides in high yield when β-anomer of peracetylated D-glucose was used. In addition, usually inactive α-anomers of peracetylated D-glucopyranose and D-galactopyranose were involved in this reaction. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 12, pp. 2401–2404, December, 2007.     

Production and Physico-chemical Characterization of a Biosurfactant Produced by <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> OBP1 Isolated from Petroleum Sludge

Pseudomonas aeruginosa strain OBP1, isolated from petroleum sludge, was used to produce biosurfactant from a modified mineral salt medium with 2% n-hexadecane as sole source of carbon. The crude biosurfactant was fractionated using TLC and HPLC. Using FTIR spectroscopy, ¹H NMR, and LC-MS analyses, the chemical structure of the purified fraction of crude biosurfactant was identified as rhamnolipid species. The LC-MS spectra show that monorhamnolipid (l-rhamnopyranosyl-β-hydroxydecanoyl-β- hydroxydecanoate, Rha-C₁₀-C₁₀) was produced in abundance with the predominant congener [M–H]⁻ ions for l-rhamnopyranosyl-l-rhamnopyranosyl-3-hydroxydecanoyl-3-hydroxydecanoate (Rha-Rha-C₁₀-C₁₀). Seven different carbon substrates and five nitrogen sources were examined for their effect on rhamnolipid production. Using n-hexadecane (20 g/l) as carbon substrate and urea along with (NH₄)₂SO₄ (2 g/l each) as nitrogen source was found to be the best, with a maximum yield of 4.8 g/l. The biosurfactant reduced the surface tension of water to 31.1 mNm⁻¹ with a critical micelle concentration of 45 mg/l. The biosurfactant showed a better emulsifying activity against a variety of hydrocarbon and achieved a maximum emulsion index of 82% for diesel. The purified biosurfactant showed a significant antibacterial activity against Staphylococcus aureus at a minimum inhibitory concentration of 8 μg/ml.     

Electrochemical preparation and <Emphasis Type="Italic">in situ</Emphasis> characterization of poly(3-methylpyrrole) and poly(3-methylpyrrole-cyclodextrin) films on gold electrodes

A Electrosynthesis of conducting poly(3-methylpyrrole) (P3MPy) and poly(3-methylpyrrole-2,6-dimethyl-β-cyclodextrin) (poly(3MPy-β-DMCD)) films on a gold electrode in acetonitrile electrolyte solution containing lithium perchlorate has been carried out by potential cycling. Products were characterized with cyclic voltammetry CV, in situ UV-Vis spectroscopy, and in situ resistance measurements. Electrosynthesis of poly(3MPy-β-DMCD) started with a (1:1) (3MPy-β-DMCD) supramolecular cyclodextrin CD complex of 3-methylpyrrole characterized with proton NMR spectroscopy. The oxidation peak of poly(3MPy-β-DMCD) in CVs is shifted to more positive values than P3MPy. In situ resistance measurements show that the resistance of poly(3MPy-β-DMCD) is higher than of P3MPy by approximately an order of magnitude. Minimum resistance can be observed for P3MPy and poly(3MPy-β-DMCD) at 0.40 < E _Ag/AgCl < 1.10 V and 0.60 < E _Ag/AgCl < 1.10 V, respectively. The higher resistance of P3MPy compared with polypyrrole may result from the presence of the methyl group substituent resulting in a decreased conjugation length. When CD is present during synthesis, resistance is even higher. In situ UV-Vis spectroelectrochemical data for both films prepared potentiodynamically by cycling the potential in the range −0.20 < E _Ag/AgCl < 1.10 V in acetonitrile electrolyte show major effects of CD presence during electrosynthesis.