Electrodeposition of Fe-Sn from the chloride-based electrolyte

The conditions for electrodeposition of Fe-Sn alloys from a novel, environmentally friendly, ferrous chloride-based electrolyte were studied. The influence of the pH on the electrolyte stability and deposit stoichiometry was studied and discussed. Anodic stripping voltammetry (ASV), XRD and SEM/EDX were used to characterise the electrodeposited phases. The results from ASVs indicated the possibility to deposit at least two different phases at high overpotentials. Hull cell depositions in an electrolyte with a Sn to Fe ratio 1:1 and a pH value of 2.8 showed regular deposition of Fe-Sn. Fe-rich deposits (54.84 at.-%) were obtained from an electrolyte with Sn to Fe ratio 1:10. The XRD results were compatible with the electrochemical investigations. In all studied samples β-Sn, FeSn2 and Fe5Sn3 were detected. The presence of ferromagnetic Fe5Sn3 was not influenced by the Sn to Fe ions ratio in the electrolyte

Alkoxylated β-Naphthol as an Additive for Tin Plating from Chloride and Methane Sulfonic Acid Electrolytes

Beta-naphthol was one of the first additives introduced for smooth and homogeneoustin electrodeposition. Although it can be oxidized under the plating conditions, forming either1,2-napthoquinone or polymeric materials based on naphthioxides, it is still in use. In this work,an investigation of its more stable form, alkoxylated beta-naphthol (ABN), on tin plating is undertaken.For this purpose, chloride based (pH ~5) and methane sulfonic acid (MSA, pH ~0.5) electrolytes,including ABN, were prepared. Reaction kinetics were studied by polarization, Tafel measurements,and cyclic voltammetry. Tin electrodeposits were obtained on flat brass substrates. Surfacemorphology and preferred crystal orientation were studied by Scanning Electron Microscopy (SEM)and X-ray Diffraction (XRD). In both studied electrolytes ABN acts as an inhibitor but in the case ofthe chloride electrolyte it is more pronounced. In the MSA electrolyte this effect was overlaid by thepresence of tin-citrate complexes. In the chloride-based electrolyte, ABN has a grain refining effect,while in the MSA electrolyte an increase of ABN concentration leads to a slight enlargement of theaverage grain size. X-ray analysis shows a constant decrease of the (101) intensity with increasingconcentration of ABN for the sample deposited from both baths

Chemo-biocatalytic one-pot two-step conversion of cyclic amine to lactam using whole cell monoamine oxidase

BACKGROUND: Most biocatalysts currently involved in one‐pot chemoenzymatic cascades are pure enzymes, while whole cells and crude enzyme extracts remain unexplored. This work aims to develop a chemo‐biocatalytic one‐pot two‐step system involving whole cell monoamine oxidase (MAO, EC 1.4.3.4) coupled with a Cu‐based oxidative system (CuI/H2O2) for the transformation of 1,2,3,4‐tetrahydroisoquinoline (THIQ) to 3,4‐dihydroisoquinolin‐1(2H)‐one (DHIO). RESULTS: MAO‐N variants D9 and D11 were tested as whole cell and crude lysate biocatalysts for biological oxidation. Whole Escherichia coli OverExpress C43(DE3) cells expressing MAO‐N D9 showed the best performance (Vmax = 36.58 mmol L−1 h−1, KM = 8.124 mmol L−1, maximum specific productivity 89.3 μmol min−1 g−1DCW) and were employed in combination with CuI/H2O2 in a sequential one‐pot two‐step process. The biotransformation was scaled‐up to the initial volume of 25 mL and after triple THIQ feeding, 48.2 mmol L−1 of the intermediate 3,4‐dihydroisoquinoline (DHIQ) was obtained with a yield of 71.3%. Afterwards, chemical catalysts (1 mol% CuI and 10 eq. H2O2) were added to the biologically produced DHIQ, which was transformed to ∼30 mmol L−1 DHIO at 69.4% overall yield. CONCLUSION: As MAO‐N variants have wide substrate specificity, this work broadens the portfolio of one‐pot chemoenzymatic processes employing whole cell biocatalysts, representing an alternative to using pure enzymes

Ultra-high throughput functional enrichment of large monoamine oxidase (MAO-N) libraries by fluorescence activated cell sorting

Directed evolution enables the improvement and optimisation of enzymes for particular applications and is a valuable tool for biotechnology and synthetic biology. However, studies are often limited in their scope by the inability to screen very large numbers of variants to identify improved enzymes. One class of enzyme for which a universal, operationally simple ultra-high throughput (>106 variants per day) assay is not available is flavin adenine dinucleotide (FAD) dependent oxidases. The current high throughput assay involves a visual, colourimetric, colony-based screen, however this is not suitable for very large libraries and does not enable quantification of the relative fitness of variants. To address this, we describe an optimised method for the sensitive detection of oxidase activity within single Escherichia coli (E. coli) cells, using the monoamine oxidase from Aspergillus niger, MAO-N, as a model system. In contrast to other methods for the screening of oxidase activity in vivo, this method does not require cell surface expression, emulsion formation or the addition of an extracellular peroxidase. Furthermore, we show that fluorescence activated cell sorting (FACS) of large libraries derived from MAO-N under the assay conditions can enrich the library in functional variants at much higher rates than via the colony-based method. We demonstrate its use for directed evolution by identifying a new mutant of MAO-N with improved activity towards a novel secondary amine substrate. This work demonstrates, for the first time, an ultra-high throughput screening methodology widely applicable for the directed evolution of FAD dependent oxidases in E. coli

Biofilm characterization of slaughterhouse and environmental isolates in Campylobacter jejuni

Bakterijska adhezija in tvorba biofilma sta v živilski industriji lahko zelo problematična, saj so bakterijske celice znotraj biofilma bolj odporne na različne okoljske dejavnike. Tudi za Campylobacter jejuni je sposobnot tvorbe biofilma zelo pomembna za preživetje zunaj gostitelja. Namen eksperimentalnega dela magistrske naloge je bil karakterizirati biofilm 16 različnih sevov C. jejuni, izoliranih v klavniškem okolju ter iz površinskih vod, pri različnih pogojih (v MHB gojišču pri 42 ºC in 37 ºC, mikroaerofilno ter 8 ºC, aerobno in v MHB z dodatkom piščančjega izcedka pri 37 ºC, mikroaerofilno ter 8 ºC, aerobno). Kot kontaktno površino smo izbrali nerjavno jeklo, saj je to zelo pogosto uporabljen material v živilsko-predelovalnih obratih. Formirani biofilm na površini jeklenih diskov smo kvantificirali z gojitveno metodo in kolorimetrično metodo. Ob pogojih, 37 ºC, mikroaerofilno v MHB z dodanim piščančjim izcedkom in 42 ºC, mikroaerofilno v gojišču MHB so bili, povprečno, klavniški sevi bolj filmotvorni kot okoljski. Potrdili smo, da ima izcedek piščančjega mesa pomembno vlogo pri filmotvornosti bakterij C. jejuni. V nadaljevanju smo testirali tudi protimikrobno delovanje etanolnega izvlečka kraškega šetraja (Satureja montana) in benzalkonijevega klorida ter njuno sposobnost zmanjšanja števila že formiranih biofilmskih celic. Vsi sevi C. jejuni so bili dovzetni na obe testirani spojini, vedar je bil benzalkonijev klorid veliko bolj učinkovit. Obe testirani spojini sta pokazali tudi dober učinek zmanjšanja števila biofilmskih celic. Pri sevu C. jejuni B0975, enemu izmed najboljših filmotvorcev, smo določili tudi občutljivost planktonskih in biofilmskih celic na izbrane protimikrobne spojine, testirali membransko integriteto celic in določili količino akumuliranega etidijevega bromida v celicah. S tem smo želeli primerjati odpornost planktonskih in biofilmskih celic C. jejuni. Večje odpornosti biofilmskih celic z uporabljenimi metodami ne moremo potrditi.Bacterial adhesion and biofilm formation can be very problematic in the food industry, as bacterial cells within the biofilm are more resistant to various environmental factors. For Campylobacter jejuni, biofilm formation is also very important for their survival outside the host. The purpose of the experimental part of the master\u27s thesis was to characterize the biofilm of 16 different C. jejuni strains isolated in the slaughterhouse environment and from surface waters under different conditions (in MHB medium at 42 ºC in 37 ºC in microaerobic atmosphere and 8 ºC in aerobic atmosphere and in MHB with added chicken juice at 37 ºC in microaerobic atmosphere and 8 ºC in aerobic atmosphere). Stainless steel was chosen as the contact surface because it is a very commonly used material in food processing plants. The formed biofilm on the surface of the steel disks was quantified by the cultivation method and the colorimetric method. At 37 ºC in MHB supplemented with chicken juice in microaerobic atmosphere and 42 ºC in MHB in microaerobic atmosphere on average were slaughterhouse strains more biofilm-forming than environmental strains. We have confirmed that chicken juice plays an important role in the proces of biofilm formation of C. jejuni. In addition, we tested the antimicrobial activity of the ethanol extract of S. montana and benzalkonium chloride and their ability to reduce already formed biofilms. All C. jejuni strains were susceptible to both tested compounds, but benzalkonium chloride was much more effective. Both tested compounds also showed good biofilm reduction effect. In C. jejuni B0975 strain, one of the best biofilm formers, we determined the sensitivity of planktonic and biofilm cells to selected antimicrobial compounds, tested the membrane integrity of the cells and determined the amount of accumulated ethidium bromide in the cells. With this we wanted to compare the resistance of planktonic and biofilm cells of C. jejuni. Increased resistance of biofilm cells by the methods used cannot be confirmed

Comparison of Campylobacter jejuni slaughterhouse and surface-water isolates indicates better adaptation of slaughterhouse isolates to the chicken host environment

Campylobacter jejuni is an emerging food-borne pathogen that poses a high risk to human health. Knowledge of the strain source can contribute significantly to an understanding of this pathogen, and can lead to improved control measures in the food-processing industry. In this study, slaughterhouse and surface-water isolates of C. jejuni were characterized and compared in terms of their antimicrobial resistance profiles and adhesion to stainless steel and chicken skin. Resistance of C. jejuni biofilm cells to benzalkonium chloride and Satureja montana ethanolic extract was also tested. The data show that the slaughterhouse isolates are more resistant to ciprofloxacin, and adhere better to stainless steel at 42 °C, and at 37 °C in 50% chicken juice. Additionally, biofilm cells of the isolate with the greatest adhesion potential (C. jejuni S6) were harvested and tested for resistance to S. montana ethanolic extract, benzalkonium chloride, and erythromycinand for efflux-pump activity, as compared to their planktonic cells. The biofilm cells showed increased resistance to both S. montana ethanolic extract and erythromycin, and increased efflux-pump activity. These data indicate adaptation of C. jejuni slaughterhouse isolates to the chicken host, as well as increased biofilm cell resistance due to increased efflux-pump activity