1,721,031 research outputs found
Comparison of mesoporous silicate supports for the immobilisation and activity of cytochrome c and lipase
Full text linkThe activity and stability of Candida antartica lipase B (CALB) and cytochrome c immobilised on to, SBA-15 and a porous spherical silicate material (PPS), were examined. The materials possess similar pore diameters but have different morphologies, pore volumes and surface areas. CALB exhibited higher catalytic activity and stability on SBA-15 when compared to PPS, while cytochrome c showed similar catalytic activity on both materials. The activity of CALB immobilised on SBA-15 was retained (95%) after 7 uses, while CALB immobilised on PPS retained only 43% activity. Such changes can be mainly ascribed to the different physical properties (pore volume, surface area and pore shape) of the supports
Electrolyte effects on enzyme electrochemistry
No full textThe electrolytes used in enzymatic biosensors or biofuel cells have always been considered to be inert. However, recent studies have demonstrated that this assumption is not correct and that the nature of the electrolyte needs to be considered. Ion-specific interactions can occur with the faradaic response observed in both direct and mediated electron transfer being modulated by the nature of the salt used in solution. Specific ion effects arise from the Hofmeister series, which is well established in studies of protein systems but not in electrochemical studies of redox enzymes. Recent experimental and theoretical work on explaining the Hofmeister effect is described
Specific ion effects on the mediated oxidation of NADH
No full textThe electrochemical mediated oxidation of dihydronicotinamide adenine dinucleotide (NADH) by 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) was examined in a range of electrolytes. Changes in the Faradaic response were observed that were dependent on the nature of the salt used. In the presence of 200mM chloride salts, the Faradaic response followed the trend Na+>K+>Li+>Cs+>Gnd+while in the presence of 200mM sodium salts, the trend Cl->Br->F->ClO4->SCN-was observed. The observed trends varied when the concentration of the ions was altered. The effect of the cation also changed when the counterion was changed, indicating that specific ion pair effects were present. The results indicate that the nature of the electrolyte alters the Faradaic response
High Energy Ball Milling and Liquid Crystal Template Method: A Successful Combination for the Preparation of Magnetic Nano-Platforms
No full textIn this study, we present the preparation of superparamagnetic ordered mesoporous silica (SOMS) for biomedical applications by the combination of high energy ball milling (HEBM) and the liquid crystal template method (LCT) to produce a material comprised of room temperature superparamagnetic Fe3O4 nanoparticles in a MCM-41 like mesostructured silica. In a typical synthesis, a mixture of Fe2O3 and silica was sealed in a stainless-steel vial with steel balls. Ball milling experiments were performed in a vibratory mill apparatus. The milling process produced nanocomposites with an average size ranging from ∼100-200 nm, where the Fe3O4 nanoparticles (4.8 nm size) are homogeneously dispersed into the amorphous SiO2 matrix. The obtained nanocomposite has been used for the preparation of the SOMS through the LCT method. Structural, morphological and textural characterization were performed using X-ray powder diffraction, transmission electron microscopy and nitrogen sorption analysis. Field dependence of magnetization was investigated and showed superparamagnetic behaviour at 300 K with a value of saturation magnetization (Ms) that is of interest for biomedical applications
Specific Ion Effects on the Enzymatic Activity of Alcohol Dehydrogenase from Saccharomyces Cerevisiae
Full text linkThe enzymatic activity of alcohol dehydrogenase (ADH) in the presence of a range of electrolytes is investigated. In the presence of 150 and 200 mM cations a substantial increase in activity following the series GnCl < CsCl < KCl ∼ NaCl < LiCl was observed with a 69% increase in the presence of KCl 200 mM with respect to the salt-free solution. In the presence of 150 and 200 mM anions the increase in activity followed an ion specific trend NaF ∼ NaCl ∼ NaBr > no salt > NaClO4 > NaSCN with a peak in activity increase of 75% in the presence of NaBr. The values of the Michaelis–Menten constant (Km) did not show any significant ion specific effect, while the maximum rate (Vmax) of ethanol oxidation to acetaldehyde was strongly ion specific. The changes in specific activity and Vmax in the presence of anions likely arises from ion specific interactions with charged residues in the active site of ADH. The data indicate that the enzymatic activity of alcohol dehydrogenase can be modulated by the nature of electrolytes at physiological concentration
Lipases encapsulation onto ZIF-8: A comparison between biocatalysts obtained at low and high zinc/2 methylimidazole molar ratio in aqueous medium
No full textLipase AK from Pseudomonas fluorescens and Lipase RM from Rhizomucor miehei were encapsulated into a zeolite imidazolate framework (ZIF-8) by a “one-pot” synthesis to obtain AK@ZIF-8 and RM@ZIF-8 biocatalysts. The effect of a high (1:40) and low (1:4) Zn/2-methylimidazole molar ratio on the biocatalysts synthesis was investigated. The different Zn/ligand (L) ratios affected both the surface area, the loading, and the specific activity of the biocatalysts. Samples synthesized by using a high Zn/L ratio had high values of surface area whereas those obtained by using a low Zn/L ratio had higher loadings and specific activities. The decrease of pH (from 11.6 to 9.4) during the synthesis at high Zn/L ratio produced ZIF-8 samples with features similar to those observed for low Zn/L ratio samples. The low Zn/L (1:4) ratio AK@ZIF-8 biocatalyst retained 99 % activity after storage for 15 days at 5 °C and 40 % activity after five reaction cycles
Nanoporous gold electrodes for application in trace metal sensors, biosensors and biofuel cells
No full textNanoporous Gold (NPG) is a material of emerging interest for immobilization of
biomolecules and especially enzymes. NPG materials provide a high surface area onto
which biomolecules can either be directly physisorbed, covalently linked after first
modifying the NPG with a self-assembled monolayer (SAM) or entrapped in a polymer
matrix. The immobilization of enzymes while using NPG substrate material is being
pursued for applications in sensors, assays, supported synthesis, catalysis and biofuel
cells. NPG materials can be prepared by using many different approaches. However, the
most common method used is the dealloying of a low carat gold alloy containing
between 20-50 atomic % gold in a strong acid (70% HNO3), which oxidizes the least
noble metal, removing it from the alloy. The rapid rearrangement of the gold atoms at
the solid/liquid interface leaves behind the characteristic surface morphology. The
resultant structure consists of interconnected ligaments and pores with typical widths
between 5-200 nm. The surface area of these materials can be up to 500 times higher
than their geometric area.
Surface addressability of NPG is crucial for functionalization and surface
modification for the use in sensors, biosensors and biofuel cells. Full addressability of
the surface area of NPG was observed with small molecules such as sulphuric acid. The
surfaces could also be modified using bulky anthraquinone functional groups attached
on activated diazonium salts throughout the whole structure. Surface modification of
NPG has been achieved using a variety of strategies, such as through SAM formation of
thiol compounds, electro-reduction of in situ synthesized diazonium compounds and the
drop-casting or electro-polymerization of osmium redox polymers and hydrogels.
Surface functionalized NPG could be used for a variety of applications. Bulky
negatively charged sulfonate groups could therefore attract positively charged free trace
metal ions (such as Cu2+) in solutions for direct detection at the electrode surface. The
sensor displayed a detection range from 0.2 to at least 25 µM which is within the legal
concentration limit of 20.5 µM (1300 ppb) in drinking water (United States, EPA). The
sensitivity and limit of detection (LOD) were found to be 8.18 µA cm-2 µM-1 and 18.9
nM (~1.2 ppb) respectively. The BDS surface functionalization was also capable of
blocking biofouling material from the electrode surface, making it possible to measure
in complex media such as artificial human serum. Fructose dehydrogenase (FDH) could
be covalently attached to carboxylic acid terminated diazonium compounds for the
precise detection of D-fructose concentrations in a range of natural sweeteners and
beverages. The sensor was able to give accurate readings within 5 seconds with a linear
range of 0.05 - 0.3 mM D- fructose concentration, a sensitivity of 3.7 ± 0.2 μA cm-2
mM-1 and a LOD of 1.2 μM. When combining anodic enzymes, such as glucose
dehydrogenase (GDH) and FDH, with cathodic enzymes such as bilirubin oxidase
(BOD), enzymatic biofuel cells with considerable power outputs can be obtained.
GDH/MvBOD EFCs generated power densities of up to 17.5 and 7.0 μW cm-2 in PBS
and artificial serum, respectively, at an OCV of ~0.45 V (vs Ag/AgCl) with a
concentration of 5 mM D-glucose. These EFCs retained over 60% of their initial power
density after 8 hours of continuous operation. FDH/BpBOD EFCs generated power
densities of up to 13 µW cm-2 at an operating potential of 0.18 V vs Ag/AgCl at a
concentration of 10 mM D-fructose. The half-life was found to be ca. 19 h
Nanoporous gold electrodes as matrices for enzyme immobilization for application in biosensors and biofuel cells
No full textRobust nanoporous gold electrodes were fabricated by sputtering a gold-silver
alloy onto a glass support and subsequent dealloying of the silver component. Alloys
were prepared with either a non-uniform or uniform distribution of silver alloy which
showed clear differences in morphology on characterization with scanning electron
microscopy. The surface area of these electrodes was up to 28 times that of the
geometric surface area. The surface area accessible to modification by redox proteins
was determined using cyt c as a model system. Covalent immobilization of cyt c at
SAMs modified planar and nanoporous gold electrodes resulted in ca. 9 and 11 times
higher surface coverages at uniform and non-uniform nanoporous gold, respectively,
than at planar gold electrodes.
Well defined mediatorless bioelecytrocatalytic reduction of oxygen was
obtained on nanoporous gold electrodes prepared using a vacuum method and
subsequently modified with Myrthecium verrucaria bilirubin oxidase (MvBOD).
Diffusion limited faradaic response, with current densities of 0.8 mA/cm2, was observed
when the enzyme modified electrode was stabilized with a layer the P017-epoxy
polymer. The enzyme, Trametes hirsuta laccase (ThLc) also displayes direct electron
transfer at unmodified nanoporous gold electrodes. The observed current densities of
ca. 0.03 mA/cm2 were 10 times higher than the current densities at the ThLc modified
electrode made by drop-casting and are in contrast to the absence of a response at
unmodified polycrystalline gold electrodes.
Nanoporous and planar gold electrodes modified with Aspergillus niger glucose
oxidase (GOx) and Corynascus thermophilus cellobiose dehydrogenase (CtCDH)
together with Os redox mediators and PEGDGE as a cross-linking agent resulted in
glucose and lactose detecting biosensors. The sensors had (Imax,app, KM,app, sensitivity),
limits of detection (LOD). GOx and CtCDH modified electrodes were utilized as
anodes with MvBOD and Melanocarpus albomyces Lc (rMaLc) modified cathodes in
biofuel cells. A maximal power density of 41 μA/cm2 for the CtCDH/MvBOD biofuel
cell in 5 mM lactose and O2 saturated buffer was obtained. The power densities of the
biofuel cells were also evaluated in artificial plasma, where decreased values were
observed
Tailored adsorption of enzymes onto mesoporous silicates.
No full textEnzymes often display high regio- and chemoselectivity and therefore are of great
interest for industrial processes. However, in these processes, they can show low
stability and are not reusable. Immobilisation can optimize stability and activity, providing controlled mass transport of substrate and enabling re-use of the biocatalyst. Immobilisation also allows for continuous processes. Mesoporous
silicates provide a means of immobilizing proteins and enzymes in a stable
environment while retaining physiological function. Mesoporous silicates possess
large surface areas, highly ordered pore structures and a very narrow pore size
distribution. Pore sizes range from 3-20nm making them ideal for protein
encapsulation. One recurrent problem with physi-sorption onto mesoporous silicates
is leaching of the enzyme. Covalent attachment can reduce leaching but can increase rigidity which could lead to a loss in activity. Methods of tailoring the properties of both the mesoporous silicate surface and the protein surface to improve adsorption and catalytic activity have been explored with a view to generate a stable
biocatalyst. A biocatalyst has been generated through the tailored adsorption of a tagged enzyme onto a metal functionalized mesoporous silicate. By employing a
standard method of protein purification, a His-tagged enzyme has been immobilized
onto a nickel functionalized mesoporous silicate. Enzymes that have been studied
include subtilisin E, hydroxynitrile lyase and bile salt hydrolase. Denatured Histagged
subtilisin E was successfully immobilized onto nickel functionalized SBA
15. His-tagged bile salt hydrolase was successfully immobilized onto the surface of
nickel functionalized MCF and SBA 15. Manihot esculenta hydroxynitrile lyase
could not be immobilised with a histidine tag as the tag was not exposed for binding
- …
