pH-dependent release property of dioleoylphosphatidyl ethanolamine liposomes

pH-sensitive liposomes were prepared by a detergent removal method. Dioleoylphosphatidylethanolamine (DOPE) and cholesteryl hemisuccinate (CHEMS) were combined for the preparation of the liposomes so that the molar ratios of DOPE to CHEMS are 9/1, 8/2, 6/4 and 5/5. On transmission electron micrographs, hundreds of nm sized-multilamella vesicles were observed. The degrees of fluorescence quenching were approximately 70–80%, indicating that closed vesicles were formed. According to the results of the pH-dependent release experiment with the liposome composed of DOPE/CHEMS (5/5), no significant release was observed in the pH region ranging from 6 to 8. At pH of 5, an appreciable amount of calcein was released. The patterns of pH-dependent releases from liposomes composed of DOPE/CHEMS (6/4) and DOPE/CHEMS (8/2) were almost the same as those from liposomes composed of DOPE/ CHEMS (5/5). With the liposomes composed of DOPE/CHEMS (9/1), unlike the other liposomes described above, almost 90% release was observed at pH 6. In this case, since the amount of a complementary molecule, CHEMS, is relatively small, the liposomes would be subjective to destabilization even at a small change in the degree of deionization of the carboxylic group. This may explain why the liposome of DOPE/CHEMS (9/1) exhibits a significant release at a relatively high pH, pH 6.0.     

Facile direct electron transfer in glucose oxidase modified electrodes

Glucose oxidase (GOx) is widely used in the glucose biosensor industry. However, mediatorless direct electron transfer (DET) from GOx to electrode surfaces is very slow. Recently, mediatorless DET has been reported via the incorporation of nanomaterials such as carbon nanotubes and nanoparticles in the modification of electrodes. Here we report GOx electrodes showing DET without the need for any nanomaterials. The enzyme after immobilization with poly-l-lysine (PLL) and Nafion^® retains the biocatalytic activities and oxidizes glucose efficiently. The amperometric response of Nafion-PLL-GOx modified electrode is linearly proportional to the concentration of glucose up to 10 mM with a sensitivity of 0.75 μA/mM at a low detection potential (−0.460 V vs. Ag/AgCl). The methodology developed in this study will have impact on glucose biosensors and biofuel cells and may potentially simplify enzyme immobilization in other biosensing systems.     

pH-controllable bioelectrocatalysis of glucose by glucose oxidase loaded in weak polyelectrolyte layer-by-layer films with ferrocene derivative as mediator

Oppositely charged poly(allylamine hydrochloride) (PAH) and hyaluronic acid (HA) were assembled into {PAH/HA}_n layer-by-layer (LBL) films on pyrolytic graphite (PG) electrodes. Glucose oxidase (GOD) in solution was then loaded into the films, designated as {PAH/HA}_n-GOD. When the {PAH/HA}_n-GOD film electrodes were placed in pH 5.0 buffers containing ferrocenedicarboxylic acid (Fc(COOH)₂) and glucose, a well-defined and large cyclic voltammetric (CV) oxidation wave of glucose catalyzed by GOD immobilized in the films and mediated by Fc(COOH)₂ in solution was observed. However, when the films were placed in pH 9.0 buffers containing the same amount of Fc(COOH)₂ and glucose, the electrocatalytic response was quite small. The bioelectrocatalysis for the film system was at the “on” state at pH 5.0 and at the “off” state at pH 9.0. This pH-sensitive “on-off” behavior was reversible and could be repeated for several times. The possible mechanism of the pH-switchable bioelectrocatalysis was explored and discussed, and should be mainly attributed to the different electrostatic interaction between Fc(COOH)₂ and the films at different pH. This work provides a novel model to realize pH-controllable bioelectrocatalysis based on the enzyme-loaded LBL assembly films, and may guide us to develop the tunable electrochemical biosensors based on electrocatalysis with immobilized enzymes.     

Direct electrochemistry of glucose oxidase assembled on graphene and application to glucose detection

The direct electrochemistry of glucose oxidase (GOx) integrated with graphene was investigated. The voltammetric results indicated that GOx assembled on graphene retained its native structure and bioactivity, exhibited a surface-confined process, and underwent effective direct electron transfer (DET) reaction with an apparent rate constant (k_s) of 2.68 s⁻¹. This work also developed a novel approach for glucose detection based on the electrocatalytic reduction of oxygen at the GOx-graphene/GC electrode. The assembled GOx could electrocatalyze the reduction of dissolved oxygen. Upon the addition of glucose, the reduction current decreased, which could be used for glucose detection with a high sensitivity (ca. 110 ± 3 μA mM⁻¹ cm⁻²), a wide linear range (0.1-10 mM), and a low detection limit (10 ± 2 μM). The developed approach can efficiently exclude the interference of commonly coexisting electroactive species due to the use of a low detection potential (−470 mV, versus SCE). Therefore, this study has not only successfully achieved DET reaction of GOx assembled on graphene, but also established a novel approach for glucose detection and provided a general route for fabricating graphene-based biosensing platform via assembling enzymes/proteins on graphene surface.     

脂质体与化妆品

文章概述了脂质体的性能、作用原理、制备及在化妆品中的应用。     

Direct electron transfer and electrocatalysis of glucose oxidase immobilized on glassy carbon electrode modified with Nafion and mesoporous carbon FDU-15

In this paper, it was found that glucose oxidase (GOD) has been stably immobilized on glassy carbon electrode modified with mesoporous carbon FDU-15 (MC-FDU-15) and Nafion by simple technique. The sorption behavior of GOD immobilized on MC-FDU-15 matrix was characterized by transmission electron microscopy (TEM), ultraviolet-visible (UV-vis), FTIR, respectively, which demonstrated that MC-FDU-15 could facilitate the electron exchange between the active center of GOD and electrode. The direct electrochemistry and electrocatalysis behavior of GOD on the modified electrode were characterized by cyclic voltammogram (CV) which indicated that GOD immobilized on Nafion and MC-FDU-15 matrices display direct, reversible and surface-controlled redox reaction with an enhanced electron transfer rate constant of 4.095 s⁻¹ in 0.1 M phosphate buffer solution (PBS) (pH 7.12). Furthermore, it was also discovered that, in the presence of O₂, GOD immobilized on Nafion and MC-FDU-15 matrices could produce a linear response to glucose. Thus, Nafion/GOD-MC-FDU-15/GC electrode is hopeful to be used in glucose biosensor. In addition, GOD immobilized on MC-FDU-15 and Nafion matrices possesses an excellent bioelectrocatalytic activity for the reduction of O₂. So, the Nafion/GOD-MC-FDU-15/GC electrode can be utilized as the cathode in biofuel cell.     

Dual oxygen and Ir oxide regeneration of glucose oxidase in nanostructured thin film glucose sensors

A nanoparticulate iridium oxide (IrOx) thin film has been developed as a redox-active matrix material for an advanced generation glucose biosensor, in which IrOx serves as the non-physiological mediator, replacing oxygen in the enzymatic re-oxidation of glucose oxidase (GOx). Ethanolic solutions of Nafion and an Ir sol were mixed with an aqueous GOx solution and then deposited on a Au support. The Ir nanoparticles were then oxidized electrochemically to IrOx and the resulting films (IrOx-GOx-Nafion) were tested for their glucose response in both oxygen- and argon-saturated solutions, with the oxygen content in both solutions monitored by a Pt electrode. The sensors that are regenerated largely by O₂ are characterized by a Michaelis-Menten K^′m value of ∼30 mM or more and i_max values of at least 20 μA cm⁻². Under fully deareated conditions, the sensors lose only ∼50% of their response to glucose, clearly indicating that a dual oxygen-regeneration and IrOx mediation mechanism is operative for the biosensor under these conditions. Under optimized conditions, involving a controlled GOx:Ir ratio, only the Ir oxide sites in the film serve to mediate GOx regeneration, giving K^′m (10-15 mM) and i_max values that are independent of the O₂ content of the solution.     

葡萄糖氧化酶在修饰碳纳米管上的电化学

将葡萄糖氧化酶（GOx）分别固定在多壁碳纳米管（MWNT）、氨基化碳纳米管（AMWNTs）和羧基化碳纳米管（MWNTs-COOH）修饰的电极表面,电化学测量表明固定在羧基和氨基碳纳米管上的GOx式量电位基本没变,而峰电流得到了很大提高。尤其是氨基化碳纳米管上的GOx的峰电流是未功能化碳管上GOx的4倍多。进一步研究Nafion/GOx-AMWNTs/GC电极的电化学行为,发现固定在AMWNTs上的GOx可进行直接准可逆的氧化还原反应,而且固定在AMWNTs上的GOx有良好的稳定性。氨基改性碳纳米管电极载体材料有望显著提高GOx生物燃料电池性能。     

Direct electron transfer from glucose oxidase immobilized on a nano-porous glassy carbon electrode

A pair of well-defined and reversible redox peaks was observed for the direct electron transfer (DET) reaction of an immobilized glucose oxidase (GOx) on the surface of a nano-porous glassy carbon electrode at the formal potential (E°′) of −0.439 V versus Ag/AgCl/saturated KCl. The electron transfer rate constant (k_s) was calculated to be 5.27 s⁻¹. The dependence of E°′ on pH indicated that the direct electron transfer of the GOx was a two-electron transfer process, coupled with two-proton transfer. The results clearly demonstrate that the nano-porous glassy carbon electrode is a cost-effective and ready-to-use scaffold for the fabrication of a glucose biosensor.     

Viscoelastic and swelling properties of glucose oxidase loaded polyacrylamide hydrogels and the evaluation of their properties as glucose sensors

Here we report the immobilization of glucose oxidase in polyacrylamide hydrogels carried out by aqueous crosslinking copolymerization of acrylamide and N,N′ methylene bisacrylamide in the presence of the enzyme. The swelling and viscoelastic properties of the hydrogels were evaluated as a function of the content of crosslinker of the polymer chains and enzyme concentration. Amperometric measurements were also carried out to evaluate the system as a glucose biosensor.     

Photo-electro-chemical properties of TiO2 mediated by the enzyme glucose oxidase

Electrochemical measurements show that the enzyme glucose oxidase (GO) is adsorbed on the surface of TiO₂ without apparently changing the flat band potential of the semiconductor, indicating that it does not cause a change of the energy of conduction band electrons. On the other hand, it is observed that GO markedly increases the efficiency of the two electron reduction of O₂ to H₂O₂ which is accumulated in the solution phase.

ESR spin trapping investigations indicate that GO favors the formation of OH radicals, due to either the inhibition of charge recombination processes or to H₂O₂ reduction by conduction band electrons. Accordingly, photo-oxidation of different alcohols to the corresponding radical species is also enhanced in the presence of GO.

The photo-oxidation of 1,2-propandiol on TiO₂/GO is regioselective in that (i) partial oxidation to hydroxyacetone is observed and (ii) no mineralization (full combustion to CO₂) of the substrate occurs. These facts are of particular interest in the field of studies concerning the design of new photocatalytic systems with enhanced activity and controllable oxidative power.     

Immobilization of glucose oxidase and platinum on mesoporous silica nanoparticles for the fabrication of glucose biosensor

In this study, amino group modified mesoporous silica nanoparticles (MSN) were prepared and used to immobilize both platinum nanoparticles (PtNP) and glucose oxidase (GOx). The prepared MSN–PtNP demonstrated high stability and reactivity for catalyzing H₂O₂ electro-reduction, mainly due to the large amount of PtNP immobilized, the high surface area of these catalysts and the unique nanostructures formed through the synthetic route. Working at −0.2 V, the linear range in response to H₂O₂ by the prepared MSN–PtNP can be 5 × 10⁻⁷ to 6 × 10⁻² mol L⁻¹. After immobilizing GOx onto MSN–PtNP, the resulting MSN–PtNP–GOx was capable of interference-free determination of glucose with the linear range as wide as 1 × 10⁻⁶ to 2.6 × 10⁻² mol L⁻¹. Furthermore, the fabricated glucose biosensor can offer significant advantages compared with its conventional counterparts, typically like the high sensitivity, good reproducibility and stability, and rapid response ability as well. The fabricated glucose biosensor demonstrated its potential in clinical applications, so as to enable the determination of glucose in real serum samples.     

Electrochemistry of glucose oxidase immobilized on the carbon nanotube wrapped by polyelectrolyte

A more stably dispersing of multi-wall carbon nanotube composite (noted as PDDA-MWNT), which was obtained by wrapping the MWNT with poly(diallydimethylammonium) chloride (PDDA), was used for the immobilization of glucose oxidase (GOD) and its bioelectrochemical studies. The morphologies and structures of the PDDA-MWNT composite were characterized by environment scanning electron microscopy (ESEM) and X-ray photoelectron spectroscopy (XPS). Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry were used to feature the GOD adsorbed onto the electrode modified by PDDA-MWNT composite. The immobilized GOD at the PDDA-MWNT films exhibited a pair of well-defined nearly reversible redox peaks and a fast heterogeneous electron transfer rate with the rate constant (k_s) of 2.76 s⁻¹. In addition, GOD immobilized in this way retained its bioelectrocatalytic activity for the oxidation of glucose. The method of immobilizing GOD without any additional cross-linking agents presented here is easy and facile, which provides a model for other redox enzymes and proteins.     

Effects of 1-butyl-3-methylimidazolium tetrafluoroborate on the oxidation of glucose catalyzed by glucose oxidase

The effects of room temperature ionic liquids (ILs) on the conformation and electrocatalytic activity of enzymes were studied using glucose oxidase (GOx) and 1-butyl-3-methylimidazolium tetrafluoroborate ([bmim]BF₄) as models. UV-vis and circular dichroic (CD) spectra indicated that [bmim]BF₄ did not affect the conformation of the enzyme, and the secondary structure of GOx in [bmim]BF₄-PBS mixtures (the content of the IL is from 0 to 20 vol%) was essentially the same as that of the native one. The Raman spectra showed that no interaction existed between glucose and [bmim]BF₄. The oxidation of glucose catalyzed by GOx was investigated under a substrate-saturated condition in [bmim]BF₄-PBS mixtures using ferroceneacetic acid (FcA) as a mediator. The voltammetric results showed that electrocatalytic current (i_cat) decreased with the increase of the content of [bmim]BF₄ in the mixtures. The reason causing the decrease of i_cat was analyzed. The reduction in the diffusion rate of FcA due to the increase of the viscosity after the addition of the IL was a key factor of causing the decrease of i_cat. The results presented here will be useful for the designing of the related biosensor used in ILs-containing system.     

Immobilization of glucose oxidase and acetylcholinesterase onto modified polyamide sorbent

Two types of polyamide (PA) sorbents with high specific area were prepared. The effects of solvent type, concentrations of formic acid, and polymer on the porosity characteristics were studied. The sorbent with the highest specific area was obtained by using C₂H₅OH—HCOOH solvent (60% HCOOH) and the rest of the experiments were carried out with this type of sorbent. The possibility of applying the PA sorbent as carrier for immobilization of glucose oxidase (GOD) and acetylcholinesterase (AChE) was investigated. In order to increase the active groups content (necessary for enzyme immobilization), the sorbent was modified with dimethylaminoethylmethacrylate (DMAEM) and 2-acrylamido-2-methylpropensulfonic acid. The amount of the active groups introduced during the modification and the degree of hydrophilicity were determined. The quantity of bound protein and relative activity of GOD and AChE immobilized onto unmodified and modified sorbents were studied. Optimum pH and temperature of the immobilized GOD and AChE were also determined. The influence of three phosphoroorganic compounds on the activity of the immobilized AChE was investigated. Tetrachlorvinvos was found to be the strongest inhibitor, while AChE immobilized onto PA sorbent modified with DMAEM showed the highest stability. The possibility of using immobilized GOD and AChE in a flow-injection system for determination of the concentrations of glucose and phosphoroorganic compounds was studied. © 1998 John Wiley & Sons, Inc. J Appl Polym Sci 68:323–329, 1998     

Layer by layer assembly of glucose oxidase and thiourea onto glassy carbon electrode: Fabrication of glucose biosensor

For the first time a novel, simple and facile approach is described to construct highly stable glucose oxidase (GOx) multilayer onto glassy carbon (GC) electrode using thiourea (TU) as a covalent attachment cross-linker. The layer by layer (LBL) attachment process was confirmed by cyclic voltammetry, electrochemical impedance spectroscopy and Fourier transform infrared reflection spectroscopy (FT-IR-RS) techniques. Immobilized GOx shows excellent electrocatalytic activity toward glucose oxidation using ferrocenemethanol as artificial electron transfer mediator and biosensor response was directly correlated to the number of bilayers. The surface coverage of active GOx per bilayer, heterogeneous electron transfer rate constant (k_s) and Michaelis–Menten constant (K_M), of immobilized GOx were 1.50 × 10⁻¹² mol cm⁻², 9.2 ± 0.5 s⁻¹ and 3.42(±0.2) mM, respectively. The biosensor constructed with four-bilayers of TU/GOx showed good stability, high reproducibility, long life-time, fast amperometric response (5 s) with the high sensitivity of 5.73 μA mM⁻¹ cm⁻² and low detection limit of 6 μM at concentration range up to 5.5 mM.     

Surface activity of hydrophobically modified alternating copolymers

Surface activity of N-monopropyl maleamic acid-alt-styrene (NPMA-alt-S) and N-monodecyl maleamic acid-alt-styrene (NDMA-alt-S) was studied. Surface pressure-area isotherms (Π-A) at the air/water interface were determined. It was found that the pH of the water subphase and the chemical structure of the copolymers studied strongly influence the type of isotherms. It was also found that the driving force for the adsorption of NPMA-alt-S, water soluble polymer at the air/water interface arises from an entropic contribution.     

可膨胀石墨催化合成丙烯酸甲酯

可膨胀石墨对丙烯酸甲酯的合成具有较高的催化活性。最佳条件为：甲醇与丙烯酸的物质的量比为1．15：1，催化剂用量为醇和酸总质量的5．56％，反应时间为55min，丙烯酸甲酯收率可达98％以上。     

Glucose-Sensitive Liposomes Incorporating Hydrophobically Modified Glucose Oxidase

Glucose-sensitive liposomes were prepared by incorporating hydrophobically modified glucose oxidase (EC 1.1.3.4.) into the liposomal bilayer of dioleoylphosphatidylethanolamine and cholesteryl hemisuccinate. For the release test, calcein, a fluorescence marker, was entrapped in the liposomes. The liposomes were stable under neutral conditions in terms of calcein release but an extensive release was observed under acidic conditions. In the experiment of glucose concentration-dependent calcein release, no release was observed for 180 min when the suspension of liposome was free of glucose. With a glucose concentration of 50 mg/dL, no appreciable amount of calcein was released for the first 20 min, and then the release rate was accelerated. At 200 mg/dL glucose concentration which is diagnostic and indicative for insulin-dependent diabetes, the lag time of calcein release became shorter and a faster response was obtained. When glucose concentration further increased to 400 mg/dL, the calcein release rate and the degree of release in 180 min were almost the same as the values when the glucose concentration was 200 mg/dL. The glucose concentration-dependent release is due to pH change, since the suspension of liposomes became acidic during the release experiments.     

Photo‐responsive monoolein cubic phase incorporating hydrophobically modified poly(vinyl alcohol)–coumarin conjugate

Photo‐responsive monoolein cubic phases were developed by immobilizing hydrophobically modified poly (vinyl alcohol)–epoxypropoxy coumarin conjugate (HmPVA–EPC) in the water channels of the cubic phase. On the NMR spectrum of HmPVA‐EPC, the molar ratio of PVA/decanoyl chloride (a hydrophobic anchor)/EPC was determined to be about 1:0.64:1.45. The interfacial activity of PVA–EPC conjugate was much lower than that of PVA, but the interfacial activity of HmPVA–EPC was as high as that of PVA. The critical micelle concentrations of HmPVA–EPC and PVA were about 0.005%. Under the ultraviolet (UV) irradiation (λ = 254 nm), EPC residues of HmPVA were readily dimerized and the dimerization degree in 180 min was about 33%. The irradiation of UV light had no significant effect on the release of fluorescein isothiocyanate–dextran (a fluorescence marker) from cubic phase free of polymer and cubic phase incorporating HmPVA, but it suppressed the release from cubic phase incorporating HmPVA–EPC. The photo crosslinking of HmPVA–EPC in the water channels is believed to be responsible for the suppressed release. POLYM. ENG. SCI., 54:227–233, 2014. © 2013 Society of Plastics Engineers