Fabrication of nanoporous nanocomposites entrapping Fe3O4 magnetic nanoparticles and oxidases for colorimetric biosensing

A nanostructured multicatalyst system consisting of Fe(3)O(4) magnetic nanoparticles (MNPs) as peroxidase mimetics and an oxidative enzyme entrapped in large-pore-sized mesoporous silica has been developed for convenient colorimetric detection of biologically important target molecules. The construction of the nanocomposites begins with the incorporation of MNPs on the walls of mesocellular silica pores by impregnating Fe(NO(3))(3)·9H(2)O, followed by the immobilization of oxidative enzymes. Glutaraldehyde crosslinking was employed to prevent enzymes leaching from the pores and led to over 20 wt% loading of the enzyme. The oxidase in the nanocomposite generates H(2)O(2) through its catalytic action for target molecules and subsequently activates MNPs to convert selected substrates into colored products. Using this strategy, two different biosensing systems were constructed employing glucose oxidase and cholesterol oxidase and their analytical capabilities were successfully verified by colorimetrically detecting the corresponding target molecules with excellent selectivity, sensitivity, reusability, and stability. Future potential applications of this technology range from biosensors to multicatalyst reactors.     

A novel colorimetric determination of reduced glutathione in A549 cells based on Fe3O4 magnetic nanoparticles as peroxidase mimetics

In this paper, a novel and simple colorimetric method for the determination of reduced glutathione (GSH) based on Fe(3)O(4) magnetic nanoparticles (MNPs) as peroxidase mimetics was developed. The Fe(3)O(4) MNPs prepared via a coprecipitation method, which possess intrinsic peroxidase-like activity, were used as a catalyst in the color development reaction of a peroxidase substrate 2,2'-azino-bis(3-thylbenzo-thiazoline-6-sulfonic acid) diammonium salt (ABTS) and H(2)O(2). The existence of GSH can consume H(2)O(2) and cause a color change of the reaction system which can be detected by the naked eye. Accordingly, the GSH can be detected by measuring the wastage of H(2)O(2). A good linear relationship was obtained from 3.0 to 30.0 μM for GSH. Good recoveries ranging from 96.7 to 107% were obtained. Furthermore, it was used to detect GSH in A549 cells.     

G-Quadruplex-based DNAzyme for colorimetric detection of cocaine: using magnetic nanoparticles as the separation and amplification element

The appearance of the aptamer provides good recognition elements for small molecules, especially for drugs. In this work, by combining the advantages of magnetic nanoparticles (MNPs) with colorimetric drug detection using hemin-G-quadruplex complex as the sensing element, we report a simple and sensitive DNAzyme-based colorimetric sensor for cocaine detection in a 3,3,5,5-tetramethylbenzidine sulfate (TMB)-H(2)O(2) reaction system. The whole experimental processes are simplified. Cocaine aptamer fragments, SH-C2, are covalently labeled onto the amine-functionalized MNPs. When the target cocaine and another cocaine aptamer fragments (C1) grafted with G-riched strand AG4 (i.e. C1-AG4) are present simultaneously, the C2 layer on MNPs hybridizes partly with C1-AG4 to bind the cocaine. The C1-AG4 can be combinded with hemin to form DNAzyme which can effectively catalyze the H(2)O(2)-mediated oxidation of TMB, giving rise to a change in solution color. Importantly, using MNPs as the separation and amplification elements could effectively reduce the background signal and the interference from the real samples. A linear response from 0.1 μM to 20 μM is obtained for cocaine and a detection limit of 50 nM is achieved, which provides high sensitivity and selectivity to detect cocaine.     

Stable-isotope dilution analysis of galactose metabolites in human erythrocytes

An established gas chromatography/mass spectrometry (GC/MS) method, devised for stable-isotope dilution analysis of plasma galactose, was developed to allow determination of erythrocyte (red blood cell, RBC) concentrations of galactose-1-phosphate and other primary metabolites relevant in galactosaemia. Galactose-1-phosphate was enzymatically converted to galactose, and the aldononitrile pentaacetate derivative was separated by gas chromatography and determined by mass spectrometry using chemical ionisation and selected ion monitoring of the [MH-60](+) ion. U-(13)C-Labelled standard was used for quantification. Comparative measurements were conducted using established fluorimetric and radiometric enzymatic methods. The GC/MS analysis for galactose-1-phosphate was linear (range examined 0-600 micromol/L(RBC), packed cells), of acceptable repeatability at low and high concentrations (within and between run CVs <15%), with a limit of quantification of 0.01 micromol/L(RBC). With samples from patients with classical galactosaemia there was a linear correlation with conventional enzymatic assays (r(2) > 0.927). In erythrocytes from post-absorptive patients under treatment, Q188R-heterozygous parents, and healthy subjects, galactose-1-phosphate concentrations (mean +/- SD) were found to be 142 +/- 38 (n = 41), 1.4 +/- 0.2 (n = 8), and 1.9 +/- 0.5 (n = 33) micromol/L(RBC), respectively. In comparison, free galactose concentrations were 3.8 +/- 1.7, 0.49 +/- 0.19, and 0.43 +/- 0.20 mol/L(RBC), respectively. The procedure allowed simultaneous galactitol analysis and proved to be useful to trace incorporation of (13)C-label into erythrocyte galactose metabolites in a D-[1-(13)C]galactose in vivo turnover study.     

Development of a new diagnostic method for galactosemia by high-performance anion-exchange chromatography with pulsed amperometric detection

We developed a new non-derivatization analytical method for the determination of galactose in the diagnosis of galactosemia by high-performance anion-exchange chromatography (HPAEC)-pulsed amperometric detection (PAD). With an anion-exchange column, the analytes were separated efficiently using 3mM NaOH containing 1mM NaOAc, and 200mM NaOH was added for post-column reagent. The limit of detection (S/N=3) and limit of quantification (S/N=10) for galactose were 25ng/mL and 83ng/mL, respectively. Linear dynamic range was from 4.67mg/dL to 53.46mg/dL (r(2)=0.9999). The mean recovery of galactose for intra-, inter-day assays were found to be of satisfactory results (98.14-101.42%).     

Determination of galactose in human plasma by HPLC with electrochemical detection

Galactose in plasma from patients with hepatic diseases who had undergone low level galactose infusion was determined by using HPLC with electrochemical detection (LCEC). Agreement between galactose concentration determined by the LCEC and a fluorometric method was remarkably good at moderate levels of galactose in plasma. However, the fluorometric method is not suitable for samples containing very small amounts of galactose (blood from hepatic veins) and even for a few samples at moderate galactose content (blood from peripheral veins), suggesting the presence of an endogenous interference. There was no interference for the quantitation of galactose by the LCEC method, by virtue both of the specificity involved in the electrochemical detection and the separation by liquid chromatography. The detection limit of the LCEC method was 0.4 mg galactose/L blood.     

Magnetic solid‐phase extraction for determination of sulpiride in human urine and blood using high‐performance liquid chromatography

A novel and efficient sample preconcentration technique based on the Fe₃O₄ magnetic nanoparticles (Fe₃O₄ MNPs) coated with silica (SiO₂) has been developed for extraction and determination of sulpiride. The functionalized MNPs showed excellent dispersibility in aqueous solution and were applied to magnetic solid‐phase extraction of sulpiride from human urine and blood prior to high‐performance liquid chromatography analysis. The separation, preconcentration and desorption procedure was completed in 10 min. Optimal experimental conditions, including sample pH, the amount of the MNPs, eluent type and volume, and the ultrasonication time were studied and established. The method showed good linearity for the determination of sulpiride in the concentration range of 10–1000 ng/mL in urine and blood. The recovery of the method was in the range between 91.2 and 97.5%, and the limit of detection was 2 ng/mL for sulpiride in human blood and urine. The results indicated that the present procedure is a suitable pretreatment method for biological samples. Copyright © 2015 John Wiley & Sons, Ltd.     

Heterogeneous asymmetric Henry reaction using a chiral bis(oxazoline)-copper complex immobilized on magnetically separable mesocellular mesoporous silica support

A chiral bis(oxazoline) ligand was immobilized onto a magnetically separable hierarchically-ordered mesocellular mesoporous silica (M-HMMS) and this new catalytic system was examined in the asymmetric Henry reaction between various aldehydes and nitromethane at ambient temperature. Good enantioselectivity (upto 86.0% ee) was observed when the free silanol groups of the mesoporous silica were capped by trimethylsilyl group. This catalyst was separated magnetically and reused several times without significant loss of reactivity or enantioselectivity. This magnetic separation of catalysts could lead to further development toward practical industrial scale application due to simplicity without cumbersome filtration.     

配体交换法制备水中高分散稳定性的多羧基修饰Fe3O4纳米颗粒

采用油相高温分解法制备了粒径可控且单分散的油溶性Fe3O4磁性纳米粒子(MNPs-OA), 并通过配体交换对其表面进行了亲水性修饰, 制备了柠檬酸(CA)、 N-(三甲氧基硅丙基)乙二胺三乙酸钠(SiCOOH)、 丁烷四羧酸(BTCA)和乙二胺四乙酸 (EDTA)四钠4种多羧基配体修饰的水溶性Fe3O4磁性纳米粒子(MNPs-CA, MNPs-SiCOOH, MNPs-BTCA 和MNPs-EDTA), 其中首次选用四羧基配体BTCA和EDTA四钠来修饰Fe3O4磁性纳米粒子(MNPs). 对油溶性MNPs和4种水溶性MNPs的形貌、 结构、 化学组成和磁性能进行了表征, 并对4种多羧基配体修饰的水溶性MNPs在水相中的稳定性和分散性进行了表征. 结果表明, 所得MNPs的平均粒径为15 nm, 具有超顺磁性, 配体交换后的水溶性MNPs具有良好的亲水性, 并在弱酸～碱性很宽的pH范围内具备良好的分散稳定性. 此类多羧基修饰的水溶性MNPs可与适当的阳离子聚电解质进行组装, 从而得到在磁靶向载体和磁共振造影(MRI)显影中具有良好应用前景的磁性自组装微囊.     

磁固相萃取/气相色谱一质谱法分析水样中的16种邻苯二甲酸酯类化合物

合成了一种由Fe3 O4磁性纳米粒子(MNPs)和多壁碳纳米管(MWCNTs)组成的复合纳米材料,用于水样中16种邻苯二甲酸酯类化合物(PAEs)的磁固相萃取(MSPE),并结合气相色谱-质谱(GC - MS)法进行定量分析.合成的纳米材料用傅立叶变换红外光谱表征.为提高萃取效率,优化了解析溶剂的种类和用量、解析时间、...     

8-羟基喹啉功能化磁性粒子的制备及其应用于伏安法测定水样中痕量铅

将所制备的磁性Fe3O4颗粒先后分别在乙醇和水介质中与3-氨基丙基三乙氧基硅烷(APS)和10%(体积分数)戊二醛溶液反应使Fe3O4颗粒的表面形成醛基功能化,最后在pH 6.8的磷酸盐缓冲溶液中与8-羟基喹啉(Ox)反应,生成了8-羟基喹啉功能化的磁性Fe3O4颗粒。试验表明经Ox修饰的Fe3O4颗粒对Pb2+有很强的络合作用,据此将其作为痕量铅的富集试剂应用于阳极溶出伏安法测定水样中铅含量。取水样50.0mL,加入Fe3O4/Ox颗粒在pH 6.8磷酸盐缓冲溶液中的悬浮液1.5mL,充分搅拌20min后,用磁铁在反应烧杯底部吸着磁性颗粒使与水溶液分离。于磁性颗粒中加入1mol·L^-1盐酸溶液1mL,将其表面吸着的Pb^2+溶解,用水将溶液稀释至5.0mL。以玻碳电极为工作电极,Ag/AgCl为参比电极,铂电极为对电极进行阴极电沉积50s,然后进行阳极溶出伏安测定。用铅标准溶液绘制标准曲线,Pb^2+的质量浓度在0.50 mg·L^-1以内与所测得的对应氧化电流导数值之间呈线性关系,检出限(3S/N)为0.7μg·L^-1。应用此方法分析了6个环境水样,测定值与已知值相符,测定值的相对标准偏差(n=5)为0.48%~3.5%。按标准加入法进行回收试验,测得回收率为91.0%~138%。     

Synthesis and Study of the Properties of Magnetic Adsorbents Based on Hydrophobized Silica

Moscow University Chemistry Bulletin - We proposed a method for producing magnetic nanocomposite adsorbents, based on the adsorption of Fe3O4 magnetic nanoparticles (MNPs) on hydrophobized silicas...     

Synthesis of Small‐Sized,Porous, and Low‐Toxic Magnetite Nanoparticles by Thin POSS Silica Coating

In this communication, we report the synthesis of small‐sized (<10 nm), water‐soluble, magnetic nanoparticles (MNPs) coated with polyhedral oligomeric silsesquioxanes (POSS), which contain either polyethylene glycol (PEG) or octa(tetramethylammonium) (OctaTMA) as functional groups. The POSS‐coated MNPs exhibit superparamagnetic behavior with saturation magnetic moments (51–53 emu g^?1) comparable to silica‐coated MNPs. They also provide good colloidal stability at different pH and salt concentrations, and low cytotoxicity to MCF‐7 human breast epithelial cells. The relaxivity data and magnetic resonance (MR) phantom images demonstrate the potential application of these MNPs in bioimaging.     

Prussian blue nanoparticles doped nanocage for controllable immobilization and selective biosensing of enzyme

A conductive nanocage composed of Prussian blue (PB) nanoparticles doped mesocellular silica–carbon foam was prepared. This nanocage possessed ink-bottlelike structure with narrow and controllable pore-size distribution, good biocompatibility and favorable conductivity. The Prussian blue nanoparticles dispersed homogeneously on the mesowalls and provided the nanocage with highly catalytic ability toward the reduction of hydrogen peroxide. The nanocage could be used for volume-selective entrapment of enzyme to prepare a biosensor. The use of the Prussian blue nanoparticles doped nanocage would open new horizons for fabrication of biosensors and biocatalysts.     

Determination of galactose and mannose residues in natural galactomannans using a fast and efficient high-performance liquid chromatography/UV detection

The present work describes the validation of an easy, fast and efficient precolumn derivatization method for the quantification of oligosides, mannose and galactose obtained by degradation of galactomannans. This work combines an acid hydrolysis and an enzymatic degradation of natural galactomannans with the quantification of released residues by reversed-phase HPLC-UV, the most usual HPLC system in laboratories. In case of enzymatic degradation, mannotetraose has been detected and quantified for the first time, and an application to the evaluation of the galactosyl distribution in galactomannans is proposed. After an acidic hydrolysis, this method also allowed to obtain the mannose/galactose (M/G) ratio.     

Preparation of highly fluorescent magnetic nanoparticles for analytes-enrichment and subsequent biodetection

Bifunctional nanoparticles with highly fluorescence and decent magnetic properties have been widely used in biomedical application. In this study, highly fluorescent magnetic nanoparticles (FMNPs) with uniform size of ca. 40 nm are prepared by encapsulation of both magnetic nanoparticles (MNPs) and shell/core quantum dots (QDs) with well-designed shell structure/compositions into silica matrix via a one-pot reverse microemulsion approach. The spectral analysis shows that the FMNPs hold high fluorescent quantum yield (QY). The QYs and saturation magnetization of the FMNPs can be regulated by varying the ratio of the encapsulated QDs to MNPs. Moreover, the surface of the FMNPs can be modified to offer chemical groups for antibody conjugation for following use in target-enrichment and subsequent fluorescent detection. The in vitro immunofluorescence assay and flow cytometric analysis indicate that the bifunctional FMNPs-antibody bioconjugates are capable of target-enrichment, magnetic separation and can also be used as alternative fluorescent probes on flow cytometry for biodetection.     

Heavily fluorinated carbohydrates as enzyme substrates: oxidation of tetrafluorinated galactose by galactose oxidase

Galactose oxidase (GOase) was shown to oxidise several C2/C3 fluorinated galactose analogues. Interestingly, the enzyme was able to distinguish between the 2,3-tetrafluorinated galactose and its epimeric glucose analogue, and this represents the first reported biotransformation of a heavily fluorinated sugar.     

Colorimetric determination of radical scavenging activity of antioxidants using Fe3O4 magnetic nanoparticles

We report the first use of iron oxide magnetic nanoparticles (Fe₃O₄ MNPs) as a novel, alternative, simple and reliable agents for colorimetric measurement of radical scavenging activity of the antioxidants. In the presence of H₂O₂ and the peroxidase colorimetric substrate, Fe₃O₄ MNPs catalyzed the oxidation of colorless peroxidase substrate to form colorimetric products via the generation of hydroxyl radicals. After adding antioxidants, the catalytic activity of Fe₃O₄ MNPs was inhibited due to scavenging of hydroxyl radicals by the antioxidants, producing less colorimetric products resulting in the reduction of color intensity. Two model antioxidant standards including gallic acid (GA) and epigallocatechin gallate (EGCG) were successfully evaluated for their hydroxyl radical scavenging activity using the developed assay. The performance of the developed method was validated against traditional antioxidant assays for 9 tea samples. Using the Spearman rank correlation coefficient method, the antioxidant activity of tea samples obtained from the Fe₃O₄ MNP assay correlated well with the traditional assays at the 95% confidence level. Furthermore, the developed assay was successfully carried out on a paper-based device to provide for high throughput analysis with low amounts of reagents and sample consumption and low analysis cost for screening of radical scavenging activity of the antioxidants. The results from the analysis of antioxidant activity in tea samples obtained from the Fe₃O₄ MNP paper-based assay were not significantly different to those obtained from the developed Fe₃O₄ MNP spectrophotometric assay indicating that the developed assay was also applicable in a low-cost analysis platform.     

共价固定亲和素的磁性纳米粒子用于快速测定蛋白A含量

采用溶剂热法制备了Fe₃O₄磁性纳米粒子(MNPs), 以戊二醛为交联剂, 将亲和素共价固定于MNPs表面. 用透射电子显微镜(TEM)、 X射线衍射(XRD)、 紫外-可见吸收光谱(UV-Vis)、 傅里叶变换红外光谱(FTIR)和荧光光谱等手段对蛋白固定过程进行了监控和表征. 采用荧光光谱法评价了固定亲和素的磁性纳米粒子(Avi-MNPs)的活性, 并将Avi-MNPs应用于分光光度法测定蛋白A的含量. TEM结果表明, 功能化前后MNPs的粒度分布均匀, 粒径大小分别约为30和50 nm. XRD分析结果表明, MNPs与Fe₃O₄的特征衍射峰完全一致, 晶体纯度良好. UV-Vis, FTIR和荧光光谱结果表明, 亲和素已固定在MNPs表面. Avi-MNPs活性评价结果表明, 其结合生物素的活力为4.706 U/mg Avi-MNPs, 低于游离的亲和素活力(14.1 U/mg D-biotin). 该方法用于检测蛋白A含量比传统酶联免疫法省时、 省力, 且对检测仪器要求低.