多孔磁性硅胶微球的制备及其在基因组脱氧核糖核酸提取中的应用

采用模板法制备了多孔磁性硅胶微球,用于生物样品中基因组脱氧核糖核酸(DNA)的分离纯化。以球形和无定型硅胶为对照,考察了吸附液组成和洗脱时间等实验参数对小牛胸腺基因组DNA在磁性硅胶固相载体上的提取回收率的影响。实验结果表明:20%(W/V)聚乙二醇和2 mol/L氯化钠,洗脱10 m in,DNA的回收率可达80%;采用简单的细胞裂解体系和合适的吸附液组成,磁性微球应用于酿酒酵母中基因组DNA的提取,得到了平均长度约为5 kb、A260/A280大于1.77的高纯度DNA片段。     

表面电性可控的磁性微球的制备、表征及其在基因组DNA萃取中的应用

运用水热法、St?ber法制备了具有核壳型结构的超顺磁性Fe3 O4@SiO2微球,运用氨丙基三乙氧基硅烷( APTES)对Fe3 O4@SiO2微球表面进行修饰,得到表面电性可控的氨基化Fe3 O4@SiO2磁性微球,运用透射、扫描电镜表征微球形貌,采用Zeta电位研究微球表面的荷电特性。将氨基化Fe3 O4@SiO2微球用于人类全血中基因组脱氧核糖核酸( DNA)的萃取,开发了固相萃取方法,并探究微球与DNA的作用机理,对提取产物进行凝胶电泳和PCR实验。结果表明,自制的微球成功地从全血中提取出纯度较高的基因组DNA,提取率约70%,微球对基因组DNA的饱和吸附量约为40 ng/μg,提取液可直接用于进一步的生物分析。     

原子转移自由基聚合法制备表面聚合物接枝四氧化三铁纳米粒子

为制备表面具有柔性高分子链的磁性微球,采用化学共沉淀法制备了具有超顺磁性的Fe3O4纳米微球,用KH550对Fe3O4纳米微球进行化学改性得到表面氨基化的Fe3O4纳米微球,与2-溴代异丁酰溴反应后制得含有引发官能团的Fe3O4纳米微球,随后将含溴的Fe3O4纳米微球与小分子单体与之通过原子转移自由基聚合(ATRP)法共聚。测试结果表明聚合物链成功地接枝到了Fe3O4纳米微球表面。     

改性果胶磁性微球的制备及吸附性能研究

为了提高果胶磁性微球的分散性及成球性,采用油酸对磁性微球进行表面改性后,制备改性果胶磁性微球。通过红外光谱、扫描电镜、XRD和磁性分析等对样品进行表征。油酸和果胶在纳米四氧化三铁表面形成了良好的修饰层,且油酸的加入提高了果胶磁性微球的分散性及成球性。研究了改性果胶磁性微球用量和溶液pH值等对其吸附Cu(Ⅱ)性能的影响,考察了其吸附动力学和吸附等温线。25℃时,改性果胶磁性微球对Cu(Ⅱ)的吸附达平衡需要2 h,饱合吸附容量为52.36 mg/g,Freundlich模型和Langmuir模型可以较好的拟合实验结果,最大吸附量为119.05 mg·g~(-1)。     

磁性壳聚糖硅胶复合微球的制备及其吸附Cu~(2+)的性能研究

以壳聚糖与正硅酸四乙酯为原料,采用溶胶-凝胶法,用戊二醛辅助交联合成了磁性壳聚糖硅胶复合微球。通过红外光谱、扫描电镜、X-射线衍射等方法对磁性壳聚糖硅胶复合微球的形态和组成特性进行分析,制备的磁性复合微球中壳聚糖与硅胶材料复合均匀,材料粒径均一,机械强度较高。考察了制备的磁性壳聚糖硅胶复合微球对Cu~(2+)的吸附性能,结果表明微球对Cu~(2+)具有较好的吸附性能,吸附容量达到98.7mg/g。     

磁性Fe_3O_4微粒表面有机改性

在分散聚合法制备复合磁性微球过程中 ,采用硅烷偶联剂 KH- 570对磁性 Fe_3O_4微粒进行表面改性 .红外光谱 (FTIR)、光电子能谱 (XPS)分析结果表明 ,偶联剂与磁性微粒表面以化学键形式结合 .改性后 ,Fe_3O_4微粒与单体及其聚合物之间具有良好的亲和性 ,采用改性后的磁性微粒可以显著改善磁性微球的性能指标 .     

不同表面官能团和化学组成共聚物微球的表面生物矿化研究

采用具有不同共聚物组成和端基官能团的聚己内酯-b-聚乙二醇共聚物(PCL-b-PEG),通过双乳液溶剂挥发法制备了一系列具有不同表面性质的生物降解高分子微球.采用生物模拟矿化的方法以磷灰石修饰微球表面.进一步通过扫描电镜、热重分析仪、X-射线衍射仪和光电子能谱仪对微球表面磷灰石的形貌、含量、结构和组成进行了分析.研究了微球表面亲水性、粗糙度、官能团以及矿化时间对于磷灰石形成的影响.最终实验结果表明,随着共聚物中PEG含量增加,微球表面粗糙度和亲水性增加,因此微球表面磷灰石含量增加.同时微球表面官能团以及矿化时间的不同也会对磷灰石的形成和分布产生明显影响.     

磁性Fe3O4微粒表面有机改性

在分散聚合法制备复合磁性微球过程中,采用硅烷偶联剂KH 570对磁性Fe3O4微粒进行表面改性.红外光谱(FTIR)、光电子能谱(XPS)分析结果表明,偶联剂与磁性微粒表面以化学键形式结合.改性后,Fe3O4微粒与单体及其聚合物之间具有良好的亲和性,采用改性后的磁性微粒可以显著改善磁性微球的性能指标.     

DNA改性聚砜多孔微球选择性清除环境激素

将DNA视为生物大分子材料,制成DNA改性聚砜微球,在生物体外考察改性微球对环境激素,包括重金属离子的选择性清除.制备了2种改性聚砜微球:DNA共混微球(将DNA共混入聚砜溶液,当混合溶液滴入水中时,发生相分离而沉淀)和DNA表面改性聚砜微球(先制备聚砜多孔微球,再通过紫外线照射的方法将DNA接枝到聚砜微球表面).两种DNA改性聚砜微球在水中比较稳定,在酸或碱性条件下,由于DNA水解而不稳定.对环境激素的清除结果表明,DNA改性聚砜微球可以选择性清除包括联苯、苯酚和二恶荚类环境荷尔蒙物质;同时选择性吸附锌、铜、银、铅和铬等重金属离子.     

用于生物探测的PAMAM树形高分子增强免疫磁性微球

采用悬浮聚合的方法制备羧基修饰的聚苯乙烯磁性微球(CMM)，研究了CMM和CMM-PAMAM(第5代)两者与牛血清蛋白(BSA)的结合状况。结果显示；CMM与BSA的结合量为22.2μg／mg。PAMAM偶联在磁性微球表面后，提高了磁性微球表面官能团的密度，因而CMM-PAMAM(第5代)与BSA的结合量可达54．4μg／mg。     

Preparation of magnetite-loaded silica microspheres for solid-phase extraction of genomic DNA from soy-based foodstuffs

Solid-phase extraction has been widely employed for the preparation of DNA templates for polymerase chain reaction (PCR)-based analytical methods. Among the variety of adsorbents studied, magnetically responsive silica particles are particularly attractive due to their potential to simplify, expedite, and automate the extraction process. Here we report a facile method for the preparation of such magnetic particles, which entails impregnation of porous silica microspheres with iron salts, followed by calcination and reduction treatments. The samples were characterized using powder X-ray diffractometry (XRD), scanning electron microscopy (SEM), nitrogen adsorption/desorption isotherms, and vibrating sample magnetometry (VSM). XRD data show that magnetite nanocrystals of about 27.2 nm are produced within the pore channels of the silica support after reduction. SEM images show that the as-synthesized particles exhibit spherical shape and uniform particle size of about 3 μm as determined by the silica support. Nitrogen sorption data confirm that the magnetite-loaded silica particles possess typical mesopore structure with BET surface area of about 183 m²/g. VSM data show that the particles display paramagnetic behavior with saturation magnetization of 11.37 emu/g. The magnetic silica microspheres coated with silica shells were tested as adsorbents for rapid extraction of genomic DNA from soybean-derived products. The purified DNA templates were amplified by PCR for screening of genetically modified organisms (GMOs). The preliminary results confirm that the DNA extraction protocols using magnetite-loaded silica microspheres are capable of producing DNA templates which are inhibitor-free and ready for downstream analysis.     

Cross-coupling reactions of nucleoside triphosphates followed by polymerase incorporation. Construction and applications of base-functionalized nucleic acids

Construction of functionalized nucleic acids (DNA or RNA) via polymerase incorporation of modified nucleoside triphosphates is reviewed and selected applications of the modified nucleic acids are highlighted. The classical multistep approach for the synthesis of modified NTPs by triphosphorylation of modified nucleosides is compared to the novel approach consisting of direct aqueous cross-coupling reactions of unprotected halogenated nucleoside triphosphates. The combination of cross-coupling of NTPs with polymerase incorporation gives an efficient and straightforward two-step synthesis of modified nucleic acids. Primer extension using biotinylated templates followed by separation using streptavidine-coated magnetic beads and DNA duplex denaturation is used for preparation of modified single stranded oligonucleotides. Examples of using this approach for electrochemical DNA labelling and bioanalytical applications are given.     

应用于全氟辛烷磺酸萃取富集的全氟癸基修饰毛细管硅胶整体柱的制备及应用

利用溶胶-凝胶法,经过烷氧基硅烷的水解、硅羟基的缩聚、凝胶化、陈化、中孔制备、干燥和表面修饰等步骤制备了全氟癸基修饰的毛细管硅胶整体柱。采用该整体柱对全氟辛烷磺酸(PFOS)进行萃取富集,考察其富集特性和效率,并与传统的C18毛细管硅胶整体柱进行对比。结果表明,全氟癸基修饰毛细管硅胶整体柱(15 cm×75μm)对PFOS具有更高的吸附量和更好的富集选择性,其平均吸附量可以达到75 ng;样品中PFOS的质量浓度为0.25 mg/L时,富集倍数平均可以达到29倍。此全氟癸基修饰毛细管硅胶整体柱对PFOS具有良好的萃取富集性能,可用于水质中痕量PFOS的萃取富集。     

Preparation of phenyl‐functionalized magnetic mesoporous silica microspheres for the fast separation and selective enrichment of phenyl‐containing peptides

Peptide enrichment before mass spectrometry analysis is essential for large‐scale peptidomic studies, but challenges still remain. Herein, magnetic mesoporous silica microspheres with phenyl group modified interior pore walls were prepared by a facile sol–gel coating strategy, and were successfully applied for selective enrichment of phenyl‐containing peptides in complex biological samples. The newly prepared nanomaterials possessed abundant silanol groups in the exterior surface and numerous phenyl groups in the interior pore walls, as well as a large surface area (592.6 m²/g), large pore volume (0.33 cm³/g), uniform mesopores (3.8 nm), strong magnetic response (29.3 emu/g), and good dispersibility in aqueous solution. As a result of the unique structural properties and size‐exclusion effect, the core–shell phenyl‐functionalized magnetic mesoporous silica microspheres exhibited excellent performance in fast separation and selective enrichment of phenyl‐containing peptides, and the adsorption capacity for bradykinin reached 22.55 mg/g. In addition, selective enrichment of phenyl‐containing peptides from complex samples that are consist of peptides, large proteins, and human serum were achieved by using the as‐prepared microspheres, followed by high‐performance liquid chromatography with ultraviolet detection and electrospray ionization quadrupole time‐of‐flight mass spectrometry analysis. These results demonstrated the as‐prepared microspheres would be a potential candidate for endogenous phenyl‐containing peptides enrichment and biomarkers discovery in peptidome analysis.     

应用于全氟辛烷磺酸萃取富集的全氟癸基修饰毛细管硅胶整体柱的制备及应用

利用溶胶-凝胶法,经过烷氧基硅烷的水解、硅羟基的缩聚、凝胶化、陈化、中孔制备、干燥和表面修饰等步骤制备了全氟癸基修饰的毛细管硅胶整体柱。采用该整体柱对全氟辛烷磺酸(PFOS)进行萃取富集,考察其富集特性和效率,并与传统的C18毛细管硅胶整体柱进行对比。结果表明,全氟癸基修饰毛细管硅胶整体柱(15 cm×75 μm)对PFOS具有更高的吸附量和更好的富集选择性,其平均吸附量可以达到75 ng;样品中PFOS的质量浓度为0.25 mg/L时,富集倍数平均可以达到29倍。此全氟癸基修饰毛细管硅胶整体柱对PFOS具有良好的萃取富集性能,可用于水质中痕量PFOS的萃取富集。     

Quantitative 1H NMR analysis of reacted silanol groups in silica nanoparticles chemically modified with monochlorosilanes

Quantitative analysis of reacted silanol groups in silica nanoparticles modified chemically with monochlorosilanes was performed by ¹H NMR after treatment with cesium fluoride. Silica nanoparticles were modified chemically by the reaction between the silanol groups and monochlorosilanes, and the structure of the organic moiety anchored onto the silica surface was confirmed with solid‐state ¹³C NMR. As monochlorosilanes react with silanol groups at 1:1 ratio unlike di‐ or trichlorosilanes, the number of the silanes introduced into silica nanoparticles equals that of reacted silanol groups. Organically modified silica nanoparticles were dissolved using cesium fluoride, and the amount of the soluble organic compounds originated from the introduced silanes was determined by a ¹H NMR internal standard method using pyrene as the reference. Those values determined by ¹H NMR were in good agreement with those determined by elemental analysis. Thus, the number of reacted silanol groups per one particle was calculated on the basis of the results obtained by the ¹H NMR method, and the values were highly dependent on the steric structure of the introduced silanes. Copyright © 2010 John Wiley & Sons, Ltd.     

Magnetic Latex Particles in Nanobiotechnologies for Biomedical Diagnostic Applications: State of the Art

Summary: The main objective of this short state-of-the art is to report on the preparation of reactive magnetic latexes for biomedical applications. The main advantage of colloidal magnetic particles is their separation upon applying an external permanent magnetic field. Then magnetic particles bearing reactive groups or specific receptors are used in biomedical diagnosis such as immunoassay, molecular diagnosis for specific capture and detection of nucleic acids, viruses extraction and detection, cell sorting and more recently in numerous biotechnological applications. The preparation of magnetic latexes requires the use of well-defined iron oxide nanoparticles in appropriate medium. In this direction, aqueous ferrofluids were largely used to prepare functional magnetic latexes, whereas few approaches have been devoted to the use of organic ferrofluids.     

Adsorption of organic molecules on silica surface

The adsorption behaviour of various organic adsorbates on silica surface is reviewed. Most of the structural information on silica is obtained from IR spectral data and from the characteristics of water present at the silica surface. Silica surface is generally embedded with hydroxy groups and ethereal linkages, and hence considered to have a negative charged surface prone to adsorption of electron deficient species. Adsorption isotherms of the adsorbates delineate the nature of binding of the adsorbate with silica. Aromatic compounds are found to involve the pi-cloud in hydrogen bonding with silanol OH group during adsorption. Cationic and nonionic surfactants adsorb on silica surface involving hydrogen bonding. Sometimes, a polar part of the surfactants also contributes to the adsorption process. Styryl pyridinium dyes are found to anchor on silica surface in flat-on position. On modification of the silica by treating with alkali, the adsorption behaviour of cationic surfactant or polyethylene glycol changes due to change in the characteristics of silica or modified silica surface. In case of PEG-modified silica, adsolubilization of the adsorbate is observed. By using a modified adsorption equation, hemimicellization is proposed for these dyes. Adsorptions of some natural macromolecules like proteins and nucleic acids are investigated to study the hydrophobic and hydrophilic binding sites of silica. Artificial macromolecules like synthetic polymers are found to be adsorbed on silica surface due to the interaction of the multifunctional groups of the polymers with silanols. Preferential adsorption of polar adsorbates is observed in case of adsorbate mixtures. When surfactant mixtures are considered to study competitive adsorption on silica surface, critical micelle concentration of individual surfactant also contributes to the adsorption isotherm. The structural study of adsorbed surface and the thermodynamics of adsorption are given some importance in this review.