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The surface of nano-scale magnetic beads is modified with specific active functional groups that can adsorb nucleic acids. By using different lysing solutions, binding solutions, and washing solutions, they can specifically bind to the target nucleic acid under specific conditions, while using magnetic beads With its own magnetism, it can easily achieve directional movement and enrichment under the action of an external magnetic field, so as to achieve the purpose of separating nucleic acid and impurities, and then achieve the separation and purification of target substances to obtain purified nucleic acid.
The magnetic bead method does not require centrifugation, is simple to operate, and is suitable for automated extraction. The entire extraction process takes a short time. At present, most nucleic acid extractors on the market use magnetic bead separation to achieve automation.
Magnetic solid phase extraction (MSPE) has many advantages over conventional SPE methods as the magnetic or magnetizable sorbent can be dispersed in sample solution thereby increases the interfacial area between sorbent and sample.
MSPE for biological samples (i.e., blood plasma, urine, etc.) is another crucial application area of magnetic nanoparticles. In a study carried out by Heidari and Limouei-Khosrowshahi , magnetic Fe3O4 nanoparticles modified with carbon were prepared for the extraction of various pharmaceutical compounds including carvedilol, losartan, and amlodipine besylate from plasma samples. In this study, the detection limit values varied in the range between 0.09 and 0.69ngmL1.
In another important work , Asgharinezhad et al. developed magnetic nanocomposite composed of Fe3O4 nanoparticles and polyaniline. The developed magnetic nanocomposite was successfully applied for the efficient extraction of benzodiazepine drugs including nitrazepam and lorazepam from human plasma and urine samples. The schematic depiction of the extraction process for the target drugs from biological samples using magnetic nanocomposites is shown in Fig. 5.6. The detection limit values for nitrazepam and lorazepam were obtained in the range from 0.5 to 1.8gL1 and 0.2 to 2.0gL1, respectively.
Feng and colleagues reported the design and preparation of Fe3O4 nanoparticle-modified multi-walled carbon nanotubes (MWCNTs) . The prepared MWCNT-based nanocomposites were successfully used for the efficient extraction of brucine (a neurotoxic alkaloid existing in the Nux-vomica tree) from human urine samples. In their study, the detection limit and quantification limit values were achieved as 6 and 21ngmL1, respectively.
In another interesting study conducted by Chen and coworkers , magnetic Fe3O4 nanoparticles having a layer composed of 1,3,5-triformylbenzene(Tb) and benzidine (Bd) ([email protected]) were prepared and successfully applied for the selective extraction of seven estrogens from human urine samples. The schematic demonstration of the preparation of magnetic [email protected] nanocomposite and extraction process for the target estrogens is shown in Fig. 5.7. In this study, the achieved detection limit values varied in the range from 0.2 to 7.7ngL1.
Mirzapour et al. reported the preparation of organic dendrimer-modified magnetic Fe3O4 nanoparticles for the efficient extraction of rosuvastatin from human urine, blood plasma, and tablet samples . For this purpose, first, magnetic Fe3O4 nanoparticles were synthesized by using FeCl2 and FeCl3. Then the surface of the synthesized magnetic Fe3O4 nanoparticles was modified with organic dendrimers containing ethylene diamine and methyl methacrylate. The prepared organic dendrimer-modified magnetic Fe3O4 nanoparticles were successfully used for the extraction of the target pharmaceutical compound rosuvastatin from human urine, blood plasma, and tablet samples. In this study, the highest extraction capacity of the prepared organic dendrimer-modified magnetic Fe3O4 nanoparticles was achieved as 61mgg1.
In another study reported by Ji and colleagues , magnetic Fe3O4 nanoparticles having MIP layer were successfully prepared and applied for the selective extraction of 9-hydroxyrisperidone and risperidone from human urine samples. The obtained detection limit values for 9-hydroxyrisperidone and risperidone were 0.24 and 0.21ngmL1, respectively.
Azodi-Deilami and coworkers prepared magnetic Fe3O4 nanoparticles coated with molecularly imprinted polymeric shell for the selective extraction of paracetamol from human plasma samples . In their study, the detection limit and quantification limit values for the target pharmaceutical compound paracetamol in human plasma were achieved as 0.17 and 0.4gL1, respectively.
In another interesting study , Jiang et al. reported the development of magnetic [email protected] nanoparticles modified with octadecyl (C18) functional groups for the efficient extraction of aromatic amines including 1-aminonaphthalene, 4-aminobiphenyl, 4,4-diaminodiphenylmethane and 4-aminophenylthioether from human urine samples. The detection limit values for 1-aminonaphthalene, 4-aminobiphenyl, 4,4-diaminodiphenylmethane, and 4-aminophenylthioether were obtained as 1.3, 0.88, 1.1, and 1.1ngmL1, respectively.
Zhang and colleagues prepared polydopamine-coated magnetic Fe3O4 nanoparticles modified with MWCNTs for the extraction of antiepileptic drugs including phenytoin, oxcarbazepine, and carbamazepine from human urine, plasma, and cerebrospinal fluid samples . In their study, the achieved detection limit values for the target drug compounds were in the range between 0.4 and 3.1ngmL1.
For magnetic solid-phase extraction (MSPE), the magnetic sorbents employed have a great influence on the separation and enrichment speed, enrichment factor, selectivity, antiinterference ability, and reproducibility of the MSPE-based methodologies. An ideal magnetic sorbent should have the following advantages: (i)strong magnetism to achieve fast magnetic separation; (ii) good dispersion, so as to improve the adsorption/desorption kinetics; (iii) large specific surface area, suitable porosity, and easy to modify, to provide abundant adsorption sites, which help to improve the adsorption capacity and extraction efficiency/recovery of target compounds; (iv) good selectivity/antiinterference capability, which helps to improve the ability of the method to tolerate complex matrixes; (v)good stability, can withstand acid and alkali environments, ultrasound, stirring, and oscillation treatment; (vi) reusable, namely, reversible adsorption; (vii) mild adsorption and desorption conditions; (viii) easy preparation and good reproducibility of preparation; (ix) low cost, and easy to obtain raw materials and low sorbents consumption during extraction; (x)environmentally friendly with low reagent consumption in its preparation and in the extraction process.
Magnetic nanoparticle (MNP) sorbents used for MSPE are nanometer-sized structures with good magnetism and available with various functional groups. MNP sorbents with superparamagnetism can be easily and rapidly separated from aqueous solution with the aid of an external magnetic field, facilitating the isolation of the target compounds adsorbed on the MNPs from the sample matrix. MNP sorbents can be functionalized with specific groups, e.g., hydroxyl, carboxyl, sulfonic acid, amino, mercapto, or chelating functional groups, resulting in favorable selectivity for various target compounds. So far, a variety of materials have been introduced, including silica-based, carbon-based, metal/metal oxides, metal-organic frameworks, porous organic polymers, mesoporous materials, and imprinted and restricted access materials.
In the last two decades, magnetic solid phase extraction (MSPE) methods using magnetic adsorbents have become one of the most commonly used methods for the separation and enrichment of organic, inorganic, and bioactive species at the matrix level. In 1973, Robinson and coworkers proposed the first magnetic separation device . afaikova and afaik used magnetic solid phase extraction as an analytical application in 1999 . They prepared a magnetic charcoal sorbent for the magnetic solid phase extraction of safranin O and crystal violet in water samples . About a 460-fold preconcentration factor was obtained for the analytes.
The MSPE method is based on the adsorption and desorption of analytes on magnetic adsorbents that are added to the sample solution containing the analytes. In this method, different type of polymers, nanomaterials, metals, and metal oxides that can be used as adsorbents are modified by magnetic particles, such as nanosized Fe3O4, -Fe2O3, ZnFe2O4, and ZnFe2O4. In this way, adsorbents that do not show magnetic properties are given magnetic properties .
Magnetic nanoparticles such as Fe3O4 and -Fe2O3 have low stability in solution medium, especially in acidic conditions, which cause the decomposition of materials in a short time and the loss of their magnetic properties. To prevent this drawback, the materials obtained are modified by silica, alumina oxides, or different groups resistant to harsh working conditions .
In this method, the magnetic sorbent is added to the sample solution containing the analyte or analytes of interest. Then, in order to adsorb the analytes on the sorbent, the sorbent with the sample solution is allowed to interact for a certain period of time. To make this interaction more effective, faster, and simpler, the resulting mixture consists of sample solution, and the magnetic sorbent is mixed for a certain time by means of devices such as vortex, magnetic stirrer, and shaker. After completion of the adsorption process, sorbent is isolated from the sample solution by an external magnetic field. Next, an eluent is added to the sorbent for elution of the target analytes, and the sorbent is isolated from the eluent phase by an external magnetic field. The concentration of target analytes in the eluent phase was analyzed by a suitable detection system. The main interactions between sorbent and analytes are dipoledipole, dipole-induced dipole, hydrogen bonding, dispersion forces, and ionic .
Selection of the suitable sorbent used MSPE is the most important step in this method, which affects the extraction efficiency as in other methods. Some of the most often used new-generation solid phase microextraction sorbents in MSPE applications are :
Environmental analysis: Extraction and preconcentration of metals, dyes, pesticides, pharmaceutical active ingredients, surfactants, PAHs, mutagenic, and carcinogenic analytes in water and sewage samples.
In 2002, afaikova and afaik tested the applicability of the magnetic solid phase extraction procedure for high volumes of urine samples. For this purpose, they have fabricated a reactive copper phthalocyanine dye-immobilized magnetite particles for the separation and preconcentration of crystal violet dye as a model analyte in high volumes of urine samples since crystal violet leads to an increased risk of cancer for living cells . In 2005, the same research group used the MSPE method to separate and preconcentrate nonionic surfactants based on aliphatic alcohols, hydrogenated fatty acid methyl esters, and oxyethylated nonylphenol in water samples .
Huang and Hu fabricated, characterized, and used -mercaptopropyltrimethoxysilane- (-MPTMS-) modified silica-coated magnetic nanoparticles (SCMNPs) as an innovative SPME sorbent for the separation and preconcentration of Pb, Hg, Cu, and Cd at trace levels in environmental and biological samples. In this method, 50mg of magnetic sorbent was added to the sample solution including metal ions (pH 6.0), and the mixture obtained was ultrasonicated for 10min to ensure the adsorption of analytes on the magnetic sorbent. Then the sorbent was isolated from the sample solution phase by applying an external magnetic field, and analytes on the sorbent were eluted with 1.0molL1 HCl and 2% (m/v) thiourea elution solution by using ultrasonication power. Analytes concentrations in the eluent phase were measured by ICP-MS. The limits of detection for analytes were between 24 and 56pgL1 . Suleiman et al. have used bismuthiol-II-immobilized silica-coated magnetic nanoparticles for the separation and preconcentration of trace amounts of Pb, Cu, and Cr in lake and river water samples. Analytes in the aqueous phase were extracted to 100mg of the magnetic nanosorbent phase at pH 7.0 by using an ultrasonic irritation source. A solution of 1.0molL1 HNO3 was used to desorp analytes from the sorbent. Concentrations of analytes were measured by ICP-OES .
A important application of the magnetic sorbentson-chip, online solid phase extractionwas reported by Li et al. in 2009. The authors fabricated a poly(dimethylsiloxane)(PDMS)/glass hybrid microchip for online solid phase extraction (SPE) and electrophoresis separation of the trace amount of fluorescence isothiocyanate (FITC)-labeled phenylalanine (Phe). The extraction phase was prepared by modifying the magnetic microspheres with hydroxyl-terminated poly-dimethylsiloxane (PDMS-OH). The extraction phase was conveniently immobilized into the solid phase extraction channel by magnetic field. In this system, injection of the sample solution into the SPE channel (PDMS-OH microspheres bed), and the desorption of analyte from the sorbent phase into the electrophoresis channel was electrically driven . Cheng et al. have used 1-hexadecyl-3-methylimidazolium bromide (C16mimBr)-coated Fe3O4 magnetic nanoparticles (NPs) as a magnetic adsorbent for mixed hemimicelles solid phase extraction of trace amounts of 2,4-dichlorophenol and 2,4,6-trichlorophenol compounds in environmental waters, followed by HPLC-UV analysis. The new nanosized sorbent provided a high surface area that led to high adsorption capacity and high extraction efficiencies (74%90%) at a minimum level of sorbent (40mg) . Jiang et al. have used zincon-immobilized silica-coated magnetic Fe3O4 nanoparticles for the magnetic solid phase extraction of trace amounts of lead in water samples prior to graphite furnace atomic absorption spectrometric (GFAAS) determination. The detection limit (LOD), enrichment factor (EF), and recovery results of the proposed method were found as 10ngL1, 200, and 84%104%, respectively . Cui et al. have prepared chitosan-modified magnetic nanoparticles by an emulsion method for the magnetic separation and preconcentration of Cr(III) and Cr(VI) in lake and tap water samples prior to ICP-OES detection. The LOD, PF, and RSD% for Cr(III) and Cr(total) were found as 100, 0.02, and 0.03ngmL1 and as 4.8% and 5.6%, respectively .
Ferrofluid-based SPME method is another important application of solid phase extraction. Ferrrofluid is a magnetic fluid that is prepared by dispersing magnetic nanoparticles in the ionic liquids or surfactants homogeneously. The fluids obtained in this way show magnetic features and can be controlled by an external magnetic field. Hence ferrofluids are used in many applications such as SPE, magnetic resonance imaging (MRI), magnetophoretic control, drug delivery, and the like. Gharehbaghi et al. have prepared an ionic liquid ferrofluid and have used it as the extraction phase for dispersive magnetic solid phase extraction of Cd(II) ions in water samples, followed by FAAS detection. In this procedure, prepared ferrofluid was added to the sample solution, and the mixture obtained was shaken to ensure the dispersion of the ferrofluid phase into the sample solution. At this stage, the hydrophobic analyte complex was adsorbed/extracted to magnetic sorbent particles. After completion of the extraction stage, magnetic particles were collected by applying an external magnetic field, and analyte was eluted for FAAS analysis. When compared with other MSPE procedures, this method is faster, and its applicability and usability are simpler for opaque/dark samples, in which observation and separation of the extraction phase are difficult .
Wang et al. have used a hydrothermal reaction procedure to synthesize a Fe3O4-functionalized metal-organic framework (m-MOF) composite as a MSPE sorbent. They have synthesized the metal-organic framework from Zn(II) and 2-aminoterephthalic acid. X-ray diffraction, FT-IR, TGA, SEM, and magnetization methods were used for the characterization of m-MOF composite. The new magnetic sorbent was used for the separation and preconcentration of trace amounts of copper, followed by ETAAS detection . Azodi-deilami has used magnetic molecularly imprinted polymer nanoparticles (m-MIPs) as a magnetic solid phase extraction sorbent for trace amounts of paracetamol in human blood plasma samples. In the synthesis of the m-MIPs, magnetite (Fe3O4) as the magnetic component, 2-(methacrylamido) ethyl methacrylate as a cross-linker and methacrylic acid as a functional monomer were used. The m-MIPs synthesized were characterized by TEM, FT-IR, XRD, and vibrating sample magnetometry methods. Analysis of paracetamol in the last phase was measured with HPLC. The LOD, LOQ, PF, and RSD% recoveries for paracetamol were 0.17gL1, 0.4gL1, 40, and 4.5%, respectively .
Pan et al. have prepared a planar-structure amine-functional magnetic polymer-modified graphene oxide nanocomposite sorbent (NH2[emailprotected]) for the magnetic solid phase extraction of trace amounts of pentachlorophenol (PCP), 2-chlorophenol (2-CP), 2,4,6-trichlorophenol (2,4,6-TCP), 2,4-dichlorophenol (2,4-DCP), and 2,3,4,6-tetrachlorophenol (2,3,4,6-TeCP). Concentrations of analytes were measured by LC-MS/MS. The MSPE method provided a very high preconcentration factor (1000) and good recovery results between 86.4% and 99.8% . In a different MSPE application, scientists used magnetic multiwalled carbon nanotube composites (MMWCNTs) for the separation and preconcentration of trace amounts of linear alkylbenzene sulfonates from environmental water samples. They have prepared MMWCNTs by using a one-pot chemical coprecipitation procedure. In this procedure, analytes in 500mL of water sample were extracted with 100mg of sorbent into 1.0mL of eluent, and analysis of trace amounts of linear alkylbenzene sulfonates in eluent was carried out by HPLC .
Magnetic polymers are another important functionalized magnetic sorbent for MSPE. Polymers are often employed to passivate the surface of the MNPs during or after the synthesis to avoid agglomeration. In general, polymers can be chemically anchored or physically adsorbed on MNPs to form a single or double layer, which creates repulsive forces to balance the magnetic and the van der Waals attractive forces acting on the MNPs. Thus, by steric repulsion, the MNPs are stabilized in suspension. Table5.3 shows some examples of the application of different polymer-based magnetic materials for the extraction of organic contaminants in foods by MSPE.
Polymers containing different functional groups can bind to the surface of MNPs. For example, poly(divinylbenzene-co-methacrylic acid)-coated Fe3O4 coreshell MNPs were prepared by Li etal.  and applied for the extraction of estrogenic endocrine-disrupting compounds in natural waters, followed by HPLC-MS/MS analysis. Recoveries of the target analytes were between 71% and 108% in spiked tap water. Another different preparation strategy, based on a simple comixing method to obtain magnetic one-dimensional polyaniline (1D-PANI) NPs, was reported by Gao etal. . First, 1D-PANIs and MNPs were prepared by chemical oxidation and solvothermal methods, respectively. Then, they were comixed and could be assembled spontaneously to achieve FexOy/1D-PANIs. The prepared nanocomposite was used to develop a simple, rapid, and effective method for the preconcentration of trace amounts of four fluoroquinolones in honey samples (recoveries 86%121%, RSD<16%) by HPLC-FLD. In other works, molecular imprinting technique has been used to build selective recognition sites in stable polymers for the preparation of selective polymer-based magnetic sorbents. In addition, to prevent interference from template leaching of MIPs in trace analysis, the use of dummy templates as substitutes for the analyte of interest has been proposed in some works. He etal.  prepared a [email protected] using methacrylic acid as functional monomer, ethylene glycol dimethacrylate as crosslinking agent, and melamine as template. The magnetic material developed was used for the selective extraction of melamine in milk. Under optimal conditions, successful recoveries in the range of 88%96% were achieved. In another work , the extremely large surface of GO and the magnetic properties of Fe3O4 NPs were combined to develop magnetic GO nanocomposites. With this purpose, GO/Fe3O4 was applied as support to develop a novel nanocomposite based on dummy-surface MIP. Propionamide was used as dummy template and the resulting material (GO/[email protected]) was applied for the rapid and selective quantification of acrylamide in heat-processed foods.
MNPs stabilized by single or double layers of polymer are not air stable, and easily leached by acidic solution, resulting in the loss of their magnetization. For these reasons, the development of other methods for protecting MNPs against deterioration has been explored. In that respect, as indicated in Section220.127.116.11, considerable attention has been paid to the preparation and application of silica-coated magnetic composites ([email protected]). Thus, to improve the performance of MNPs, novel composites with trilayer structure that combines the advantages of [email protected] and [email protected] or @DMIP have been prepared and applied in the last years for food sample preparation. In some works, molecularly imprinted techniques are based on preliminary construction of MNPs and their subsequent application as supporter for the synthesis of the [email protected]@polymer nanocomposite. For example, Gao etal.  prepared a novel [email protected][emailprotected](MAA-co-EDGMA) material that was applied for the MSPE of 11 sulfonamides in milk samples. Combined with HPLC-MS/MS the method was rapid, convenient, and efficient for the determination of the target analytes with good recoveries (88%116%). In another study, Liu etal.  developed supermagnetic surface molecularly imprinted NPs for perfloxacin mesylate (PEF-M), via surface-initiated atom transfer radical polymerization. The [email protected] [email protected] material was directly used to selectively enrich PEF-M from eggs before further HPLC-UV analysis (recoveries 93%97%. RSD<4%). Xie etal.  analyzed BPA, hexestrol, dienestrol, and diethylstilbestrol in milk by using [email protected][emailprotected] as sorbent. Diethylstilbestrol was employed as the template molecule. Recoveries of the endocrine-disrupting compounds in spiked milk samples were satisfactory (from 78% to 93%), with the exception of BPA (around 67%). On the other hand, BPA could be specifically extracted with recoveries near 100% and good reproducibility in milk samples by using [email protected][emailprotected] via RAFT. The imprinting method was based on binary functional monomers, so 4-vinylpyridine and -cyclodextrin were chosen for BPA imprinting . A strategy for producing magnetic DMIPs for simultaneous enrichment and separation of four aflatoxins was developed by Tan etal. . Vinyl functionalized superparamagnetic Fe3O4 NPs as support, 5,7-dimethoxycoumarin as dummy template, and methacrylic acid and 4-vinyl pyridine as functional monomers were used to prepare the [email protected][emailprotected] sorbent. Aflatoxins were extracted and analyzed by UHPLC-MS/MS in corn samples. The method was validated and its applicability for high throughput routine food analysis was demonstrated. Finally, in another study, Arabi etal.  developed a simple method for the preparation of magnetic DMIP NPs. First, Fe3O4 was prepared and, subsequently, [email protected][emailprotected] was constructed via solgel strategy using 3-aminopropyl trimethoxysilane as the functional monomer in the presence of TEOS as decrosslinker and urethane as dummy template. The MNPs demonstrated excellent selectivity toward the target analyte (acrylamide) in the analysis of potato chips with good recoveries (94%98%).
Different efforts have been made to design on-line MSPE methodologies. A first approach is the magnetic in-tube solid phase micro extraction which is based on the immobilization of magnetic silica nanoparticles inside a fused-silica capillary used as a loop in the injection valve of a liquid chromatography system. The sample containing the analytes flows through the capillary and the analytes are retained on the magnetic solid, the internal phase is then washed and finally the analytes are eluted with an adequate solvent previous to their analysis.
Immobilization of magnetic solids in liquid chromatography and electrophoretic methods has been proposed employing permanent magnets close to the column and the capillary, respectively. The magnetic solids are then retained on the inner wall to perform the separation the analytes. Separation efficiency is a consequence of the large surface area of the magnetic particles and the magnetic field strength.
Magnetic solids have been immobilized by different protocols in order to integrate them as extraction phase in lab-on valve systems, microfluidic chip manifolds, flow injection and sequential injection protocols. In all cases, the particles are immobilized rather than being dispersed in the sample as MSPE batch mode.
MNPs possess unique features that make them suitable for many applications including sample preparation techniques. Magnetic SPE (MSPE), one of the trends in the field of sample extraction and preconcentration, is typically performed by adding functionalized MNPs to sample matrices. Samples are, then, incubated and centrifuged or sonicated for a predetermined period of time until the target analytes are adsorbed to the MNPs. The functionalized MNPs are easily isolated from the solution by applying an external magnetic field for reuse. The analytes are, desorbed by an appropriate solvent. The ease of separating suspended magnetic nanoparticles from the samples by use of a magnet field is one of its advantages, which simplifies and accelerates the isolation process. Typically, magnetic SPE sorbents are composed of Fe3O4 as a core material to impart magnetism and support nanoparticles to extract the targeted analytes. Various NPs have been proposed as the extracting materials in MSPE (Table7.1). Super paramagnetic Fe3O4 diphenyl NPs having an average diameter of 200nm were prepared and applied in the extraction and preconcentration of unmetabolized PAHs from urine samples . A magnetic SPE sorbent made of a carbon-ferromagnetic nanocomposite designed with a hydrophobic sublayer and a hydrophilic surface was applied for the preconcentration of PAHs from environmental samples for GC-MS analysis . The dual functionality feature imparts the sorbent the ability to efficiently extract PAHs with a benign compatibility with the sample matrix. Hexane was used as desorption solvent under sonication without the need for sample stirring or shaking. The obtained enrichment factors were 35- to 133-fold for the targeted analytes. Coreshell structured carbon-encapsulated magnetic NPs (CMNPs) were used to preconcentrate bisphenol A, 4-n-nonylphenol, 4-tert-octylphenol, diethyl phthalate, dipropyl phthalate, dibutyl phthalate, dicyclohexyl phthalate, dioctyl phthalate, sulfonamide, tetracyclines, and quinolones antibiotics organic compounds from water samples with subsequent quantitation using HPLC with fluorescence detector (HPLC-FLD) and HPLC with photodiode array detector (HPLC-PDA) . It is proposed that interaction between the sorbent and the analytes took place via stacking interaction, hydrophobic interaction, and hydrogen bonding, and the extraction ability of CMNPs is controlled by the content of oxygen-containing species and graphitized carbon on the carbon shell . It was found that tuning the temperature of sorbent preparation controls the sorbent content of oxygen-containing species and graphitization, which in turn governs the degree of sorbent hydrophobicity. CMNPs prepared at 850C were highly graphitized (80%) and had a strong adsorption affinity to highly polar and moderately polar analytes. Reasonable extraction of nonpolar and moderately nonpolar compounds was achieved with CMNPs prepared at 300500C with the graphitization efficiency of carbon shell lower than 50%.
The prepared CMNPs sorbent had ample oxygen-containing species (80%) on its surface at 200C heating temperature and favored the extraction of quinolones antibiotics over other analytes. A magnetic carbon nanomaterial for Fe3O4 enclosure hydroxylated multiwalled carbon nanotubes (Fe3O4-EC-MWCNTs-OH) was used for magnetic SPE of aconitines (aconitine, hypaconitine, and mesaconitine) from human serum samples followed by their quantitation by HPLC with diode array detector (HPLC-DAD) . Under the optimized experimental conditions, the recoveries of spiked serum samples were between 98.0% and 103.0%. Fig.7.3 depicts the experimental process of the Fe3O4-EC-MWCNTs-OH SPE method.
Modifications from conventional SPE were developed to lower consumption of toxic reagents, decrease sample preparation time, and, most importantly, lower the detection limits of analytes. The following methods describe the new advances in SPE for CE. Magnetic SPE (MSPE) was proposed by afakov and Safark22 where magnetized sorbents were dispersed in the sample solution. The agglomeration and redispersion of the adsorbent were driven by the application or removal of an external electric field. The MSPE method was used for the preconcentration of illegal drugs in human urine,23,24 glyphosate in guava fruit,24 benzimidazole drugs in swine tissue,25 genomic deoxyribonucleic acid (DNA) from genetically modified soybeans,26 and quinolones and tetracyclines in milk samples14,27 prior to CE analysis.
Molecularly imprinted polymers (MIPs) are synthetic materials with recognition sites or cavities specially designed for a specific analyte. The use of MIP as chromatographic material in SPE was first demonstrated by Sellergren28 for the molecular recognition of pentamidine in human urine samples. Template molecules such as digoxin,29 trichlorfon,30 and ochratoxin A31 were then used for sample preparation prior to CE analysis in various sample matrices. SPE cartridges packed with MIPs were also employed for the extraction of heavy polycyclic aromatic hydrocarbons from edible oils.32,33
Stir-bar sorptive extraction was pioneered by Baltussen and coworkers34 for the analysis of volatile and semivolatile pollutants from aqueous samples in GCMS. A stir bar that is coated with an adsorbent layer is introduced into the sample solution. The analytes are extracted from the sample during stirring and are released by heat or solvents before analysis. An excellent application was done by Li and coworkers35 where a zirconia-coated stir bar was used for the extraction of chemical warfare degradation products alkyl alkylphosphonic acids and methylphosphonic acids in environmental waters. The high affinity of nanometer-sized zirconia molecules with the phosphonate group caused the effective extraction. The analytes were released by ultrasonication in NH4OH and methanol before CE analysis.
Simultaneous extraction and derivatization in the SPE cartridge were also developed with immobilized chromophores on the sorbent.36 The target amines were extracted by covalently bonding with the chromophore.
The book contains the valuable source of information about the wide spectrum of application fields of molecularly imprinted polymers as a green sorption medium employed in various types of analytical procedures. It describes the application potential of currently molecular imprinting technologies, associated with the SPE, magnetic imprinted microspheres, sorbents in mass spectrometry or imprinted polymer electrochemical sensors, in the field of determining the specific organic and inorganic pollutants in environmental, biological and food samples. Moreover, this book puts into consideration the aspect related to the employment of green chemistry features in MIPs preparation process. This book is mainly dedicated for the young scientist who would like to start their adventure with application of MIPs in everyday analytical practice and increase their knowledge about recent advances in molecular imprinting technologies. Moreover, this scientific project might be the valuable source of information for more experienced researchers which would like to enrich their knowledge with green analytical chemistry features and surface imprinted micro- and nanoparticles as well as magnetic microspheres. In summary, newly designed and appropriate prepared MIPs devoted for the selective recognition of an individual analyte or a specific group of chemical compounds in environmental, biological and food samples might be considered as reasonable way out for analytical chemistry, especially in a case where the preparation of proper sorption mediums with acceptable morphological properties and physicochemical characteristics contain a significant challenge.
At this point it should be highlighted that the preparation process of a new type of MIPs gives a possibility to introduce to the laboratory practice the computer molecular modelling. This solution might give a possibility to design a new type of sorption material without the need for a series of syntheses and, in a consequence, the consumption of a significant amount of solvents and reagents. This example of interdisciplinary studies (physical chemistry, polymer chemistry, analytical chemistry, and environmental chemistry) gives an additional point to considered the MIP sorption materials as a green analytical solution in the field of sample preparation techniques. Combining the knowledge from the different fields of chemistry it is possible to design and synthetize polymer sorption mediums which might contain a powerful solution for selective isolation and/or preconcentration of chemical compounds characterized by the very specific properties, in environmental, biological and food samples. In addition, the application of correctly prepared and well characterized MIP sorption material at the stage of sample preparation or separation of analytes might have a significant impact on a selectivity and the sensitivity of entire analytical procedure causing a decrease in the numerical values of method detection (MDL) and method quantification limits (MQL).
A surfactant-coated dodecyl-functionalized magnetic nanoparticle (MNP) was prepared for the SPE of steroid hormones from environmental and biological samples.5 The MNPs were synthesized by coprecipitation and were functionalized with dodecyltriethoxysilane, which resulted in dodecyl-grafted MNPs (C12Fe3O4). They were further modified with nonionic surfactants. Tween surfactants coating the C12Fe3O4 were able to be used as magnetic SPE sorbents. The developed method was successfully used with HPLCUV for the detection of steroid hormone compounds in environmental and urine samples.
Further, a restricted access magnetic core-mesoporous shell microsphere with C8-modified interior pore-walls ([email protected]) was developed and used for the detection of diazepam in rat plasma by LCMS.24
Recently, Hu et al. prepared restricted accessed [email protected]@4-(2-pyridylazo)resorcinol MNPs for direct magnetic SPE of trace metal ions in human fluids, followed by inductively coupled plasma mass spectrometry (ICPMS).25
This is modified SPE in which capturing of target molecule is carried out by dispersing the solid sorbents in the complex solution containing the analytes followed by centrifugal separation of the adsorbent containing the captured analytes and their subsequent elution . MNPs are most commonly used in d-SPE technique which is advantageous to conventional SPE in terms of faster capturing, large volume capacity, no blockage, and devoid of high back pressure issue. Magnetite (Fe3O4) and maghemite (-Fe2O3) are the most frequently used MNPs in d-SPE which can also be termed magnetic solid-phase extraction (MSPE). These MNPs can either be applied alone as observed in the separation of nerve agents from water sample or in conjugation with other NMs, such as silica-coated MNP which has been applied for polycyclic aromatic hydrocarbons (PAHs) extraction from water samples [95,207209]. Extraction of various sulfonamide antibiotics from water samples has also been achieved using this method . Higher adsorption capacity of the sorbent material is ideal for such technique like graphene-based MNPs have been used for identification of carbamate pesticides residue in tomatoes . MNP layered over coreshell made by polydopamine , poly(divinylbenzene-cometacrylic acid) , and palmitate  has also yielded better result in estrogen extraction from water. Similarly, another group of MNPs, such as CTAB-MNPs  and hexadecyldimethyl amine-MNPs  which are primarily surfactant NPs, has been used for isolation of anionic materials like perfluorinated compounds. Another example of such MNPs is europium and terbium-coated NPs that have been used in MSPE for extraction of tetracyclines and quinolones from various samples. Further advantage of this technique is its compatibility with downstream analysis through chromatography or mass spectrometry (MS)-based platforms 
Various studies have been conducted in high-gradient magnetic separation (HGMS) technology over the past few years. The major advantages of this technology are obvious when processing highly complex feedstocks, such as blood. The current state-of-the-art technology for protein fractionation is liquid chromatography, which needs several time-consuming and costly upstream purification steps to be able to process the feedstock. In contrast, HGMS allows the extraction of one protein fraction directly from the non-purified complex feedstock. With HGMS technology, the efforts of downstream processing can be decreased drastically, while the yield is increased.
The principle of the technology is to bind a specific protein fraction to magnetic beads with a highly selective functionalized surface. By using a magnetic field, it is possible to extract the magnetic beads and also the specific fraction from a non-purified feedstock together with these beads in a single unit operation. Application fields for this technology are slurries involving high downstream effort, extremely low titer, and highly valuable components like hormones, antibodies, enzymes, or simply the functionalized particles themselves.
Research was conducted on the magnetic beads, the selective binding, and the elution process. The first rotor-stator magnetic separator systems were built in order to evaluate the separation principle. The separation, particle life time as well as elution processes were evaluated and proven
To take the HGMS to the next stage, a new design concept was developed for the rotor-stator technology, a quantum leap in terms of yield and harvesting efficiency as well as cleaning and sterilization. The ANDRITZ high-gradient magnetic separator design is compliant to:
The valorization of bauxite residue generated by the Bayer process is a major challenge for the alumina industry, for both economical and environmental reasons. Iron oxides, mainly goethite and hematite, are major constituents of bauxite residue that also have a potential economic value. Many attempts have been reported in the literature, of reducing these species to magnetic iron oxides, such as magnetite, followed by its recovery using magnetic separation. No commercial success has however been reported to date. The initial steps of our detailed study, undertaken on the magnetic separation process to extract the iron compounds from calcined bauxite residue, have shown that a basic magnetic separator cannot recover the magnetite content. A homemade lab-scale disc magnetic separator has been fabricated and the influence of key parameters, such as solids concentration, feed flow rate, rotational speed, etc., was quantified. However, regardless of the equipment performance, the results are highly dependent on the material submitted to magnetic separation. In order to properly evaluate the various parameters influence, and the efficiency of the separation process, an ideal mixture of magnetic material (magnetite) and non-magnetic material (as is bauxite residue) was prepared and submitted to the disc magnetic separator. The collected material is enriched to more than 90 wt% magnetic content, from an initial 30 wt%.
The authors would like to thank Mr. Eric Lemay and Mr. Pascal Vandal for their assistance in the laboratory. Thanks are also due to the Government of Canada and the Government of Qubec for the granted MITACS scholarship. Finally, thanks to Rio Tinto for financial support and for providing the calcined bauxite residue used in the experiments.
Superparamagnetic particles have been used extensively in diagnostics and other research applications for the isolation of cells and biomolecules such as antibodies, nucleic acids, and polypeptides. The biomagnetic separators available from Bangs accommodate a complete range of magnetic separation applications, including cell sorting, mRNA and DNA isolation, and the purification of proteins and protein complexes.Our biomagnetic separators feature high-energy neodymium-iron-boron [Nd-Fe-B] magnets that ensure fast separation rates, excellent wall or well retention, and maximum yield. They are suitable for use with each of our magnetic particle lines. See our Biomagnetic Separators brochure.
The 1.5 mL Magnetic Separator is a durable plastic magnetic separation unit designed to accommodate a standard 1.5 mL microcentrifuge tube for a complete range of magnetic separation applications, including cell sorting, mRNA and DNA isolation, and the purification of biomolecules. By placing the tube into the separator, our high energy neodymium-iron-boron [Nd-Fe-B] magnet will quickly draw superparamagnetic particles out of the solution. Once separation is complete, with particles firmly held to the wall, the supernatant may be removed. The 1.5 mL Magnetic Separator is simple and easy to use with most established bead protocols. See magnetizing force here.
The BioMag MultiSep Magnetic Separator accommodates three different tube sizes, which makes it a convenient and economical solution for processing magnetic microsphere samples. The MultiSep can be used with 50mL, 15mL, or 1.5mL centrifuge tubes for individual separations in a variety of applications.
The BioMag Multi-6 Microcentrifuge Tube Separator is a durable plastic magnetic separation unit designed to accommodate six 1.5mL microcentrifuge tubes. The magnets and tube holders are positioned to efficiently magnetically separate microliter volumes common in molecular biology applications.
The BioMag 96-Well Plate Separator (bottom pull) is a durable magnetic separation unit designed to accommodate most 96-well plates. The magnetic separator consists of permanent magnets enclosed in a plastic frame.
The BioMag 96-Well Plate Side Pull Separator is a durable magnetic separation unit designed to accommodate 96-well plates that allow the magnetic pins to fit between the individual wells. This separator allows particles to be pulled to the side of the walls, giving access to the bottom of the wells for more complete fluid removal and less chance of particle aspiration. The magnetic separator consists of 24 permanent neodymium-iron-boron rod magnets embedded in a plastic frame.
The BioMag Flask Separator is a 12.5cm x 6cm rectangular magnetic separation unit designed for use with tissue culture flasks. Test tubes may be used with this unit by securing tubes against the magnet with a rubber band (or by using one magnet on either side of the flask, as shown in the image to the left). The unit consists of permanent magnets enclosed in a plastic frame. The magnetic strength contained in each disc magnet is 27 megagauss Oersteads. Hurry! The BioMag Flask Separator is being offered at a discounted price, so stock up today!
One of the greatest challenges facing the mining industry is the separation of unwanted material generated by the extraction process from the valuable material. Mining, whether done through open seam or underground means, creates a huge amount of waste product in the form of worthless or low value minerals and unusable man-made materials. These materials can be extremely difficult to separate from the valuable materials miners are after. Perhaps the most efficient way of separating these materials is through magnetic separation.
Magnetic separation machines consist of a vibratory feeding mechanism, an upper and lower belt and a magnet. The bulk material is fed through the vibrating mechanism onto the lower belt. At this point, the magnet pulls any material susceptible to magnetic attraction onto the upper belt, effectively separating the unwanted metals from the rest of the bulk.
Magnetic separation has been used in the mining industry for more than 100 years, beginning with John Wetherills Wetherill Magnetic Separator, which was used in England in the late nineteenth century.
Magnetic separation is most commonly used in the mining industry to separate tramp ore, or unwanted waste metals, from the rest of the bulk material. Tramp ore typically consists of the man-made byproducts created by the mining process itself, such as wires from explosive charges, nuts and bolts, nails, broken pieces from hand tools such as jack hammers and drills or tips off of heavy duty extraction buckets.
Magnetic separation machines are usually placed at the beginning of a mines materials processing line to remove tramp ore before it can cause harm to downstream equipment such as ore crushers and conveyor belts, which can be easily damaged by metal shards or other sharp objects.
The type of magnetic separator used by a mine depends on what material they are extracting and how much tramp ore is generated by their process. As a result, separators of different magnetic flux, or power, can be used. There are 2 types of magnetic separators; electromagnetic and permanent.
Electromagnetic separators generate a magnetic field by switching power from alternating current to direct current. Electromagnetic separators are useful for removing large pieces of tramp ore from the bulk material. These separators are typically suspended over a conveyor belt and draw the unwanted material upward. Electromagnetic separators are easy to clean as removing the tramp ore that they separate from the bulk is as simple as turning off the power that creates their magnetic field.
Permanent magnets consist of materials that generate their own magnetic field. Though not as powerful as electromagnetic separators, permanent magnets are better at attracting strongly magnetized materials such as nickel, cobalt, iron and some rare earth metals. Some permanent magnets are now being made with rare earth metals that have the ability to attract even stainless steel, which is typically not susceptible to magnetic pull. In order to clean permanent magnets, a stainless steel scraper must be used to remove any metal parts from the magnets surface.
Magnetic separation definitely is one of the most important parts of this process. I think magnetic separators are often taken for granted when it comes to processing, whether that processing is in mining or in food processing. Many people dont even know the work that goes into making food safe or mining materials pure.
Bunting is committed to helping customers across the mineral processing industry to break ground as our equipment assists them in solving challenging problems. We dig down to find the root cause of their problems and use specialty software to custom-design the best solutions to fit the customers needs. We offer exceptionally rugged magnets and heavy-duty magnetic separation equipment specifically to cater to customers in the aggregate, mining, and mineral industries. Working with these materials is tough, but Bunting equipment is even tougher. Our magnetic separation equipment will protect the other equipment utilized in your facility and allow you to deliver the highest purity product to your customers.
Bunting provides equipment to help mineral processing companies by using magnetic separation technology to remove metal contaminants from product lines and providing metal detection equipment to alert operators of ferrous and non-ferrous metal hazards trapped within greater product mass. All of our equipment is designed to be low-maintenance and operator friendly to increase efficiency and decrease downtime in your production.
Magnetic separation utilizes powerful magnets to pull out ferrous metal contaminants from a product line, trapping them against the face of the magnet and ensuring they cannot re-enter the product stream. The strength of the magnetic field means that continued product flow, no matter how rapid, will not be able to dislodge the ferrous material from where it has been initially trapped. Bunting offers magnetic separation equipment suitable for heavy-duty applications, with rugged construction designed for handling large amounts of material.
Metal detection systems generate an electromagnetic field that material is flowed through, sensing and rejecting any ferrous or non-ferrous metal that is hidden within the product. In situations where a magnetic separator may be unable to remove a non-ferrous contaminant, such as aluminum, a metal detector is able to sense this metal and remove the contaminated section of product from the greater flow of operations.
All of our products are custom designed according to the customers specifications, allowing for them to integrate seamlessly into the existing production environment. Our team of engineers works with each customer to deliver a personalized piece of equipment with the physical dimensions to best fit your space and the magnetic components that best suit your separation and detection needs.
Induced roll magnetic separators are used for the continuous extraction of small magnetic particles from certain minerals to produce mineral purification for a wide range of mineral and ceramic processing industries.
Induced roll magnetic separators are used for the continuous extraction of small magnetic particles from certain minerals to produce mineral purification for a wide range of mineral and ceramic processing industries.
The Magnetic Drum Separator is normally installed at product discharge points and incorporates a 150 180 degree magnet system, encased in a stainless steel shell, or manganese wear plates for severe application.
The Magnetic Drum Separator is normally installed at product discharge points and incorporates a 150 180 degree magnet system, encased in a stainless steel shell, or manganese wear plates for severe application.
The separation of ferrous metal happens thanks to the principle of magnetic attraction: the magnetic field generated by permanent or electrical magnets attracts the ferromagnetic intrusions, extracting them from the waste flow.
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Magnetic bead separation makes nucleic acid and protein purification more effective and easier to automate. With advancements in lab technology and reduced costs, magnetic separation has become a leading technique for sample purification. Read on below to know more about magnetic bead separators.
A magnetic bead separator uses superparamagnetic beads or particles. They are made up of tiny particles of iron oxides ranging from 20 nm to a few m with superparamagnetic properties, meaning that they exhibit magnetic properties only in the presence of an external magnetic field. Their small size enables them to stay separated when needed, along with any material theyre bound to.
The magnetic bead separator then applies an external magnetic field to attract the beads to the tubes outer edge containing the sample. This separates the beads from the suspension and immobilizes them with the target DNA.
The sample is then washed to remove the unwanted suspension, and an elution buffer is added to the tube. This elution buffer releases the DNA from the magnetic beads and is removed from the tube or microplate well by pipetting. This produces a purified DNA sample, ready for downstream assays or quantitation. Applications that benefit from using a magnetic bead separator include immunoprecipitation, cell isolation, nucleic acid isolation, and other downstream processes from PCR to ELISA.
Modern magnetic bead separator stations come in compact sizes that can be integrated with other lab instruments and robots to create a semi or fully automated system. Below are some components that you can integrate with your magnetic bead separator when designing an automated extraction system.
Magnetic bead separation is a quick and effective method of extracting DNA and other molecules. By integrating your magnetic bead separator with other instruments in an automated workcell, you can further increase your lab efficiency.
Nucleic acid separation is an increasingly important tool for molecular biology. Before modern technologies could be used, nucleic acid separation had been a time- and work-consuming process based on several extraction and centrifugation steps, often limited by small yields and low purities of the separation products, and not suited for automation and up-scaling. During the last few years, specifically functionalised magnetic particles were developed. Together with an appropriate buffer system, they allow for the quick and efficient purification directly after their extraction from crude cell extracts. Centrifugation steps were avoided. In addition, the new approach provided for an easy automation of the entire process and the isolation of nucleic acids from larger sample volumes. This review describes traditional methods and methods based on magnetic particles for nucleic acid purification. The synthesis of a variety of magnetic particles is presented in more detail. Various suppliers of magnetic particles for nucleic acid separation as well as suppliers offering particle-based kits for a variety of different sample materials are listed. Furthermore, commercially available manual magnetic separators and automated systems for magnetic particle handling and liquid handling are mentioned.
Magnetic separation is an emerging technology that uses magnetism for the efficient separation of micrometre-sized para- and ferromagnetic particles from chemical or biological suspensions. Enrichment of low-grade iron ore, removal of ferromagnetic impurities from large volumes of boiler water in both conventional and nuclear power plants, or the removal of weakly magnetic coloured impurities from kaolin clay are typical examples of magnetic separation in traditional industries. The application of these techniques in biosciences had been restricted and of limited use up to the 1970s. The idea of using magnetic separation techniques to purify biologically active compounds (nucleic acids, proteins, etc.), cells, and cell organelles led to a regrowing interest over the last decade. New magnetic particles with improved properties were developed for the partly complicated separation processes in these fields [see reviews: Olsvik et al. 1994; Safarik and Safarikova 1999; Franzreb et al. 2006].
Magnetic separation of nucleic acids has several advantages compared to other techniques used for the same purpose. Nucleic acids can be isolated directly from crude sample materials such as blood, tissue homogenates, cultivation media, water, etc. The particles are used in batch processes where there are hardly any restrictions with respect to the sample volumes. Due to the possibility of adjusting the magnetic properties of the solid materials, they can be removed relatively easily and selectively even from viscous sample suspensions. In fact, magnetic separation is the only feasible method for the recovery of small particles (diameter approx. 0.051m) in the presence of biological debris and other fouling material of similar size. Furthermore, the efficiency of magnetic separation is especially suited for large-scale purifications (Safarik et al. 2001; Franzreb et al. 2006).
These upcoming separation techniques also serve as a basis of various automated low- to high-throughput procedures that allow to save time and money. Centrifugation steps can be avoided and the risk of cross-contamination when using traditional methods is no longer encountered. Various types of magnetic particles are commercially available for nucleic acid purification, magnetic separators working in the manual and automated mode are offered. A short description of traditional and magnetic separation methods for nucleic acid isolation, together with a short overview of batch and automated separators, will be given below.
The isolation of DNA or RNA is an important step before many biochemical and diagnostic processes. Many downstream applications such as detection, cloning, sequencing, amplification, hybridisation, cDNA synthesis, etc. cannot be carried out with the crude sample material. The presence of large amounts of cellular or other contaminating materials, e.g. proteins or carbohydrates, in such complex mixtures often impedes many of the subsequent reactions and techniques. In addition, DNA may contaminate RNA preparations and vice versa. Thus, methods for the efficient, reliable and reproducible isolation of nucleic acids from complex mixtures are needed for many methods that are used today and rely on the identification of DNA or RNA, e.g. diagnosis of microbial infections, forensic science, tissue and blood typing, detection of genetic variations, etc.
A range of methods are known for the isolation of nucleic acids in the fluid phase, but they are generally based on complex series of precipitation and washing steps and are time-consuming and laborious to perform. Thus, classical methods for the isolation of nucleic acids from complex starting materials such as blood or tissues, involve the lysis of the biological material by a detergent or chaotropic substance, possibly in the presence of protein-degrading enzymes, followed by several processing steps applying organic solvents such as phenol and/or chloroform or ethanol, which in general are highly toxic and require special and, hence, expensive disposal. For example, the complete removal of proteins from nucleic acids can be achieved by the addition of sodium perchlorate (Wilcockson 1973). The separation of RNA from DNA requires selective precipitation steps with LiCl or a specific nuclease-free isolation with guanidinium hydrochloride or guanidinium thiocyanate, combined with phenol extraction and ethanol precipitation (Bowtell 1987). Such methods are not only cumbersome and time-consuming, but the relatively large number of steps required increases the risk of degradation, sample loss or cross-contamination of samples especially when several samples are processed simultaneously. In the case of RNA isolation, the risk of DNA contamination is comparatively high.
Apart from laborious and time-consuming traditional methods, alternative separation techniques have been developed. Sorption processes based on (a) hydrogen-binding interaction with an underivatised hydrophilic matrix, typically silica, under chaotropic conditions, (b) ionic exchange under aqueous conditions by means of an anion exchanger, (c) affinity and (d) size exclusion mechanisms were used for DNA purification. Solid-phase systems which adsorb DNAsilica-based particles (Vogelstein and Gillespie 1979; Boom et al. 1990, 1999; Melzak et al. 1996; Tian et al. 2000; Breadmore et al. 2003), glass fibres, and anion-exchange carriers (Ferreira et al. 2000; Endres et al. 2003; Teeters et al. 2003)are used in chromatographic separation columns [e.g. DE 41 43 639 C2 (Qiagen GmbH)] for example.
These carriers are applied for DNA isolation or purification together with highly concentrated chaotropic salt solutions (e.g. sodium iodide, sodium perchlorate, guanidinium thiocyanate). In US 5,075,430 (BioRad), for instance, usage of diatomaceous earth as a carrier material is described. Again, bonding takes place in the presence of a chaotropic salt. Other approaches are based on detergence together with a nucleic-acid-binding material (EP 0 796 327 B1, Dynal) or on the usage of a solid carrier with DNA-binding functional groups combined with polyethylene glycol and salts at high concentrations (WO/1999/058664, Whitehead Institute for Biomedical Research).
The increasing use of magnetic solid carriers in biochemical and molecular biology processes has many advantages compared to other non-magnetic separation processes. The term magnetic means that the support obtains a magnetic moment when placed in a magnetic field. Thus, it can be displaced. In other words, particles having a magnetic moment may be removed readily by the application of a magnetic field, e.g. by using a permanent magnet. This is a quick, simple and efficient way to separate the particles after the nucleic binding or elution step (see Fig.1) and a far less rigorous method than traditional techniques, such as centrifugation, that generate shear forces which may lead to the degradation of the nucleic acids. It is also possible to isolate components of the cell lysate, which inhibit for example the DNA polymerase of a following PCR reaction like polysaccharides, phenolic compounds or humic substances (Demeke and Adams 1992; Watson and Blackwell 2000).
Usually, it is sufficient to apply a magnet to the side of the vessel containing the sample mixture for aggregating the particles near the wall of the vessel and pouring away the remainder of the sample (see Fig.1).
Magnetic carriers with immobilised affinity ligands or prepared from a biopolymer exhibiting affinity to the target nucleic acid are used for the isolation process. Many magnetic carriers are commercially available and can also be prepared in the laboratory. Such materials are magnetic particles produced from different synthetic polymers, biopolymers, porous glass, or magnetic particles based on inorganic magnetic materials such as surface-modified iron oxide. Especially suited are superparamagnetic particles, which do not interact among each other in the absence of a magnetic field. These particles will magnetise under a strong magnetic field, but retain no permanent magnetism once the field is removed. When magnetic aggregation and clumping of the particles are prevented during the reaction, easy suspension of the particles and uniform nucleic acid extraction are ensured.
The diameter of the particles is approximately between 0.5 and 10m. Materials with a large surface area are preferred for binding the nucleic acids. Without going into theoretical details, the nucleic-acid-binding process may be assisted by the nucleic acid wrapping around the support. Such supports generally have an irregular surface and may be porous for example. Particulate materials, e.g. beads and in particular polymer beads, are generally preferred due to their larger binding capacity. Conveniently, a particulate solid support used will comprise spherical beads.
In the laboratory, colloidal magnetite Fe3O4 (or similar magnetic material such as maghemite Fe2O3 or ferrites) particles usually are surface-modified by silanisation. Naked iron oxide (Fe3O4) has the capacity of adsorbing DNA (Davies et al. 1998), but aggregates due to attractive forces reduce the surface area that can be used for adsorption. Silane compounds coupled to magnetite derivatised with carboxyl groups are known to have a DNA extraction ability in solutions containing PEG (Hawkins et al. 1994). Modified bacterial magnetite particles in the presence of amino silane compounds and hyperbranched polyamidoamine dendrimer are used for DNA extraction by Yoza et al. (2002, 2003). Modified magnetic cobalt ferrite particles have been investigated for DNA isolation under high sodium chloride and PEG concentrations by Prodelalova et al. (2004).
Surface modification of magnetic nanoparticles with alkoxysilanes (Bruce et al. 2004; Tan et al. 2004; Bruce and Sen 2005) or polyethyleneimine (Chiang et al. 2005; Veyret et al. 2005) is also useful. The above-mentioned magnetic colloids are not easy to separate using classical magnets. This is due to a small particle size, at which Brownian motion forces are higher than the exerted magnetic force. To enhance phase separation, various magnetic latexes that may interact with nucleic acids were prepared.
Magnetic micro-beads can be prepared in a number of ways, but usually magnetically susceptible particles (e.g. iron oxide) are coated with synthetic or biological polymers. Elaissari et al. (2003) describe the interaction of nucleic acids and different polymers. Biopolymers such as agarose, chitosan, -carrageenan, and alginate, can be prepared easily in a magnetic form (Levison et al. 1998; Prodelalova et al. 2004). In the simplest case, the biopolymer solution is mixed with magnetic particles and, after bulk gel formation, the magnetic gel formed is broken into fine particles. Alternatively, the biopolymer solution containing dispersed magnetite is dropped into a mixed hardening solution or a water-in-oil suspension technique is used to prepare spherical particles. Basically, the same process can be used to prepare magnetic particles for nucleic separation from synthetic polymers such as hydrophobic polystyrene (Ugelstad et al. 1992) and hydrophilic polyacryl amide (Elaissari et al. 2001) or poly(vinyl alcohol) (Oster et al. 2001). Genomic DNA was also successfully isolated from cell lysate on weak acid derivatives of magnetic P(HEMA-co-EDMA) and P(HEMA-co-GMA) microparticles in the presence of PEG and sodium chloride (Horak et al. 2005).
The first approach to synthesising micro-sized particles was published by Ugelstad et al. They developed an interesting methodology leading to monosized polystyrene magnetic microspheres, which were studied in various biomedical applications (Ugelstad et al. 1993). These particles have an excellent size distribution and spherical shape, but their surface is very hydrophobic and results in a high amount of unspecific protein binding on the particle surface.
Another possibility consists in combining different polymer matrix materials with silica components (Grttner et al. 2001; Mller-Schulte et al. 2005) that specifically interact with the nucleic acids.
Depending on the support and the nature of the subsequent processing required, it may or may not be desirable to release the nucleic acid from the support. The direct use of magnetic beads, e.g. in PCR or other amplifications, without eluting the nucleic acid from the surface is not trivial. The enzymatic detection and amplification methods will be inhibited by the magnetic beads, their stabilisers, or their metal oxides (Spanova et al. 2004), which decrease PCR sensitivity or lead to false negative PCR results. For many DNA detections or identification methods, elution is not necessary. Although the DNA may be randomly in contact with the bead surface and bound at a number of points by hydrogen binding or ionic or other forces, there generally will be sufficient lengths of DNA available for hybridisation to oligonucleotides and for amplification. If desired, however, elution of the nucleic acid may be achieved using known methods, e.g. higher ionic strength, heating or pH changes.
Commercially available magnetic particles that are suited for nucleic acid separation can be obtained from a variety of companies. Mostly, the matrixes are based on silica, porous glass, cellulose, agarose, polystyrene and silane (see Tables1 and 2). Moreover, some important patents exist that describe the synthesis of magnetic carriers not only for nucleic separation:
One of the first patents for particle synthesis is the Ugelstad polymerization process, which is described, for example, in EP 0 003 905 B2, US 5,459,378, and US 4,530,956 (SINTEF). It leads to monodisperse magnetic particles by several swelling and polymerisation steps. WO/1992/016581 (Cornell Research Foundation) also describes the preparation of monodisperse particles, particularly macroporous polymer beads. The process proposed uses a three-phase emulsion containing soluble polymer particles, a monomer phase and water. Nucleic acid separation using magnetic beads is described in (Alderton et al. 1992) and in WO/1991/012079 as well as in US 5,523,231 (Amersham). These magnetic beads are able to absorb the nucleic acid after a salt-ethanol precipitation. The approaches are not nucleic-acid-specific, i.e. the magnetic beads adsorb other bio-substances in parallel. Of course, this is a drawback of these approaches.
In the declaration WO/1996/041811 (Boehringer; Roche) mainly non-porous glass particles comprising mica and magnetite particles are described (Bartl et al. 1998). During their production, magnetic particles and a surrounding glass coating are superimposed on a mica core. The disadvantage of these products is their affinity to sedimentation. Furthermore, the production process is time-consuming and based on a complex spray process. Another approach to the production of particles from spherical magnetite kernels with a surface coating of silicon dioxide is covered by the European patent application EP 1 468 430 A1.
Monodisperse magnetic beads are described in WO/1998/012717 (Merck). They consist of a SiO2 core, which is given magnetic properties by a ferric-oxide coating. After a subsequent silanisation of the ferric-oxide coating, the particles can bind nucleic acids.
Many patents concerning nucleic acid separation are from the Dynal company. They developed monodisperse polymer magnetic particles with different sizes (coefficient of variation less than 5%) (see EP 0 796 327 B1), which are sold with a polystyrene matrix under the name of Dynabeads. The small-size distribution ensures reproducible separation properties. Protocols for nucleic acid separation with these particles are described by EP 0 512 439 B1 and with oligonucleotide-linked particles for specific nucleic acid separation in US 5,512,439.
Magnetic beads based on mica or polystyrene and coated by a magnetic oxide reach a high specific density, which leads to a fast sedimentation. Thus, additional mechanical mixing is necessary. The main drawback of the coated particles consists in the fact that the metal oxides may be in direct contact with the analytical solutions despite silanisation. All state-of-the-art approaches to the production of magnetic beads are laborious; the production process time amounts to several hours. To overcome this problem, the US patents 6,204,033 and 6,514,688 (chemagen Biopolymer Technologie AG) describe spherical, magnetic polymer particles based on polyvinyl alcohol particles, which can be produced in short terms using inverse suspension polymerisation. The polymer particles contain reactive hydroxyl groups to which other molecules can be coupled. Due to their hydrophilic surface, the particles exhibit small unspecific bindings only. Together with an at least partly silanised surface (DE 100 13 955 A1 and EP 1 274 745 A1) or a germanium-containing compound (DE 101 03 652 A1), they can be used for specific nucleic acid separation.
The inverse suspension process for the separation of nano- and micro-sized silica particles is suggested in WO/2002/009125 (Fraunhofer-Gesellschaft). The main idea is the dispersion of aqueous silica-sole containing magnetic colloids, which are hardened to spherical hydrophilic gel particles by adding a suited base. These particles can be used for nucleic acid separation with high binding capacities (WO/2005/50 52 581 A3, MagnaMedics GmbH).
Both total DNA and RNA are separated by the same magnetic beads. For the purpose of removing RNA from DNA, the RNA is destroyed before the DNA separation step. Adding of an RNAse or an alkali such as NaOH is an appropriate process. Vice versa, RNA can be separated if the DNA is degraded with DNAse.
The primary method considered for plasmid purification is the separation of plasmid DNA (pDNA) from the chromosomal DNA and cellular RNA of the host bacteria. Stadler et al. (2004) show that even in the case of a high copy plasmid, pDNA represents not more than 3% of the cleared lysate and that most of the critical contaminants are negatively charged (RNA, cDNA, endotoxin) and similar in size (cDNA, endotoxins) and hydrophobicity (endotoxins). A number of methods have been developed to generate a cleared lysate, but they are not able to remove proteins and lipids. Alkaline lysis of harvested bacterial cells with a subsequent neutralisation, as originally described by Birnboim and Doly (1979), is the process of choice. Cleared lysate protocols may vary slightly from each other as regards salt concentrations, volume, pH, temperature, and process step durations (Hirt 1967; Holmes and Quigley 1981; Birnboim 1983). These techniques make use of the differences in denaturation and renaturation characteristics of covalently closed circular plasmid DNA and chromosomal DNA fragments.
Table1 shows some commercially available magnetic particles used for DNA, RNA and pDNA isolation. Many magnetic particles are available with optimised buffers and protocols for small lab scale and automated systems. There are also some companies offering particles for nucleic acid purification without any further information.
The magnetic carrier is provided with binding solutions to assist in the selective capture of nucleic acids. For example, complementary DNA or RNA sequences (Satokari et al. 2005) or DNA-binding proteins may be used as well as viral proteins binding to viral nucleic acids. In this review, a short overview of eukaryotic mRNA and viral DNA/RNA will be given.
There are several companies (see Table2) offering oligodeoxythymidine immobilised with magnetic particles, which can be used effectively for the rapid isolation of highly purified mRNA from eukaryotic cell cultures or total RNA preparations (Jacobsen et al. 2004). These procedures are based on the hybridisation of the oligonucleotide dT sequence with the stable polyadenylated 3 termini of the eukaryotic mRNA. The length of the complementary sequence differs between 20 and 30 oligonucleotides. This sequence is directly bound covalently to the particle surface or indirectly by biotinylated oligonucleotides and the interaction of streptavidin-coated particles. CPG and Dynal (now Invitrogen) offer MPG and Dynabeads with already immobilised biotinylated oligonucleotide, but also other companies offer streptavidin-modified particles, which can be used for mRNA isolation, as described, e.g. by the mRNA isolation kit with MagneSphere from Promega. Nearly all magnetic particles (except for MagaCell oligo-dT30 and Sera-Mag oligo-(dT)30) are available together with an optimised buffer system and helpful protocols.
Automated extraction of viral RNA and DNA from the plasma mini-pool is performed by the chemagic Viral DNA/RNA Kit and chemagic Magnetic Separation Module I (Hourfar et al. 2005a,b; Pichl et al. 2005).
A rapid diagnosis of enterovirus infection by magnetic bead extraction has been established by Muir et al. (1993). Enterovirus RNA can be separated from large-volume water samples using the NucliSens miniMAG System (Rutjes et al. 2005). Hei and Cai (2005) developed a system for purifying SARS coronavirus RNA by a hybridisation of a specific oligonucleotide sequence, which is immobilised on the magnetic bead surface.
A variety of magnetic separators are available on the market, ranging from very simple concentrators for one tube to complicated fully automated devices. The isolation of nucleic acids is mostly performed in the batch mode using commercially available lab-scale magnetic separators (particle concentrators). Separators are usually made of strong rare-earth permanent magnets designed to hold various amounts of micro-tubes or tubes.
Particles with a diameter larger than 1m can be separated easily using simple magnetic separators, while separation of smaller particles (magnetic colloids with a particle size ranging from ten to hundreds of nanometres) may require the use of high-gradient magnetic separators.
The racks are designed to hold various amounts of micro-tubes or tubes. Test tube magnetic separators allow to separate magnetic particles from volumes between approximately 5l and 50ml. There are many combinations with other features like a mixing function (Ademtech) or a possibility to turn the separator over for the removal of the supernatant (chemagen Biopolymer-Technologie AG). Other devices are applied for the separation of magnetic particles from the wells of standard micro-titration plates. In some of them the temperature can be pc-controlled (AGOWA), other devices may be inserted into automated separation devices.
Laboratory automation is increasingly important in molecular biology and biotechnology. Constantly increasing numbers of analyses of different sources and sample volumes have resulted in an enormous importance of flexible robots or automated systems. Automation is also required for handling a large number of samples without human errors.
Many instruments have been developed to automate PCR amplification, the sequencing reaction and the detection of nucleic acids, but automating DNA extraction by traditional methods with centrifugation and vacuum steps still is difficult. A complete separation of the solid carrier matrix by centrifugation is not possible. Supports filled with carrier materials cannot be used, as the ineluctable dead volumes of the support lead to sample material loss in case of small amounts of sample materials. Another drawback is the danger of mutual contamination of different biological samples, especially if directly neighbouring supports are emptied by the vacuum. However, the last decade shows that DNA purification using magnetic bead technology is suitable for automation systems, and several automated instruments for handling magnetic beads have been developed (Alderton et al. 1992; Wahlberg et al. 1992; Rolfs and Weber 1994; Fangan et al. 1999; Obata et al. 2001; Akutsu et al. 2004; Vuosku et al. 2004).
More and more vendors offer commercially automated devices for the handling of magnetic particles, e.g. for the purification of nucleic acid (see Table4). Most systems are offered together with system-specific optimised particles, buffer systems and protocols.
The devices are able to process between six and 96 samples in parallel and commonly customised for small buffer volumes. For larger volumes, the chemagic Magnetic Separation Module I (<10ml) (see Fig.2) or the Magtration System 8l(7ml) can be used.
chemagic Magnetic Separation Module I consisting of (A) separation head with magnetizable rods [here 12-well format for large (50ml) volumes; 96-well format for MTPs also available], (B) electro magnet, (C) chemagic dispenser for parallel filling of all required buffer solutions (accessory) and (D) tracking unit. The principle functionality regarding separation and resuspension of magnetic beads is shown in the scheme
The present review has shown that the separation of nucleic acid is a highly dynamic field of research and development. An increasing number of commercial vendors offer magnetic particles, also in the form of a kit that is optimally suited for the application desired. The increasing number of publications shows that magnetic particles of higher potential are currently under research. Materials with more specific-binding properties and a better separability are promising approaches. A higher degree of automation leads to systems analysing a larger number of samples and higher sample volumes at the same time.
Akutsu J-I, Tojo Y, Okochi M, Yohda M, Segawa O, Obata K, Tajima H (2004) Development of an integrated automation system with a magnetic bead-mediated nucleic acid purification device for genetic analysis and gene manipulation. Biotechnol Bioeng 86:667671
Boom R, Sol C, Beld M, Weel J, Goudsmit J, Wertheim-van Dillen P (1999) Improved silica-guanidinium thiocyanate DNA isolation procedure based on selective binding of bovine alpha-casein to silica particles. J Clin Microbiol 37:615619
Breadmore MC, Wolfe KA, Arcibal IG, Leung WK, Dickson D, Giordano BC, Power ME, Ferrance JP, Feldman SH, Norris PM, Landers JP (2003) Microchip-based purification of DNA from biological samples. Anal Chem 75:18801886
Fangan BM, Dahlberg OJ, Deggerdal AH, Bosnes M, Larsen F (1999) Automated system for purification of dye-terminator sequencing products eliminates up-stream purification of templates. Biotechniques 26:980983
Hourfar MK, Schmidt M, Seifried E, Roth WK (2005) Evaluation of an automated high-volume extraction method for viral nucleic acids in comparison to a manual procedure with preceding enrichment. Vox Sang 89:7176
Jacobsen N, Nielsen PS, Jeffares DC, Eriksen J, Ohlsson H, Arctander P, Kauppinen S (2004) Direct isolation of poly(A)(+) RNA from 4M guanidine thiocyanate-lysed cell extracts using locked nucleic acid-oligo(T) capture. Nucleic Acids Res 32:e64
Muir P, Nicholson F, Jhetman M, Neogi S, Banatvala JE (1993) Rapid diagnosis of enterovirus infection by magnetic bead extraction and polymerase chain-reaction detection of enterovirus RNA in clinical specimes. J Clin Microbiol 31:3138
Obata K, Segawa O, Yakabe M, Ishida Y, Kuroita T, Ikeda K, Kawakami B, Kawamura Y, Yohda M, Matsunaga T, Tajima H (2001) Development of a novel method for operating magnetic particles, Magtration Technology, and its use for automating nucleic acid purification. J Biosci Bioeng 91:500503
Rutjes SA, Italiaander R, van den Berg HHJL, Lodder WJ, de Roda Husman AM (2005) Isolation and detection of enterovirus RNA from large-volume water samples by using the nucliSens miniMAG System and real-time nucleic acid sequence-based amplification. Appl Environ Microbiol 71:37343740
Vuosku J, Jaakola L, Jokipii S, Karppinen K, Kamarainen T, Pelkonen VP, Jokela A, Sarjala T, Hohtola A, Haggman H (2004) Does extraction of DNA and RNA by magnetic fishing work for diverse plant species? Mol Biotechnol 27:209215
DE 101 03 652 A1 Magnetische Polyvinylalkoholpartikel mit modifizierter Oberflche zur Isolierung und Reinigung von Nukleinsuren (2002) Brassard L, Parker J, Smets H, Oster J; chemagen Biopolymer-Technologie AG, Germany
US 4,336,173 Process for preparing an aqueous emulsion or dispersion of a partly water-soluble material, and optionally further conversion of the prepared dispersion or emulsion to a polymer dispersion when the partly water-soluble material is a polymerizable monomer (1980) Ugelstad J; SINTEF, Norway
US 4,530,956 Process for the preparation of aqueous dispersions of organic materials and possible further conversion to a polymer dispersion when the organic material is a polymerizable monomer (1985) Ugelstad J, Berge A; SINTEF Norway
WO/2002/009125 Spherical, magnetic SiO2 particles with an adjustable particle and pore size and an adjustable magnetic content. Method for producing them and use of SiO2 particles of this type (2001) Mller-Schulte D, Fischer R; Fraunhofer-Gesellschaft zur Frderung der angewandten Forschung e.V. Germany
Multotec supplies a complete range of magnetic separation equipment for separating ferromagnetic and paramagnetic particles from dry solids or slurries, or for removing tramp metal. Multotec Dry and Wet Drum Separators, WHIMS, Demagnetising Coils and Overbelt Magnets are used in mineral processing plants across the world. We can engineer customised magnetic separation solutions for your process, helping you improve the efficiency of downstream processing and lower your overall costs of production.
Multotec provides a wide range of magnetic separators including: Permanent magnet Low Intensity Magnetic Separators (LIMS) or Medium Intensity Magnetic Separators (MIMS) and electromagnetic High Intensity Magnetic Separators (HIMS). Multotec provides unmatched global metallurgical expertise through a worldwide network of branches, which support your processing operation with turnkey magnetic separation solutions, from plant audits and field service to strategic spares for your magnetic separation equipment.
Whether you need to recover fast moving tramp metal, recover valuable metals in waste streams or enhance the beneficiation of ferrous metals, Multotec has the magnetic separator you require. Dry drum cobber magnetic separators provide an initial upgrade of feed material as well as a gangue material rejection stage. By improving the material fed to downstream plant processes, our magnetic separation solutions reduce the mechanical requirements of grinding, ultimately lowering overall costs. Our heavy media drum separators are ideally suited for dense media separation plants. Our ferromagnetic wet drum separators can be used in iron ore separation plants in both rougher or cleaner beneficiation applications. We also provide demagnetising solutions that reverse the residual effects that magnetic separation has on the magnetic viscosity of ferrous slurries, to return the mineral stream to an acceptable viscosity for downstream processing. These demagnetising coils generate a magnetic field that alters magnetic orientation at 200 Hz.
The trend towards larger and faster travelling conveyors in the African mining industry has highlighted the vital role of overbelt magnets. Solutions need to be optimised to such factors as belt speed and width, the belt troughing angle, the burden depth, the material density and bulk density, the expected tramp metal specifications, ambient operating temperatures and suspension height to provide maximum plant and cost efficiency. Multotec can supply complete overbelt magnet systems, from equipment supply to a turnkey service by means of its strategic partners, including even the gantry work.
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The MagneSphere Technology Magnetic Separation Stands can be used in conjunction with any of the PolyATtract Systems and are ideal for applications requiring multiple paramagnetic isolations of biomolecules. These stands use the same strong rare earth magnet used in the PolyATtract Systems Magnetic Separation Stands and come in a variety of sizes to accommodate 296 samples.
TheEpiMag HT (96-Well) Magnetic Separatoris a magnetic stand (magnetic rack) that allows paramagnetic bead precipitation of liquid samples from various flat-bottom or U-bottom 96-well microplates (we recommend our EpiMag 96-Well Microplates for ideal compatibility). Designed from repurposed Greiner microplates, it ensures a high degree of compatibility with microplates while remaining extremely cost friendly to accomodate any lab's budget. It can be used for isolation and purification of nucleic acids and proteins, immunoprecipitation, immunoassays (ELISA), cell sorting, and purification of biomolecules. This magnetic separation device has the following features and advantages:
Product DescriptionThe implementation of twenty-four extremely powerful magnetic rods made ofneodymium-iron boronenables fast and easy magnetic separation. Each magnetic rod addresses four wells of a 96-well plate simultaneously. With this magnetic separator, paramagnetic beads will be firmly pulled to the side of the plate wells, which ensures complete removal of the magnetic beads from the solution by a robot or with a multichannel pipette, minimizing sample loss.Get in Touch with Mechanic