- Email: [email protected]
Mechanochemical synthesis of magnetically responsive materials from non-magnetic precursors
Materials Letters 126 (2014) 202–206
Contents lists available at ScienceDirect
Materials Letters journal homepage: www.elsevier.com/locate/matlet
Mechanochemical synthesis of magnetically responsive materials from non-magnetic precursors Ivo Safarik a,b,n, Katerina Horska a, Kristyna Pospiskova b, Jan Filip b, Mirka Safarikova a a b
Department of Nanobiotechnology, Institute of Nanobiology and Structural Biology of GCRC, Na Sadkach 7, 370 05 Ceske Budejovice, Czech Republic Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 11, 783 71 Olomouc, Czech Republic
art ic l e i nf o
a b s t r a c t
Article history: Received 30 October 2013 Accepted 7 April 2014 Available online 15 April 2014
Mechanochemical synthesis of various types of magnetically responsive materials from non-magnetic powdered precursors has been developed. The preparation is based on the mechanochemical conversion of ferrous and ferric ions at the presence of alkaline hydroxide into magnetic iron oxides nanoparticles (maghemite identified by XRD measurements). The presence of powdered nonmagnetic materials during the mechanochemical process led to the efficient deposition of magnetic nanoparticles on the surface of the treated materials in the form of individual nanoparticles and their aggregates. The prepared magnetically responsive materials have been used as adsorbents for xenobiotics removal and as a carrier for enzymes immobilization. & 2014 Elsevier B.V. All rights reserved.
Keywords: Mechanochemistry Magnetic materials Magnetic adsorbents Magnetic carriers
1. Introduction Magnetic nano- and microparticles have attracted an increasing interest in various fields including nanoscience, nanotechnology, biosciences, biotechnology and environmental technology. Many chemical procedures have been used to synthesize magnetic particles, such as classical co-precipitation, reactions in constrained environments (e.g. microemulsions), sol–gel syntheses, sonochemical reactions, hydrothermal reactions, hydrolysis and thermolysis of precursors, flow injection syntheses, electrospray synthesis and microwave synthesis [1,2]. Recently mechanochemical procedures have been used to synthesize magnetic iron oxides and ferrites nanoparticles [3–5]. Mechanochemistry represents one of several ways of chemical activation. In solid-state mechanochemistry, nonthermal chemical reactions occur because of the deformation and fracture of solids, which are technically induced by milling or grinding of the materials. During this process the mechanical energy induces chemical reactions and phase transformations [6]. Up to now, mechanochemical synthesis has been employed mainly for the production of individual iron oxides and ferrites. However, mechanochemistry can be successfully used also for the preparation of magnetically responsive materials from originally
n Corresponding author at: Department of Nanobiotechnology, Institute of Nanobiology and Structural Biology of GCRC, Na Sadkach 7, 370 05 Ceske Budejovice, Czech Republic. Tel.: +420 387775608; fax: +420 385310249. E-mail address: [email protected] (I. Safarik). URL: http://www.nh.cas.cz/people/safarik (I. Safarik).
http://dx.doi.org/10.1016/j.matlet.2014.04.045 0167-577X/& 2014 Elsevier B.V. All rights reserved.
nonmagnetic powdered precursors. In this paper we present a very simple, generally applicable procedure for the preparation of magnetic materials from variety of inorganic, organic and biological precursors, together with the illustration of their possible applications as adsorbents and enzyme carriers. Mechanochemical postmagnetization can be very useful for smart magnetic modification of diverse non-magnetic materials.
2. Materials and methods Materials: FeCl3. 6H2O, FeCl2. 4H2O, montmorillonite, halloysite, Candida rugosa lipase (EC 3.1.1.3), 1,4-butanediol diglycidyl ether (BDDE), sodium (meta)periodate, 1,10 -carbonyldiimidazole (CDI), 4-nitrophenyl butyrate, dimethyl sulfoxide, Bismarck brown Y and sodium acetate were purchased from Sigma-Aldrich, USA. Microcrystalline cellulose, safranin O and 4-nitrophenol were from Lachema, Czech Republic, while the common chemicals were from Lach-Ner, Czech Republic. Finally powdered biological materials with diameters below 1 mm (spruce sawdust, scales from grass carp (Ctenopharyngodon idella), wheat straw, pistachio nut shells, peanut husks, oak acorns, spent coffee grounds), as well as potato starch and pine pollen were obtained locally. The natural ocherous sediment containing biogenic iron oxides was collected using glass vessels from a water stream in Ceske Budejovice (Czech Republic); it was sieved through a 1 mm sieve to remove larger detrital fraction, then repeatedly washed with deionized water and air dried at a temperature not exceeding 50 1C.
I. Safarik et al. / Materials Letters 126 (2014) 202–206
Mechanochemical synthesis of magnetically responsive materials: In the standard procedure a mixture of 1.35 g of FeCl3 6H2O (0.005 mol), 0.50 g of FeCl2 4H2O (0.0025 mol) and 4 g of sodium chloride (inert material used to avoid particles agglomeration) was grounded in a mortar at room temperature for 10 min. Then, appropriate amount of target nonmagnetic powdered material (usually 1–2 g) was added and after thorough mixing the process continued for another 10 min. As the last step, powdered potassium hydroxide (1.22 g) was added and after mixing the grinding continued for 10 min. After KOH addition the mixture became brown. During the grinding process the material was scraped from the mortar wall occasionally. After finishing the mechanochemical process, the magnetically modified material was thoroughly washed with water (to remove soluble impurities and free iron oxides particles) and stored in a suspension, or it was air dried. Structural characterization of iron oxide nanoparticles prepared by mechanochemical procedure: X-ray powder diffraction (XRD) patterns of selected samples were recorded on PANalytical X'Pert PRO instrument in Bragg–Brentano geometry with Fe-filtered CuKα radiation (40 kV, 30 mA). The samples were inserted into conventional front-loading cavity sample holder and scanned in the 2θ range of 10–901 (step size 0.0171). The commercial standards SRM640 (Si) and SRM660 (LaB6) from NIST were used for the evaluation of the line positions and instrumental line broadening, respectively. The acquired patterns were evaluated using the X'Pert HighScore Plus software (PANalytical) together with PDF-4þ database. Adsorption of organic dyes on magnetically responsive composites: 30 mg of selected magnetically modified adsorbents were mixed with 7.0 mL of water in a test tube. Then, 0.1–3 mL of stock water solution of a tested dye (1 mg/mL) was added and the total volume of the solution was made up to 10.0 mL with water. The suspension was mixed on a rotary mixer (Dynal, Norway) for 3 h at room temperature. The magnetic adsorbent was then separated from the suspension using a magnetic separator (MPC-1 or MPC-6, Dynal, Norway) and the clear supernatant was used for the spectrophotometric measurement. The concentration of free (unbound) dye in the supernatant (Ceq) was determined from the calibration curve. The amount of dye bound to the unit mass of the adsorbent (qeq) was calculated using the following formula: qeq ¼ ðC tot C eq Þ=3
ðmg=gÞ
ð1Þ
where Ctot is the total (initial) concentration of dye (μg/mL) used in the experiment. The value qeq was expressed in mg of adsorbed dye per 1 g of adsorbent. Equilibrium adsorption data were fit to Langmuir adsorption isotherms using SigmaPlot software. Immobilization of lipase on magnetic cellulose particles: C. rugosa lipase was immobilized on magnetically modified cellulose particles. Three various agents were used for activation of hydroxyl groups present in the cellulose structure, namely sodium periodate, butanediol diglycidyl ether and carbonyldiimidazole. To prepare samples, 30 mg of prepared magnetic cellulose particles were washed with water and magnetically separated by NdFeB magnet. In the next step, particles were treated by the activating agent. Using 1,10 -carbonyldiimidazole (CDI), 1.5 mL of 0.5% (w/v) solution in dimethyl sulfoxide was added to particles. For the periodate method, 1.5 mL of 1% (w/v) solution of sodium (meta)periodate (NaIO4) in 0.1 M sodium acetate buffer pH 4 was utilized. During activation by the epoxide method, 1.5 mL of 3% (v/v) solution of 1,4-butanediol diglycidyl ether (BDDE) in 0.25 M NaOH was used. Particles were shaken with these activating agents on an automatic rotator (20 rpm) for 24 h at room temperature in the dark. After the modification procedure, particles were magnetically separated, supernatants were poured off and particles were repeatedly washed with distilled water. Subsequently, 1.5 mL of lipase solution (1 mg/mL) in 50 mM potassium phosphate buffer,
203
pH 7.5, was added to the activated particles and shaken on automatic rotator (20 rpm) for 24 h at 4 1C. After the immobilization procedure, particles were magnetically separated, supernatant containing unbound enzyme was removed and particles with bound enzyme were repeatedly washed with buffer until no enzyme activity in the supernatant was detected. Magnetic cellulose particles with immobilized lipase were stored in buffer at 4 1C. Lipase assay: Activity of lipase immobilized on magnetic cellulose particles was determined spectrophotometrically using 0.5 mM 4-nitrophenyl butyrate (dissolved in ethanol) in 50 mM potassium phosphate buffer, pH 7.5. Particles of magnetic cellulose with attached lipase were stirred during the reaction in buffer containing the substrate, then magnetically separated to the bottom of the cuvette to stop the reaction and increasing amount of yellow-colored 4-nitrophenol was measured at 405 nm. Molar absorption (extinction) coefficient (ε) of the reaction product was determined spectrophotometrically for the specific medium conditions used in this enzyme assay. Coefficient for 4-nitrophenol in 50 mM potassium phosphate buffer, pH 7.5 at 405 nm was 13815 L/mol cm. Operational stability of immobilized lipase: Reusability of lipase immobilized on magnetic cellulose particles was tested as its operational stability; it was repeatedly used for 7 reaction cycles. Particles with attached lipase were washed with buffer between each cycle. Activity of lipase was measured spectrophotometrically as described previously. Residual activities of lipase after each cycle were determined and compared taking the initial activity in the first cycle as 100%. Time stability of immobilized lipase: Particles of magnetic cellulose with immobilized lipase were stored in the reaction buffer at 4 1C for 30 days and percentage of residual enzyme activity on the carrier was determined. Possible presence of lipase released from the support was tested during this time period by measuring the activity of free lipase in the supernatant.
3. Results and discussion Mechanochemical synthesis of magnetic composite materials: In mechanochemical synthesis, hydrated solid reactants, namely ferrous and ferric chlorides reacted with KOH during grinding in a mortar to form magnetite nanoparticles. To avoid agglomeration, the excess of sodium chloride was added to the precursors before grinding. The following reaction takes place [7]: 2FeCl3 6H2O(s) þ FeCl2 4H2O(s) þ 8KOH(s)-8KCl(s) þFe3O4(s) þ20H2O(g)
(2)
Magnetite nanoparticles as the primary product are stable only in inert atmosphere [5]. However, in the described process which
Fig. 1. Appearance of original powdered oak acorns suspension (left), suspension of oak acorns powder after magnetic modification (middle) and demonstration of magnetic separation of magnetically modified oak acorns powder (right).
204
I. Safarik et al. / Materials Letters 126 (2014) 202–206
is performed in air, the reaction-generated heat and air oxidation lead to the conversion of magnetite nanoparticles into maghemite as follows [7]: 4 Fe3O4 (s) þO2 (g)-6 γ-Fe2O3 (s)
(3)
The addition of appropriate nonmagnetic powder material to the reaction mixture and grinding before the KOH addition led to
the homogeneous distribution of ferrous and ferric ions within the material matrix. Subsequent addition of KOH followed by grinding led to the formation of magnetite and maghemite nanoparticles bound to the surface and pores of matrix according to Eqs. (2) and (3). The possible influence of the modified material and its chemical composition may lead to the formation of the non-stoichiometric maghemite during the grinding process. Anyway magnetic iron
Fig. 2. Scanning electron microscopy of native and magnetically modified materials. 1 – native potato starch (reproduced with permission from [13]); 2 – magnetic potato starch; 3 – native pine pollen (reproduced with permission from [14]); 4 – magnetic pine pollen; 5 – native pistachio nut shells (reproduced with permission from [15]); 6 – magnetic pistachio nut shells; 7 – native halloysite; 8 – magnetic halloysite.
I. Safarik et al. / Materials Letters 126 (2014) 202–206
oxides nanoparticles bound to the treated material lead to the formation of stable magnetically responsive composite materials, which can be easily separated using a simple magnet or magnetic separator (Fig. 1). The magnetic iron oxide particles deposition is usually very stable, and during the subsequent washing steps only a very low amount of particles were released and washed out. The presence of magnetic iron oxides nanoparticles and their aggregates on the surface of selected magnetically modified materials (magnetic potato starch, pine pollen, pistachio nut shells and halloysite) can be clearly seen using scanning electron microscopy (Fig. 2). It can be expected that the strong binding of magnetic iron oxides particles to the surface of non-magnetic materials has been achieved by a subtle balance of van der Waals, electrostatic and hydrophobic interactions both between the magnetic particles and the treated material surface and between the adsorbed magnetic particles [8]. The structure of iron oxide particles in selected magnetically modified materials (namely spruce sawdust) was identified using XRD as maghemite γ-Fe2O3 (Fig. 3) with observed remnants of soluble salts. The very broad diffraction lines imply nanocrystalline character of the maghemite particles – the size of X-ray coherent domains (i.e., corresponding approximately to mean particle size) calculated according to the Scherrer formula equals to 4 nm [9]. Adsorption of organic dyes on magnetic composites: Recently, large number of low cost, easily available materials have been tested and used as adsorbents for xenobiotics removal [10]. Magnetic modification of such adsorbents simplifies their separation from the treated solutions and suspensions. That is why adsorption properties of several materials magnetized with described mechanochemical procedure (namely powdered magnetically responsive montmorillonite, grass carp scales, oak acorns and biogenic iron oxides) were tested with two water-soluble organic dyes, belonging to different dye classes, namely Bismarck brown Y (diazo dye) and safranin O (safranin dye). It was shown in preliminary experiments that the adsorption of the tested dyes reached equilibrium in approximately 60–120 min. Incubation time of 3 h was used for adsorption experiments. The equilibrium adsorption isotherms for both tested dyes and four magnetic adsorbents (prepared by addition of 1 g of material during the standard mechanochemical procedure) are shown in Fig. 4. The experimental data were analyzed by means of non-linear regression calculation using SigmaPlot software and it was shown that they follow the Langmuir isotherm equation. The Langmuir model is valid for monolayer adsorption onto a surface with a finite number of identical sites. The well-known expression
205
for the Langmuir model is given by qeq ¼
Q max bC eq 1 þbC eq
ð4Þ
where qeq (expressed in mg/g) is the amount of the adsorbed dye per unit mass of magnetic adsorbent and Ceq (expressed in mg/L) is the unadsorbed dye concentration in solution at equilibrium. Qmax (expressed in mg/g) is the maximum amount of the dye per unit mass of adsorbent to form a complete monolayer on the surface bound at high dye concentration and b is a constant related to the affinity of the binding sites (expressed in L/mg) [11]. Because the adsorption of the tested dyes can be successfully described by the Langmuir isotherm, it is possible to calculate the maximum adsorption capacity which is a very important parameter describing the adsorption process (Table 1). In the case of two dyes and four adsorbents tested, the highest value was found for Bismarck brown Y and grass carp scales (43.3 mg/g), while the lowest value was obtained for safranin O and magnetic oak acorns (12.9 mg/g). The maximum adsorption capacities of the developed materials are not so high as observed in the case of some other alternative magnetic biosorbents [12]. The possible reason for lower adsorption is the high coverage of the magnetically modified material surface with magnetic iron oxides (see Fig. 2). In order to test this possibility, two batches of magnetic montmorillonite were prepared, differing by the amount of native montmorillonite added during the standard mechanochemical procedure (one or two grams); higher amount of native material during the mechanochemical process results in lower surface coverage with magnetic iron oxides. The maximum adsorption capacities were 38.6 and 52.4 mg/g for one gram and two gram batches, respectively. Immobilization of lipase on magnetic cellulose particles: Selected magnetically modified materials can be used as carriers for immobilization of enzymes and other important biologically active compounds. C. rugosa lipase was immobilized on magnetic cellulose particles prepared by mechanochemical method. Three reagents, namely sodium periodate, butanediol diglycidyl ether and carbonyldiimidazole were used for activation of hydroxyl groups present in cellulose structure. Table 2 shows the activity of immobilized lipase on one milligram of magnetic cellulose. During the study of operational stability, lipase immobilized on magnetic cellulose using carbonyldiimidazole activation retained 98% of initial activity after 7 cycles. In the case of sodium periodate
Table 1 Maximum adsorption capacities (Qmax; mg/g) of four magnetically modified adsorbents (one gram of material used during the mechanochemical process) for the tested dyes (Bismarck brown Y and safranin O). Langmuir adsorption isotherm was used for calculation. Magnetic adsorbents
Bismarck brown Y
Safranin O
Grass carp scales Oak acorns Montmorillonite Biogenic iron oxides
Qmax ¼43.3 Qmax ¼20.9 Qmax ¼38.6 Qmax ¼36.4
Qmax ¼ 30.3 Qmax ¼ 12.9 Qmax ¼ 41.4 Qmax ¼ 15.9
Table 2 Activity of lipase immobilized on magnetic cellulose particles using three immobilization methods. CDI, 1,10 -carbonyldiimidazole; BDDE, 1,4-butanediol diglycidyl ether.
Fig. 3. Typical X-ray powder diffraction pattern of magnetically modified material spruce sawdust (top) and separate maghemite nanoparticles prepared by the same mechanochemical procedure (bottom). Vertical lines indicate theoretical positions of most intense maghemite diffractions (taken from PDF card No.: 00-039-1346), asterisks indicate diffractions of remnants of soluble salts.
Immobilization method
Activity of enzyme on magnetic cellulose particles (nkat/mg)
CDI NaIO4 BDDE
1.48 4.09 4.55
206
I. Safarik et al. / Materials Letters 126 (2014) 202–206
adsorbents for organic xenobiotics removal and as a biocompatible carrier for enzymes immobilization. Low-cost, efficient and biocompatible magnetic composites can substantially improve and simplify wide range of biotechnology and environmental technology processes.
Acknowledgments The authors thank Dr. Dalibor Jancik (RCPTM, Palacky University, Olomouc, Czech Republic) and Laboratory of Electron Microscopy (Biological Centre, Ceske Budejovice, Czech Republic) for making SEM images. This research was supported by the Grant Agency of the Czech Republic (Project no. 13-13709S), by the Research Project LH12190 (Ministry of Education, Youth and Sports of the Czech Republic), by Technology Agency of the Czech Republic (Competence Centres, Project no. TE01020218) and by the Operational Program Research and Development for Innovations – European Development Fund (CZ.1.05/2.1.00/03.0058) of the Ministry of Education, Youth and Sports of the Czech Republic.
Fig. 4. Equilibrium adsorption isotherms of Bismark brown Y (top) and safranin O (bottom) on the tested magnetically modified adsorbents (one gram of material used during the mechanochemical process). Ceq: equilibrium liquid-phase concentration of the unadsorbed (free) dye (mg/L); qeq: equilibrium solid-phase concentration of the adsorbed dye (mg/g). ( ♦ ) biogenic iron oxides; ( ) oak acorns; ( ▲ ) montmorillonite; ( ■ ) grass carp scales.
Fig. 5. Operational stability of lipase immobilized on magnetic cellulose particles using three immobilization methods. Chart demonstrates relative residual activity of immobilized enzyme (%) after each cycle. ♦ - CDI activation; ■ - NaIO4 activation; ▲ - BDDE activation. CDI - 1,10 -carbonyldiimidazole; BDDE - 1,4-butanediol diglycidyl ether.
and butanediol diglycidyl ether activation procedures immobilized lipase retained 90 and 83% of initial activity after 7 cycles, respectively (Fig. 5). After storage at 4 1C for the time period of 1 month, lipase immobilized using carbonyldiimidazole retained approximately 97%, using sodium periodate 80% and using butanediol diglycidyl ether 70% of its initial activity, respectively. 4. Conclusions A fast and simple mechanochemical conversion of powdered nonmagnetic materials into their magnetic derivatives has been developed. Magnetically responsive composites have been used as
References [1] Safarik I, Safarikova M. Magnetic nanoparticles for in vitro biological and medical applications: an overview. In: Thanh NTK, editor. Magnetic Nanoparticles: From Fabrication to Biomedical and Clinical Applications. Boca Raton, FL 33487-2742, CRC Press/Taylor and Francis; 2012. p. 215–42. [2] Laurent S, Forge D, Port M, Roch A, Robic C, Elst LV, et al. Magnetic iron oxide nanoparticles: synthesis, stabilization, vectorization, physicochemical characterizations, and biological applications. Chem Rev 2008;108:2064–110. [3] Bellusci M, Aliotta C, Fiorani D, La Barbera A, Padella F, Peddis D, et al. Manganese iron oxide superparamagnetic powder by mechanochemical processing, nanoparticles functionalization and dispersion in a nanofluid. J Nanoparticle Res 2012:14 (Article No. 904). [4] Das D, Sureshkumar MK, Koley S, Mithal N, Pillai CGS. Sorption of uranium on magnetite nanoparticles. J Radioanal Nucl Chem 2010;285:447–54. [5] Lin CR, Chu YM, Wang SC. Magnetic properties of magnetite nanoparticles prepared by mechanochemical reaction. Mater Lett 2006;60:447–50. [6] Beyer MK, Clausen-Schaumann H. Mechanochemistry: the mechanical activation of covalent bonds. Chem Rev 2005;105:2921–48. [7] Lu J, Yang SH, Ng KM, Su CH, Yeh CS, Wu YN, et al. Solid-state synthesis of monocrystalline iron oxide nanoparticle based ferrofluid suitable for magnetic resonance imaging contrast application. Nanotechnology 2006;17:5812–20. [8] Saito T, Koopal LK, Nagasaki S, Tanaka S. Adsorption of heterogeneously charged nanoparticles on a variably charged surface by the extended surface complexation approach: charge regulation, chemical heterogeneity, and surface complexation. J Phys Chem B 2008;112:1339–49. [9] Patterson AL. The Scherrer formula for X-Ray particle size determination. Phys Rev 1939;56:978–82. [10] Srinivasan A, Viraraghavan T. Decolorization of dye wastewaters by biosorbents: a review. J Environ Manag 2010;91:1915–29. [11] Safarik I, Rego LFT, Borovska M, Mosiniewicz-Szablewska E, Weyda F, Safarikova M. New magnetically responsive yeast-based biosorbent for the efficient removal of water-soluble dyes. Enzyme Microb Technol 2007;40:1551–6. [12] Safarik I, Horska K, Safarikova M. Magnetically responsive biocomposites for inorganic and organic xenobiotics removal. In: Kotrba P, Mackova M, Macek T, editors. Microbial Biosorption of Metals. Dordrecht, Heidelberg, London, New York: Springer; 2011. p. 301–20. [13] Varatharajan V, Hoover R, Li JH, Vasanthan T, Nantanga KKM, Seetharaman K, et al. Impact of structural changes due to heat-moisture treatment at different temperatures on the susceptibility of normal and waxy potato starches towards hydrolysis by porcine pancreatic alpha amylase. Food Res Int 2011;44:2594–606. [14] Nakagawa T, Edouard JL, de Beaulieu JL. A scanning electron microscopy (SEM) study of sediments from Lake Cristol, southern French Alps, with special reference to the identification of Pinus cembra and other Alpine Pinus species based on SEM pollen morphology. Rev Palaeobot Palynol 2000;108:1–15. [15] Yang T, Lua AC. Characteristics of activated carbons prepared from pistachionut shells by physical activation. J Colloid Interface Sci 2003;267:408–17.
Contents lists available at ScienceDirect
Materials Letters journal homepage: www.elsevier.com/locate/matlet
Mechanochemical synthesis of magnetically responsive materials from non-magnetic precursors Ivo Safarik a,b,n, Katerina Horska a, Kristyna Pospiskova b, Jan Filip b, Mirka Safarikova a a b
Department of Nanobiotechnology, Institute of Nanobiology and Structural Biology of GCRC, Na Sadkach 7, 370 05 Ceske Budejovice, Czech Republic Regional Centre of Advanced Technologies and Materials, Palacky University, Slechtitelu 11, 783 71 Olomouc, Czech Republic
art ic l e i nf o
a b s t r a c t
Article history: Received 30 October 2013 Accepted 7 April 2014 Available online 15 April 2014
Mechanochemical synthesis of various types of magnetically responsive materials from non-magnetic powdered precursors has been developed. The preparation is based on the mechanochemical conversion of ferrous and ferric ions at the presence of alkaline hydroxide into magnetic iron oxides nanoparticles (maghemite identified by XRD measurements). The presence of powdered nonmagnetic materials during the mechanochemical process led to the efficient deposition of magnetic nanoparticles on the surface of the treated materials in the form of individual nanoparticles and their aggregates. The prepared magnetically responsive materials have been used as adsorbents for xenobiotics removal and as a carrier for enzymes immobilization. & 2014 Elsevier B.V. All rights reserved.
Keywords: Mechanochemistry Magnetic materials Magnetic adsorbents Magnetic carriers
1. Introduction Magnetic nano- and microparticles have attracted an increasing interest in various fields including nanoscience, nanotechnology, biosciences, biotechnology and environmental technology. Many chemical procedures have been used to synthesize magnetic particles, such as classical co-precipitation, reactions in constrained environments (e.g. microemulsions), sol–gel syntheses, sonochemical reactions, hydrothermal reactions, hydrolysis and thermolysis of precursors, flow injection syntheses, electrospray synthesis and microwave synthesis [1,2]. Recently mechanochemical procedures have been used to synthesize magnetic iron oxides and ferrites nanoparticles [3–5]. Mechanochemistry represents one of several ways of chemical activation. In solid-state mechanochemistry, nonthermal chemical reactions occur because of the deformation and fracture of solids, which are technically induced by milling or grinding of the materials. During this process the mechanical energy induces chemical reactions and phase transformations [6]. Up to now, mechanochemical synthesis has been employed mainly for the production of individual iron oxides and ferrites. However, mechanochemistry can be successfully used also for the preparation of magnetically responsive materials from originally
n Corresponding author at: Department of Nanobiotechnology, Institute of Nanobiology and Structural Biology of GCRC, Na Sadkach 7, 370 05 Ceske Budejovice, Czech Republic. Tel.: +420 387775608; fax: +420 385310249. E-mail address: [email protected] (I. Safarik). URL: http://www.nh.cas.cz/people/safarik (I. Safarik).
http://dx.doi.org/10.1016/j.matlet.2014.04.045 0167-577X/& 2014 Elsevier B.V. All rights reserved.
nonmagnetic powdered precursors. In this paper we present a very simple, generally applicable procedure for the preparation of magnetic materials from variety of inorganic, organic and biological precursors, together with the illustration of their possible applications as adsorbents and enzyme carriers. Mechanochemical postmagnetization can be very useful for smart magnetic modification of diverse non-magnetic materials.
2. Materials and methods Materials: FeCl3. 6H2O, FeCl2. 4H2O, montmorillonite, halloysite, Candida rugosa lipase (EC 3.1.1.3), 1,4-butanediol diglycidyl ether (BDDE), sodium (meta)periodate, 1,10 -carbonyldiimidazole (CDI), 4-nitrophenyl butyrate, dimethyl sulfoxide, Bismarck brown Y and sodium acetate were purchased from Sigma-Aldrich, USA. Microcrystalline cellulose, safranin O and 4-nitrophenol were from Lachema, Czech Republic, while the common chemicals were from Lach-Ner, Czech Republic. Finally powdered biological materials with diameters below 1 mm (spruce sawdust, scales from grass carp (Ctenopharyngodon idella), wheat straw, pistachio nut shells, peanut husks, oak acorns, spent coffee grounds), as well as potato starch and pine pollen were obtained locally. The natural ocherous sediment containing biogenic iron oxides was collected using glass vessels from a water stream in Ceske Budejovice (Czech Republic); it was sieved through a 1 mm sieve to remove larger detrital fraction, then repeatedly washed with deionized water and air dried at a temperature not exceeding 50 1C.
I. Safarik et al. / Materials Letters 126 (2014) 202–206
Mechanochemical synthesis of magnetically responsive materials: In the standard procedure a mixture of 1.35 g of FeCl3 6H2O (0.005 mol), 0.50 g of FeCl2 4H2O (0.0025 mol) and 4 g of sodium chloride (inert material used to avoid particles agglomeration) was grounded in a mortar at room temperature for 10 min. Then, appropriate amount of target nonmagnetic powdered material (usually 1–2 g) was added and after thorough mixing the process continued for another 10 min. As the last step, powdered potassium hydroxide (1.22 g) was added and after mixing the grinding continued for 10 min. After KOH addition the mixture became brown. During the grinding process the material was scraped from the mortar wall occasionally. After finishing the mechanochemical process, the magnetically modified material was thoroughly washed with water (to remove soluble impurities and free iron oxides particles) and stored in a suspension, or it was air dried. Structural characterization of iron oxide nanoparticles prepared by mechanochemical procedure: X-ray powder diffraction (XRD) patterns of selected samples were recorded on PANalytical X'Pert PRO instrument in Bragg–Brentano geometry with Fe-filtered CuKα radiation (40 kV, 30 mA). The samples were inserted into conventional front-loading cavity sample holder and scanned in the 2θ range of 10–901 (step size 0.0171). The commercial standards SRM640 (Si) and SRM660 (LaB6) from NIST were used for the evaluation of the line positions and instrumental line broadening, respectively. The acquired patterns were evaluated using the X'Pert HighScore Plus software (PANalytical) together with PDF-4þ database. Adsorption of organic dyes on magnetically responsive composites: 30 mg of selected magnetically modified adsorbents were mixed with 7.0 mL of water in a test tube. Then, 0.1–3 mL of stock water solution of a tested dye (1 mg/mL) was added and the total volume of the solution was made up to 10.0 mL with water. The suspension was mixed on a rotary mixer (Dynal, Norway) for 3 h at room temperature. The magnetic adsorbent was then separated from the suspension using a magnetic separator (MPC-1 or MPC-6, Dynal, Norway) and the clear supernatant was used for the spectrophotometric measurement. The concentration of free (unbound) dye in the supernatant (Ceq) was determined from the calibration curve. The amount of dye bound to the unit mass of the adsorbent (qeq) was calculated using the following formula: qeq ¼ ðC tot C eq Þ=3
ðmg=gÞ
ð1Þ
where Ctot is the total (initial) concentration of dye (μg/mL) used in the experiment. The value qeq was expressed in mg of adsorbed dye per 1 g of adsorbent. Equilibrium adsorption data were fit to Langmuir adsorption isotherms using SigmaPlot software. Immobilization of lipase on magnetic cellulose particles: C. rugosa lipase was immobilized on magnetically modified cellulose particles. Three various agents were used for activation of hydroxyl groups present in the cellulose structure, namely sodium periodate, butanediol diglycidyl ether and carbonyldiimidazole. To prepare samples, 30 mg of prepared magnetic cellulose particles were washed with water and magnetically separated by NdFeB magnet. In the next step, particles were treated by the activating agent. Using 1,10 -carbonyldiimidazole (CDI), 1.5 mL of 0.5% (w/v) solution in dimethyl sulfoxide was added to particles. For the periodate method, 1.5 mL of 1% (w/v) solution of sodium (meta)periodate (NaIO4) in 0.1 M sodium acetate buffer pH 4 was utilized. During activation by the epoxide method, 1.5 mL of 3% (v/v) solution of 1,4-butanediol diglycidyl ether (BDDE) in 0.25 M NaOH was used. Particles were shaken with these activating agents on an automatic rotator (20 rpm) for 24 h at room temperature in the dark. After the modification procedure, particles were magnetically separated, supernatants were poured off and particles were repeatedly washed with distilled water. Subsequently, 1.5 mL of lipase solution (1 mg/mL) in 50 mM potassium phosphate buffer,
203
pH 7.5, was added to the activated particles and shaken on automatic rotator (20 rpm) for 24 h at 4 1C. After the immobilization procedure, particles were magnetically separated, supernatant containing unbound enzyme was removed and particles with bound enzyme were repeatedly washed with buffer until no enzyme activity in the supernatant was detected. Magnetic cellulose particles with immobilized lipase were stored in buffer at 4 1C. Lipase assay: Activity of lipase immobilized on magnetic cellulose particles was determined spectrophotometrically using 0.5 mM 4-nitrophenyl butyrate (dissolved in ethanol) in 50 mM potassium phosphate buffer, pH 7.5. Particles of magnetic cellulose with attached lipase were stirred during the reaction in buffer containing the substrate, then magnetically separated to the bottom of the cuvette to stop the reaction and increasing amount of yellow-colored 4-nitrophenol was measured at 405 nm. Molar absorption (extinction) coefficient (ε) of the reaction product was determined spectrophotometrically for the specific medium conditions used in this enzyme assay. Coefficient for 4-nitrophenol in 50 mM potassium phosphate buffer, pH 7.5 at 405 nm was 13815 L/mol cm. Operational stability of immobilized lipase: Reusability of lipase immobilized on magnetic cellulose particles was tested as its operational stability; it was repeatedly used for 7 reaction cycles. Particles with attached lipase were washed with buffer between each cycle. Activity of lipase was measured spectrophotometrically as described previously. Residual activities of lipase after each cycle were determined and compared taking the initial activity in the first cycle as 100%. Time stability of immobilized lipase: Particles of magnetic cellulose with immobilized lipase were stored in the reaction buffer at 4 1C for 30 days and percentage of residual enzyme activity on the carrier was determined. Possible presence of lipase released from the support was tested during this time period by measuring the activity of free lipase in the supernatant.
3. Results and discussion Mechanochemical synthesis of magnetic composite materials: In mechanochemical synthesis, hydrated solid reactants, namely ferrous and ferric chlorides reacted with KOH during grinding in a mortar to form magnetite nanoparticles. To avoid agglomeration, the excess of sodium chloride was added to the precursors before grinding. The following reaction takes place [7]: 2FeCl3 6H2O(s) þ FeCl2 4H2O(s) þ 8KOH(s)-8KCl(s) þFe3O4(s) þ20H2O(g)
(2)
Magnetite nanoparticles as the primary product are stable only in inert atmosphere [5]. However, in the described process which
Fig. 1. Appearance of original powdered oak acorns suspension (left), suspension of oak acorns powder after magnetic modification (middle) and demonstration of magnetic separation of magnetically modified oak acorns powder (right).
204
I. Safarik et al. / Materials Letters 126 (2014) 202–206
is performed in air, the reaction-generated heat and air oxidation lead to the conversion of magnetite nanoparticles into maghemite as follows [7]: 4 Fe3O4 (s) þO2 (g)-6 γ-Fe2O3 (s)
(3)
The addition of appropriate nonmagnetic powder material to the reaction mixture and grinding before the KOH addition led to
the homogeneous distribution of ferrous and ferric ions within the material matrix. Subsequent addition of KOH followed by grinding led to the formation of magnetite and maghemite nanoparticles bound to the surface and pores of matrix according to Eqs. (2) and (3). The possible influence of the modified material and its chemical composition may lead to the formation of the non-stoichiometric maghemite during the grinding process. Anyway magnetic iron
Fig. 2. Scanning electron microscopy of native and magnetically modified materials. 1 – native potato starch (reproduced with permission from [13]); 2 – magnetic potato starch; 3 – native pine pollen (reproduced with permission from [14]); 4 – magnetic pine pollen; 5 – native pistachio nut shells (reproduced with permission from [15]); 6 – magnetic pistachio nut shells; 7 – native halloysite; 8 – magnetic halloysite.
I. Safarik et al. / Materials Letters 126 (2014) 202–206
oxides nanoparticles bound to the treated material lead to the formation of stable magnetically responsive composite materials, which can be easily separated using a simple magnet or magnetic separator (Fig. 1). The magnetic iron oxide particles deposition is usually very stable, and during the subsequent washing steps only a very low amount of particles were released and washed out. The presence of magnetic iron oxides nanoparticles and their aggregates on the surface of selected magnetically modified materials (magnetic potato starch, pine pollen, pistachio nut shells and halloysite) can be clearly seen using scanning electron microscopy (Fig. 2). It can be expected that the strong binding of magnetic iron oxides particles to the surface of non-magnetic materials has been achieved by a subtle balance of van der Waals, electrostatic and hydrophobic interactions both between the magnetic particles and the treated material surface and between the adsorbed magnetic particles [8]. The structure of iron oxide particles in selected magnetically modified materials (namely spruce sawdust) was identified using XRD as maghemite γ-Fe2O3 (Fig. 3) with observed remnants of soluble salts. The very broad diffraction lines imply nanocrystalline character of the maghemite particles – the size of X-ray coherent domains (i.e., corresponding approximately to mean particle size) calculated according to the Scherrer formula equals to 4 nm [9]. Adsorption of organic dyes on magnetic composites: Recently, large number of low cost, easily available materials have been tested and used as adsorbents for xenobiotics removal [10]. Magnetic modification of such adsorbents simplifies their separation from the treated solutions and suspensions. That is why adsorption properties of several materials magnetized with described mechanochemical procedure (namely powdered magnetically responsive montmorillonite, grass carp scales, oak acorns and biogenic iron oxides) were tested with two water-soluble organic dyes, belonging to different dye classes, namely Bismarck brown Y (diazo dye) and safranin O (safranin dye). It was shown in preliminary experiments that the adsorption of the tested dyes reached equilibrium in approximately 60–120 min. Incubation time of 3 h was used for adsorption experiments. The equilibrium adsorption isotherms for both tested dyes and four magnetic adsorbents (prepared by addition of 1 g of material during the standard mechanochemical procedure) are shown in Fig. 4. The experimental data were analyzed by means of non-linear regression calculation using SigmaPlot software and it was shown that they follow the Langmuir isotherm equation. The Langmuir model is valid for monolayer adsorption onto a surface with a finite number of identical sites. The well-known expression
205
for the Langmuir model is given by qeq ¼
Q max bC eq 1 þbC eq
ð4Þ
where qeq (expressed in mg/g) is the amount of the adsorbed dye per unit mass of magnetic adsorbent and Ceq (expressed in mg/L) is the unadsorbed dye concentration in solution at equilibrium. Qmax (expressed in mg/g) is the maximum amount of the dye per unit mass of adsorbent to form a complete monolayer on the surface bound at high dye concentration and b is a constant related to the affinity of the binding sites (expressed in L/mg) [11]. Because the adsorption of the tested dyes can be successfully described by the Langmuir isotherm, it is possible to calculate the maximum adsorption capacity which is a very important parameter describing the adsorption process (Table 1). In the case of two dyes and four adsorbents tested, the highest value was found for Bismarck brown Y and grass carp scales (43.3 mg/g), while the lowest value was obtained for safranin O and magnetic oak acorns (12.9 mg/g). The maximum adsorption capacities of the developed materials are not so high as observed in the case of some other alternative magnetic biosorbents [12]. The possible reason for lower adsorption is the high coverage of the magnetically modified material surface with magnetic iron oxides (see Fig. 2). In order to test this possibility, two batches of magnetic montmorillonite were prepared, differing by the amount of native montmorillonite added during the standard mechanochemical procedure (one or two grams); higher amount of native material during the mechanochemical process results in lower surface coverage with magnetic iron oxides. The maximum adsorption capacities were 38.6 and 52.4 mg/g for one gram and two gram batches, respectively. Immobilization of lipase on magnetic cellulose particles: Selected magnetically modified materials can be used as carriers for immobilization of enzymes and other important biologically active compounds. C. rugosa lipase was immobilized on magnetic cellulose particles prepared by mechanochemical method. Three reagents, namely sodium periodate, butanediol diglycidyl ether and carbonyldiimidazole were used for activation of hydroxyl groups present in cellulose structure. Table 2 shows the activity of immobilized lipase on one milligram of magnetic cellulose. During the study of operational stability, lipase immobilized on magnetic cellulose using carbonyldiimidazole activation retained 98% of initial activity after 7 cycles. In the case of sodium periodate
Table 1 Maximum adsorption capacities (Qmax; mg/g) of four magnetically modified adsorbents (one gram of material used during the mechanochemical process) for the tested dyes (Bismarck brown Y and safranin O). Langmuir adsorption isotherm was used for calculation. Magnetic adsorbents
Bismarck brown Y
Safranin O
Grass carp scales Oak acorns Montmorillonite Biogenic iron oxides
Qmax ¼43.3 Qmax ¼20.9 Qmax ¼38.6 Qmax ¼36.4
Qmax ¼ 30.3 Qmax ¼ 12.9 Qmax ¼ 41.4 Qmax ¼ 15.9
Table 2 Activity of lipase immobilized on magnetic cellulose particles using three immobilization methods. CDI, 1,10 -carbonyldiimidazole; BDDE, 1,4-butanediol diglycidyl ether.
Fig. 3. Typical X-ray powder diffraction pattern of magnetically modified material spruce sawdust (top) and separate maghemite nanoparticles prepared by the same mechanochemical procedure (bottom). Vertical lines indicate theoretical positions of most intense maghemite diffractions (taken from PDF card No.: 00-039-1346), asterisks indicate diffractions of remnants of soluble salts.
Immobilization method
Activity of enzyme on magnetic cellulose particles (nkat/mg)
CDI NaIO4 BDDE
1.48 4.09 4.55
206
I. Safarik et al. / Materials Letters 126 (2014) 202–206
adsorbents for organic xenobiotics removal and as a biocompatible carrier for enzymes immobilization. Low-cost, efficient and biocompatible magnetic composites can substantially improve and simplify wide range of biotechnology and environmental technology processes.
Acknowledgments The authors thank Dr. Dalibor Jancik (RCPTM, Palacky University, Olomouc, Czech Republic) and Laboratory of Electron Microscopy (Biological Centre, Ceske Budejovice, Czech Republic) for making SEM images. This research was supported by the Grant Agency of the Czech Republic (Project no. 13-13709S), by the Research Project LH12190 (Ministry of Education, Youth and Sports of the Czech Republic), by Technology Agency of the Czech Republic (Competence Centres, Project no. TE01020218) and by the Operational Program Research and Development for Innovations – European Development Fund (CZ.1.05/2.1.00/03.0058) of the Ministry of Education, Youth and Sports of the Czech Republic.
Fig. 4. Equilibrium adsorption isotherms of Bismark brown Y (top) and safranin O (bottom) on the tested magnetically modified adsorbents (one gram of material used during the mechanochemical process). Ceq: equilibrium liquid-phase concentration of the unadsorbed (free) dye (mg/L); qeq: equilibrium solid-phase concentration of the adsorbed dye (mg/g). ( ♦ ) biogenic iron oxides; ( ) oak acorns; ( ▲ ) montmorillonite; ( ■ ) grass carp scales.
Fig. 5. Operational stability of lipase immobilized on magnetic cellulose particles using three immobilization methods. Chart demonstrates relative residual activity of immobilized enzyme (%) after each cycle. ♦ - CDI activation; ■ - NaIO4 activation; ▲ - BDDE activation. CDI - 1,10 -carbonyldiimidazole; BDDE - 1,4-butanediol diglycidyl ether.
and butanediol diglycidyl ether activation procedures immobilized lipase retained 90 and 83% of initial activity after 7 cycles, respectively (Fig. 5). After storage at 4 1C for the time period of 1 month, lipase immobilized using carbonyldiimidazole retained approximately 97%, using sodium periodate 80% and using butanediol diglycidyl ether 70% of its initial activity, respectively. 4. Conclusions A fast and simple mechanochemical conversion of powdered nonmagnetic materials into their magnetic derivatives has been developed. Magnetically responsive composites have been used as
References [1] Safarik I, Safarikova M. Magnetic nanoparticles for in vitro biological and medical applications: an overview. In: Thanh NTK, editor. Magnetic Nanoparticles: From Fabrication to Biomedical and Clinical Applications. Boca Raton, FL 33487-2742, CRC Press/Taylor and Francis; 2012. p. 215–42. [2] Laurent S, Forge D, Port M, Roch A, Robic C, Elst LV, et al. Magnetic iron oxide nanoparticles: synthesis, stabilization, vectorization, physicochemical characterizations, and biological applications. Chem Rev 2008;108:2064–110. [3] Bellusci M, Aliotta C, Fiorani D, La Barbera A, Padella F, Peddis D, et al. Manganese iron oxide superparamagnetic powder by mechanochemical processing, nanoparticles functionalization and dispersion in a nanofluid. J Nanoparticle Res 2012:14 (Article No. 904). [4] Das D, Sureshkumar MK, Koley S, Mithal N, Pillai CGS. Sorption of uranium on magnetite nanoparticles. J Radioanal Nucl Chem 2010;285:447–54. [5] Lin CR, Chu YM, Wang SC. Magnetic properties of magnetite nanoparticles prepared by mechanochemical reaction. Mater Lett 2006;60:447–50. [6] Beyer MK, Clausen-Schaumann H. Mechanochemistry: the mechanical activation of covalent bonds. Chem Rev 2005;105:2921–48. [7] Lu J, Yang SH, Ng KM, Su CH, Yeh CS, Wu YN, et al. Solid-state synthesis of monocrystalline iron oxide nanoparticle based ferrofluid suitable for magnetic resonance imaging contrast application. Nanotechnology 2006;17:5812–20. [8] Saito T, Koopal LK, Nagasaki S, Tanaka S. Adsorption of heterogeneously charged nanoparticles on a variably charged surface by the extended surface complexation approach: charge regulation, chemical heterogeneity, and surface complexation. J Phys Chem B 2008;112:1339–49. [9] Patterson AL. The Scherrer formula for X-Ray particle size determination. Phys Rev 1939;56:978–82. [10] Srinivasan A, Viraraghavan T. Decolorization of dye wastewaters by biosorbents: a review. J Environ Manag 2010;91:1915–29. [11] Safarik I, Rego LFT, Borovska M, Mosiniewicz-Szablewska E, Weyda F, Safarikova M. New magnetically responsive yeast-based biosorbent for the efficient removal of water-soluble dyes. Enzyme Microb Technol 2007;40:1551–6. [12] Safarik I, Horska K, Safarikova M. Magnetically responsive biocomposites for inorganic and organic xenobiotics removal. In: Kotrba P, Mackova M, Macek T, editors. Microbial Biosorption of Metals. Dordrecht, Heidelberg, London, New York: Springer; 2011. p. 301–20. [13] Varatharajan V, Hoover R, Li JH, Vasanthan T, Nantanga KKM, Seetharaman K, et al. Impact of structural changes due to heat-moisture treatment at different temperatures on the susceptibility of normal and waxy potato starches towards hydrolysis by porcine pancreatic alpha amylase. Food Res Int 2011;44:2594–606. [14] Nakagawa T, Edouard JL, de Beaulieu JL. A scanning electron microscopy (SEM) study of sediments from Lake Cristol, southern French Alps, with special reference to the identification of Pinus cembra and other Alpine Pinus species based on SEM pollen morphology. Rev Palaeobot Palynol 2000;108:1–15. [15] Yang T, Lua AC. Characteristics of activated carbons prepared from pistachionut shells by physical activation. J Colloid Interface Sci 2003;267:408–17.