Materials Letters 74 (2012) 163–166

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Materials Letters

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Self-assembly of phosphorylated poly(ethyleneimine) for use as biomimetictemplates in the formation of hybrid hollow silica spheres

Rui Zhao a,⁎, Bao-Lian Su b

a Research Branch of Functional Polymer Composites, Institute of Microelectronic & Solid State Electronic, University of Electronic Science and Technology of China,Chengdu 610054, PR Chinab Laboratory of Inorganic Materials Chemistry, University of Namur (FUNDP), 61 rue de Bruxelles, B-5000 Namur, Belgium

⁎ Corresponding author. Tel./fax: +86 28 832 073 26E-mail address: ruizhao@uestc.edu.cn (R. Zhao).

0167-577X/$ – see front matter © 2012 Elsevier B.V. Aldoi:10.1016/j.matlet.2012.01.023

a b s t r a c t

a r t i c l e i n f o

Article history:Received 17 July 2011Accepted 6 January 2012Available online 18 January 2012

Keywords:BiomaterialsPhosphorylated polyethyleneimineHollow silica spheresSelf-assemblySol–gel preparation

The submicron scale hybrid hollow silica spheres have been fabricated by employing phosphorylated poly-ethyleneimine as template/scaffold/catalysis in a mild self-assembly biomineralization approach. The highlyphosphorylated polyethyleneimine was shown to be a necessary prerequisite for in vitro formation of hollowsilica spheres. In the presence of a neutral phosphate buffer, the monovalent anions H2PO4

− and divalentanions HPO4

2− acted not only as the buffer ions but also as ligands, regulating the shape and morphologyof the PEI aggregation, which resulted in the final hollow morphology of silica. The hollow silica particleswere very uniform in size, with a diameter of 0.8–1.0 μm and a shell thickness of ~70 nm.

© 2012 Elsevier B.V. All rights reserved.

1. Introduction

Recently, inorganic hollow nanoparticles, owing to their low den-sity, large specific area, high mechanical and thermal stabilities, andsurface permeability, have attracted much attention for a fundamentalinterest in biomineralization and potential applications in catalysis,cosmetics, coatings, composite materials, dyes, inks, artificial cells,and fillers.[1–3] Thus, variousmethods, such as templating [4–8], sono-chemical [9], reverse microemulsion [10,11], and hydrothermalmethods [12–14], have been reported as suitable procedures for thepreparation of inorganic materials with hollow sphere structures. Onthe other hand, the synthesis of inorganic materials using biomimeticapproaches has become an important area of research. Biomimetic syn-thesis is a potential route for the synthesis of inorganicmaterials undermild conditions, and numerous studies have already demonstrated theuse of bio-templates in the synthesis of inorganic materials [2,15–17].In the biological world, the organic matrix silicatein (proteins), silaffin(peptides) and long-chain polyamines in marine organisms play animportant role in the synthesis andmorphology of inorganicmaterials.

In the last decade, polyamines including repeating N-methylpropylamine fragments terminating with NH2 or NMe2 groups,isolated from siliceous cell walls of diatoms, have been successfullyused as reaction templates, not only for biosilicification [17–21] butalso for the formation of other minerals [22–23]. Polyamines are

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known to catalyze siloxane-bond formation and can act as flocculatingagents [24]. Moreover, it was discovered that polyamines form aggre-gates if phosphate ions are added to the solution and that the phosphateconcentration controls the size of the resulting silica particles [16,25]. Inaqueous solution polyamines form aggregates with positively chargedsurfaces, and multivalent anions, promoting higher-order assembliesof the emulsion droplets. Silicic acid derivatives may be adsorbed onand/or dissolved in the polyamine droplets, whereby they form a coac-ervate (a “liquid precipitate”), which finally hardens by silica formation.This mechanism may explain the observed correlation between polya-nion concentration and the size of the resulting silica nanosphere[15–16]. However, pattern formation or other specially designed struc-tures, requires a polyamine-based mechanism that produces an intri-cate frame-work of silica rather than a population of silica spheres.

The important feature of this work is that the submicron scale hybridhollow silica spheres have been fabricated exploiting highly phosphory-lated polyethyleneimine (PEI) as template/scaffold/catalysis in a mildself-assembly biomineralization approach. The highly phosphorylatedpolyethyleneimine (PEI) was shown to be a necessary prerequisite forin vitro formation of hollow silica spheres. Using a phosphate bufferat neutral pH, with a stoichiometric ratio of phosphate and PEI, tetra-methoxysilane (TMOS) rapidly polymerizes to produce polyaminehydride nanoparticles under vigorous shaking, during this phase-separation stage, according to previous research from other groups[16–25], the silica nanoparticles precipitated within several minutes.The mixture was then transferred to an autoclave and kept statically at80 °C for a defined period, within 3 days, the phosphorylated PEI stabi-lized silica nanoparticles grew and aggregated into hollow spheres.

164 R. Zhao, B.-L. Su / Materials Letters 74 (2012) 163–166

2. Experimental section

2.1. Synthesis

All chemicals were used as received without further purification.In a typical synthesis of hybrid phosphorylated PEI hollow silicaspheres, 0.08 g of branched polyethylenimine (average MW ~25,000,Sigma-Aldrich) was dissolved in 8 g of pH=6.8 phosphate buffer solu-tion [c(Na2HPO4)=c(NaH2PO4·H2O)=0.12 mol·L−1] to form a homo-geneous solution under stirring at room temperature. To this solutionwas added 150 μl of TMOS (Sigma-Aldrich), after vigorously shakingfor seconds, the white solid precipitated. The resultant mixture wasdivided; one part was kept at room temperature without agitation,the other was kept at 80 °C for a defined period. After 3 days, both ofthe two samples are filtered, washed with water at least three times,and then dried in air.

2.2. Characterization

Scanning electron microscopy (SEM), transmission electronmicros-copy (TEM) and EDX analysis were carried out with a Philips XL-20 at25 keV and a Philips CM-20 at 200 kV, respectively. The specimens forTEM observation were prepared by epoxy-resin-embedded micro

Fig. 1. SEM images of PEI hybrid hollow silica sphere

sectioning and mounted on a copper grid. Solid-state 31P magic anglespinning (MAS) nuclear magnetic resonance (NMR) spectra wererecorded on a BrukerMSL-400 spectrometer operating at resonance fre-quencies of 161.9 MHz and a 2.5 μs (θ=π/6) pulse with a repetitiontime of 10 s. Chemical shifts were indicated using an external 85%H3PO4 reference.

The specific surface areawas determined by the Brunauer–Emmett–Teller (BET) method, and the pore size distribution was obtained fromthe N2 adsorption branch of the isotherms using the Barrett–Joyner–Halenda (BJH) method.

Fourier transform infrared (FT-IR) spectroscopy was performedwith a Perkin-Elmer Spectrum 2000 spectrometer. The KBr pellettechnique was used for the framework vibration characterization.

3. Results and discussion

PEI existing in either linear or branched form is a strong coordinatorto form complexes. At low pH, the amine groups in PEI are protonatedmaking it a highly positively charged polyelectrolyte; at high valueof pH, it is essentially a neutral polymer. An aggregation of the PEImolecules is a prerequisite for their silica-precipitating function. Theobserved aggregation and phase separation can be explained by theattractive electrostatic interactions between the positively charged

s. 80 °C for 2 days (a–d); 80 °C for 3 days (e–f).

165R. Zhao, B.-L. Su / Materials Letters 74 (2012) 163–166

polyaminemolecules and the negatively charged phosphate ions and orthe formation of hydrogen bonds. As other research groups have pub-lished, silica precipitation takes place only above a threshold value[25], below this, no silica precipitates at all, and a rapid increase of theaggregation diameter is observed beyond this threshold value. In thepresent work, multivalent anions efficiently inducing the PEI aggrega-tion and microscopic phase separation are more complicated. WhenHCl/NaOH was used to keep the pH of the system at the neutral 6.8circumstances, single phosphate ions with different valences, such asmonovalent anions H2PO4

−, divalent anionsHPO42− and trivalent anions

PO43− introduced into the aqueous solutions of PEI, can easily precipi-

tate the silica from the system in a nano-spheremorphology. No hollowstructures occur regardless of crystallization time or crystallization tem-perature. Only when both monovalent anions H2PO4

− and divalentanions HPO4

2− coexist in the aqueous solution of PEI, can hollow silicaspheres be successfully obtained. In order to detect this interactionbetween PEI and phosphate buffer, the induced 31P nuclear magneticresonance (NMR) chemical shift, Δδ, caused by this interaction wasmeasured. From the 31P NMR showed only one single 31P NMR signaldue to orthophosphate molecules contained in the samples. The chem-ical shifts observed for the phosphate buffer, phosphorylated polyethy-leneimine and the hollow silica spheres are 1.71 ppm, 2.78 ppm, and2.10 ppm respectively. Obviously, the formation of the silica precipitateinfluences the state of inorganic phosphate present within the samples.Furthermore, it is interesting to note that the spectrum of the phos-phorylated polyethyleneimine does not exhibit so-called spinning

0

200

400

600

800

1000

1200

1400

1600

1800

2000

Inte

nsity

0 2 4

C

O

Si

P

Fig. 2. TEM images and EDX analysis of PEI hyb

sidebands. It is, therefore, concluded that the phosphate ions in phos-phorylated polyethyleneimine are highly mobile which results in thedisappearance of the spinning sidebands [26,27]. In contrast, the spec-trum of the silica precipitate exhibits spinning sidebands which are,however, relatively weak. That means, silicification of the polyaminedroplets causes a lower mobility of the embedded phosphate ions.Since the spinning sidebands are weak, it is, however, concluded thatthe phosphate ions are still mobile. It is possible that the charge ofthe ions cannot be the only factor determining the phase separationprocess. In other words, the phosphorylated polyethyleneimine ag-gregation is not simply an electrostatic effect. From the experimentshere, it is believed that the monovalent anions H2PO4

− and divalentanions HPO4

2− acted not only as the buffer ions but also as kinds ofregulating the shape and morphology of the PEI aggregation. Detailedinfluences of the ionic strength of monovalent anions H2PO4

− and di-valent anions HPO4

2− on PEI and the effects on the final morphologyof the hollow structures will be discussed thoroughly elsewhere.

Besides the effects of the phosphate ratio, the concentration of PEIin phosphate buffer also plays an important role in the formation ofhollow silica spheres. Herein under the fixed ionic phosphate activity,a high concentration of PEI (above 2 wt.%) in the buffer can only pre-cipitate large silica gel; low concentration of PEI (below 0.5 wt.%) inthe buffer also doesn't yield hollow structures after crystallization.In this paper, all of the samples unless otherwise stated, are synthe-sized in 1 wt.% of PEI in pH=6.8 phosphate buffer solution with amole concentration of phosphate of 0.12 mol·L−1.

6 8 10

Cu

Cu

rid hollow silica spheres. 80 °C for 2 days.

0,00,0 0,20,2 0,40,4 0,60,6 0,80,8 1,01,00

100100

200200

300300

400400

500500

6,83,8

0 5 10 15 200,00

0,05

0,10

0,15

0,20

Ad

sorb

ed V

olu

me

(cm

3 /g

)

Pore Diameter (nm)

Ad

sorb

ed V

olu

me

(cm

3 /g

)

Pression relative P/P0

Fig. 3. The N2 sorption isotherm and pore size distribution curve (inset) of hollow silicasphere.

166 R. Zhao, B.-L. Su / Materials Letters 74 (2012) 163–166

Scanning electron microscope (SEM) images in Figure 1 (a–d)show that the hollow silica spheres have an average particle diameterof 0.8–1.0 μm. From the transmission electron microscopy (TEM) im-ages in Fig. 2, one can see that the aggregated monolayer sphereshave diameters of ~800 nm and a wall thickness of ~70 nm. It is sur-prising that the surfaces of the large hollow spherical silica are notsmooth but coated with small “particles”. The detailed informationof the outer surfaces of large hollow silica spheres can be observedat a higher magnification (Fig. 1d). Both the SEM and TEM imagesclearly show that a big hollow sphere is homogeneously coveredwith small silica particles, resembling the morphology of a hollowstructure. Other exciting images can be found in Fig. 1b and Fig. 2(marked with arrows), which showed that some peanut-like hollowsilica also precipitated from the reaction systems. We here proposea colloidal interaction model, during the 2 day hydrothermal treat-ment, the silica nanoparticles, which were originally stabilized byhighly phosphorylated PEI polyamines, collide and aggregate to eachother randomly: some of them subsequently and independently growinto the single hollow spheres, others acquire a peanut morphology.Transient states of peanut-like silica, either broke into two hollowspheres, or just kept their primarymorphology. Other evidence support-ing the colloidal interaction model came from 3-day-hydrothermal-treatment samples. From Fig. 1e and f, outer surface silica has begunto peel off, and this phenomenon got more and more severe when thehydrothermal treatments increased to more than 4 days. Consideringthe FT-IR and TGA profiles, the IR spectra of the all samples showedthe characteristic bands of silica at 460 cm−1 (Si\O\Si band),791 cm−1 (Si\O\Si symmetric stretch), 980 cm−1 (Si\OH stretch),1082 cm−1 and 1200 cm−1 (Si\O\Si antisymmetric stretches). Theamide peak at 1474 cm−1 and 1637 cm−1, corresponding to vibrationsof the polypeptide chain of PEI, can be observed for the precipitateobtained at pH 6.8. From TGA profiles, which showed weight lossesdue to the PEI encapsulated within the resultant hollow spheres, totalweight losses for the precipitates obtained at pH 6.8 were calculated tobe 30%. Thus, it is reasonable to assume that PEI served as a

coaggregation factor and that it is tightly encapsulated within the silicamatrix, which weakened the mechanical and thermal stabilities of thehollow silica spheres. Elemental composition of these productswas con-firmed by energy-dispersive spectroscopy, indicating that the specimenshown in Fig. 2 consisted of silicon and oxygen and some phosphate.

The N2 sorption analysis was also employed to observe the porestructure of the hollow silica spheres. The adsorption branch in Fig. 3shows a major capillary condensation step at relative pressures of~0.8–0.9. A pore size distribution, calculated from the adsorptionbranch by the Barrett–Joyner–Halenda (BJH) method (inset of Fig. 3)centered at ~3.8 nm, is correlated to the cavity of the outer surface ofthe hollow silica, and the irregular pore size distribution, in the rangeof 6.8–8.0 nm, may be associated with the cavity of large hollow silicaspheres, in accordance with the TEM results. The Brunauer–Emmett–Teller (BET) surface area is 290 m2·g−1.

4. Conclusion

Hollow silica sphere materials have been synthesized via biomin-eralization processes using a neutral phosphate buffer media. In thisapproach, it is crucial to design a system in which PEI micelles willbe modified by specially aggregated morphology. It is anticipatedthat novel materials with complex morphologies can be designedvia a biomineralization approach.

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