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The co-effect of collagen and magnesium ions on calcium carbonate biomineralization
Materials Science and Engineering C 26 (2006) 648 – 652 www.elsevier.com/locate/msec
The co-effect of collagen and magnesium ions on calcium carbonate biomineralization Yunfeng Jiao, Qingling Feng *, Xiaoming Li Department of Materials Science and Engineering, Biomaterials Laboratory, Tsinghua University, Beijing 100084, People’s Republic of China Received 1 November 2004; received in revised form 6 July 2005; accepted 6 August 2005 Available online 10 November 2005
Abstract The process of calcium carbonate biomineralization in the solution containing collagen and magnesium ions was studied in this paper. The results were characterized by using powder X-ray diffraction (XRD) and scanning electron microscopy (SEM). The effect rules were obtained by the cooperation of collagen and magnesium ions in different concentration. The experiment results showed that in the presence of both collagen and magnesium ions, aragonite and vaterite were precipitated at low Mg/Ca ion concentration ratio, while only aragonite with regular spherical morphology was precipitated at high Mg/Ca ion concentration ratio. It indicated that collagen has a promotional effect on magnesium ions in controlling the polymorph of calcium carbonate crystal. A much wider range of calcium carbonate morphologies was observed in the presence of both collagen and magnesium ions. The experiments suggested that collagen acts in combination with magnesium ions to inhibit calcite crystal growth, while favoring the formation of aragonite crystals. D 2005 Elsevier B.V. All rights reserved. Keywords: Calcium carbonate; Collagen; Magnesium ion; Biomineralization
1. Introduction Many studies have been carried out on the mechanisms involved in biomineralization processes and several new biologically inspired synthetic routes have been designed for control of the formation of the mineral phase. One of the most intensely examined systems is calcium carbonate, which is abundant in biominerals, but also of industrial importance due to its wide use as a filler in paints, plastics, rubber, or paper. It has been shown that the polymorphism, morphology and structural properties of calcium carbonate can be controlled by the use of specific organic templates and/or additives. Langmuir monolayers [1,2], ultrathin organic films [3], self-assembled films [4], have been used as effective templates for the controlled growth of calcium carbonate crystals, focussing on the control of the polymorph and crystal orientation. Crosslinked gelatin films [5], polymer substrates [6], and crystal-imprinted polymer surfaces [7] have also recently been used to direct the
* Corresponding author. Tel.: +86 10 62782770; fax: +86 10 62771160. E-mail address: [email protected] (Q. Feng). 0928-4931/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.msec.2005.08.038
controlled growth of calcium carbonate crystals. Calcium carbonate films have been successfully prepared in the presence of both organic substrates and soluble polymeric additives [8,9]. Interestingly, a designed peptide has been synthesized and used for the conformation-dependent control of the calcite morphology [10]. Proteins isolated from mollusk shells [11,12] and intracrystalline macromolecules from sea urchin spines [13] have shown distinct control on the polymorph of calcium carbonate crystals. The modulation of collagen on crystal morphology of calcium carbonate has been studied in vitro [14]. Manoli induced calcium carbonate crystals by chitin and elastin and obtained calcite crystals in both cases [15,16]. Investigations show that proteins can modulate calcium carbonate morphology and also the polymorph [17 – 19]. Magnesium is known to exert a significant effect on calcium carbonate precipitation and, when present in sufficient concentration, generally results in the precipitation of aragonite rather than the thermodynamically favoured phase, calcite [20 – 24]. Recently, many groups have studied the role of magnesium in calcium carbonate precipitation. Meldrum studied the role of magnesium in stabilising
Y. Jiao et al. / Materials Science and Engineering C 26 (2006) 648 – 652
amorphous calcium carbonate and controlling calcite morphologies [25]. Dawe studied the influence of magnesium on the kinetics of calcite precipitation and calcite crystal morphology [26]. Many studies on the combined effect of organic additives and magnesium ions on calcium carbonate crystal growth have been reported recently [27 – 29], but little research has been carried out on the cooperative influence of protein and magnesium ions on calcium carbonate precipitation. This paper mainly discusses the cooperative influence of collagen and magnesium ions on calcium carbonate precipitation. The aim of these experiments is to find the effect on the polymorph and morphology of calcium carbonate by combined collagen and magnesium ions.
2.1. Preparation of collagen Acid-soluble collagen type I was obtained from rat-tail tendon using a protocol identical to that described by Pins et al. [30]. The rat-tail tendon was dissolved in 10mM HCl at room temperature for 7 h. The solution was centrifuged at 15,000 rpm at 4 -C for 45 min, and then filtered. 0.7 M NaCl was added to induce precipitation, then the precipitate was collected by centrifugation (15,000 rpm, 4 -C, 30 min) and redissolved in 10 mM HCl. Acid-soluble mixture was gathered by dialysis against an aqueous phosphate buffer (20 mM disodium hydrogen phosphate, pH 7.4) at 4 -C for 12 h. The precipitated collagen was collected by centrifugation (15,000 rpm, 4 -C, 45 min), and then dissolved into a concentrated solution by dialyzing the pellets against a large volume of 10 mM HCl at room temperature for 12 h. The collagen solution was stored at 4 -C. 2.2. Method of synthesizing crystals In our experiment, the method of synthesizing crystals was according to the report of Aizenberg et al. [31]. The crystals were grown by slow diffusion (about 7 days) of NH4HCO3 vapor into cell-culture dishes containing magnesium and calcium chloride solutions mixed with collagen solutions at room temperature in a closed desiccator. The amounts of Ca and Mg and the Mg:Ca ratio in the initial solutions are given in Table 1. The solutions were combined in a beaker and were stirred for 2 min before placing glass cover slips on the bottom of the cell-culture dishes. The stirring was stopped and the solution put in a closed dessiccator for crystal precipitation. After crystalli-
Table 1 Composition of precipitation solutions 2
CaCl (mol) MgCl2 (mol)
zation the cover glass-slips were dried in an oven at 40 -C for 12 h. 2.3. Characterization of synthesized crystals X-ray diffraction (XRD),using a D/max-RB X-ray diffractometer with 40 keV CuKa radiation, was employed to analyze the crystal structure and orientation. The morphologies of the crystals were observed under a Hitachi S-450 scanning electron microscope (SEM) after the crystal had grown for 7 days and deposited with gold by sputtering. 3. Results and discussion 3.1. The influence of collagen and magnesium on polymorph of CaCO3
2. Experimental procedure
Mg : Ca
649
0:1
1:1
2:1
3:1
4:1
5:1
0.02 0
0.02 0.02
0.02 0.04
0.02 0.06
0.02 0.08
0.02 0.1
From XRD it can be seen that the polymorph of CaCO3 changed with increasing the concentration of magnesium in the solution containing collagen. When the ratio of Mg/Ca is 0 or 1 only calcite crystals are formed (Fig. 1a), and when the ratio of Mg/Ca is 2 some vaterite and aragonite crystals are precipitated besides calcite (Fig. 1b). The proportion of vaterite and aragonite crystals increased with the increased concentration of magnesium, and when the ratio of Mg/Ca attain 4 most crystals are aragonite, only a few are calcite crystals (Fig. 1c.). While nearly all crystals are aragonite when the ratio of Mg/Ca is 5 (Fig. 1d.). When there is no collagen present, only at ratio of Mg/Ca of 4 or above that aragonite can be seen (Fig. 1e), in other instance only calcite can be seen, which is in agreement with conventional theoretical results. Magnesium is known to induce aragonite formation from sea water and in vitro at ratio of Mg / Ca equal to or greater than 4, while at lower Mg / Ca ratio mostly calcite and magnesian calcite are formed [32]. When collagen is present alone the growth of CaCO3 crystals is greatly inhibited, but there is no obvious change in CaCO3 polymorph and only calcite crystals are formed [14]. The above results show that when collagen and magnesium are both present, aragonite crystals precipitated at a lower Mg / Ca ratio of 2. This indicates that collagen has a promotional effect on magnesium ions in controlling the polymorph of CaCO3 crystals. The reason of this kinetic phenomenon is considered as contribute to two aspects: on the one hand, magnesium ions inhibit the growth of calcite. The partially dehydrated magnesium ions attach to the surface of the nascent calcite nucleus, the strongly bound residual hydration sphere poisons the surface [33], and inhibits subsequent growth. At the same time, the collagen acts with magnesium and increase the magnesium hydrate absorbed in calcite nucleus. On the other hand, owing to the polymorph of calcium carbonate is relate to its energy state [34], magnesium is likely also to react with collagen and to change the stereochemical structure of collagen molecules, and thus induces the aragonite or vaterite with higher energy in high energy state structure.
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Fig. 1. XRD data for calcium carbonate crystals modulated by collagen and magnesium ions (C: calcite, V: vaterite, A: aragonite the concentration of collagen is 0.1 g/l, calcium ion is 0.02 mol/l). (a) Calcite crystals at Mg/Ca:0 or 1. (b) Calcite crystals with some vaterite and aragonite at Mg/Ca:2. (c) Aragonite crystals with a few calcite at Mg/Ca:4. (d) Aragonite crystals at Mg/Ca:5 (e) Aragonite crystals form besides calcite at Mg/Ca:5 when no collagen present.
3.2. The influence of collagen and magnesium on morphology of CaCO3 A much wide range of calcium carbonate morphologies is generated in the presence of both collagen and magnesium ions. When the concentration of protein is 0.1 g/l. The crystals grow in absence of magnesium are irregular rhombohedral calcite with little distortion, about 120 um in size (Fig. 2a). At low magnesium concentration (Mg / Ca ratio is 1), irregular lumpish calcite crystals are formed with lamellar growth structure (Fig.
2b). By increasing the concentration of magnesium, discoid and dumbbell crystals are obtained (Fig. 2c, d, e). When the concentration of magnesium is high enough (Mg / Ca ratio attains to 5), spherical aragonite crystals are precipitated, about 150 um in size (Fig. 2f). Keeping magnesium concentration unchanged but increasing the concentration of collagen (from 0.1 g/l to 0.4 g/l) spherical aragonite crystals with more regular shape can be obtained(Fig. 2g). In contrast to this, when there is no collagen in the solution we get clusters of aragonite crystals with needle shape at higher magnesium concentration (Fig. 2h).
Y. Jiao et al. / Materials Science and Engineering C 26 (2006) 648 – 652
651
Fig. 2. SEM morphologies of the calcium carbonate crystals precipitated in the solution, collagen concentration: 0.1 g/l. (a) Irregular rhombohedral calcite crystal grown in the solution without magnesium. (b) Irregular lumpish crystals with lamellar growth structure (Mg/Ca:1). (c, d, e) Discoid and dumbbell calcium carbonate crystals. (f) Spherical aragonite crystals at higher magnesium concentration (Mg/Ca:5). (g) Spherical aragonite crystals with more regular shape (Mg/Ca:5, collagen concentration:0.4 g/l). (h) Aragonite crystals with needle shape without collagen (Mg/Ca:5).
Above results showed that in the presence of collagen irregular rhombohedral calcite with lamellar structure can be obtained. When magnesium is added, the morphology of crystals changed greatly, from lumpish to dumbbell, and spherical. The morphological changes of calcium carbonate crystals reveal that in the presence of both collagen and magnesium ions, we tend to obtain spherical aragnite spherulites instead of other forms, especially when the concentration of magnesium is high. Through controlling the concentration of collagen and magnesium, spherical crystals with more regular shapes could be obtained.
itation. The results indicated that collagen has a promotional effect on magnesium ions in controlling the polymorph of CaCO3 crystals. By cooperation of collagen and magnesium ions in different concentration, crystals showed a sequence of morphology changes and especially, aragonite crystals with regular spherical morphology were precipitated at high Mg/Ca ion concentration ratio. The experiments suggested that collagen acts in combination with magnesium ions to inhibit calcite crystal growth, while favor the formation of aragonite crystals.
4. Conclusion
Acknowledgements
The present paper mainly studied the co-effect of collagen and magnesium ions on calcium carbonate precip-
This work was supported by the National Natural Science Foundation of China, Grant No.50272035.
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Y. Jiao et al. / Materials Science and Engineering C 26 (2006) 648 – 652
References [1] D.J. Ahn, A. Berman, D. Charych, J. Phys. Chem. 100 (1996) 12455. [2] A.L. Litvin, S. Valiyaveettil, D.L. Kaplan, S. Mann, Adv. Mater. 9 (1997) 124. [3] D.D. Archibald, S.B. Qadri, B.P. Gaber, Langmuir 12 (1996) 538. [4] J. Aizenberg, A.J. Black, G.M. Whitesides, Nature 398 (1999) 495. [5] G. Falini, S. Fermani, M. Gazzano, A. Ripamonti, Chem. Eur. J. 4 (1998) 1048. [6] E. Dalas, P. Klepetsanis, P.G. Koutsoukos, Langmuir 15 (1999) 8322. [7] S.M. D_Souza, C. Alexander, M.J. Whitcombe, E.N. Vulfson, Nature 398 (1999) 312. [8] G. Xu, N. Yao, I.A. Aksay, J.T. Groves, J. Am. Chem. Soc. 120 (1998) 11977. [9] S. Zhang, K.E. Gonsalves, Langmuir 14 (1998) 6766. [10] D.B. DeOliveira, R.A. Laursen, J. Am. Chem. Soc. 119 (1997) 10627. [11] G. Falini, S. Albeck, S. Weiner, L. Addadi, Science 271 (1996) 67. [12] Y. Levi, S. Albeck, A. Brack, L. S.Weiner, Chem. Eur. J. 4 (1998) 389. [13] J. Aizenberg, J. Hanson, S. Weiner, L. Addadi, J. Am. Chem. Soc. 119 (1997) 881. [14] F.H. Shen, Q.L. Feng, C.M. Wang, J. Cryst. Growth 242 (2002) 239. [15] F. Manoli, E. Dalas, J. Cryst. Growth 201 (1999) 369. [16] F. Manoli, S. Koutsopoulos, E. Dalas, J. Cryst. Growth 182 (1997) 116.
[17] T. Kato, T. Suzuki, T. Amamiya, N. Komiyama, Supramol. Sci. 5 (1998) 411. [18] T. Kato, T. Suzuki, T. Irie, Chem. Lett. 2 (2000) 186. [19] M. Vucak, J. Peric, M.N. Pons, S. Chanel, Power Technol. 101 (1999) 1. [20] M.M. Reddy, K.K. Wang, J. Cryst. Growth 50 (1980) 470. [21] A. Mucci, J.W. Morse, Geochim. Cosmochim. Acta 47 (1983) 217. [22] G. Falini, M. Gazzano, A. Ripamonti, Chem. Commun. (1996) 1037. [23] G. Falini, S. Fermani, M. Gazzano, A. Ripmonti, Chem. Eur. J. 3 (11) (1997) 1807. [24] S. Raz, S. Weiner, L. Addadi, Adv. Mater. 12 (1) (2000) 38. [25] R.M. Wilson, R. Seshadri, F.C. Meldrum, J. Cryst. Growth 254 (2003) 206. [26] Y. Zhang, R.A. Dawe, Chem. Geol. 163 (2000) 129. [27] G. Falini, M. Gazzano, A. Ripamonti, J. Cryst. Growth 137 (1994) 577. [28] A. Sugawara, T. Kato, Chem. Commun. (2000) 487. [29] F.C. Meldruma, S.T. Hyde, J. Cryst. Growth 231 (2001) 544. [30] G.D. Pins, D.L. Chirstiansen, R. Patel, F.H. Silver, Biophys. J. 73 (1997) 2164. [31] J. Aizenberg, J. Hanson, T.F. Koetzle, S. Weiner, L. Addadi, J. Am. Chem. Soc. 119 (1997) 881. [32] Y. Kitano, Bull. Chem. Soc. Jpn. 35 (1962) 1973. [33] R.M. Noyes, J. Am. Chem. Soc. 84 (1962) 513. [34] J.W. McCanley, R. Roy, Am. Mineral. 59 (1974) 947.
The co-effect of collagen and magnesium ions on calcium carbonate biomineralization Yunfeng Jiao, Qingling Feng *, Xiaoming Li Department of Materials Science and Engineering, Biomaterials Laboratory, Tsinghua University, Beijing 100084, People’s Republic of China Received 1 November 2004; received in revised form 6 July 2005; accepted 6 August 2005 Available online 10 November 2005
Abstract The process of calcium carbonate biomineralization in the solution containing collagen and magnesium ions was studied in this paper. The results were characterized by using powder X-ray diffraction (XRD) and scanning electron microscopy (SEM). The effect rules were obtained by the cooperation of collagen and magnesium ions in different concentration. The experiment results showed that in the presence of both collagen and magnesium ions, aragonite and vaterite were precipitated at low Mg/Ca ion concentration ratio, while only aragonite with regular spherical morphology was precipitated at high Mg/Ca ion concentration ratio. It indicated that collagen has a promotional effect on magnesium ions in controlling the polymorph of calcium carbonate crystal. A much wider range of calcium carbonate morphologies was observed in the presence of both collagen and magnesium ions. The experiments suggested that collagen acts in combination with magnesium ions to inhibit calcite crystal growth, while favoring the formation of aragonite crystals. D 2005 Elsevier B.V. All rights reserved. Keywords: Calcium carbonate; Collagen; Magnesium ion; Biomineralization
1. Introduction Many studies have been carried out on the mechanisms involved in biomineralization processes and several new biologically inspired synthetic routes have been designed for control of the formation of the mineral phase. One of the most intensely examined systems is calcium carbonate, which is abundant in biominerals, but also of industrial importance due to its wide use as a filler in paints, plastics, rubber, or paper. It has been shown that the polymorphism, morphology and structural properties of calcium carbonate can be controlled by the use of specific organic templates and/or additives. Langmuir monolayers [1,2], ultrathin organic films [3], self-assembled films [4], have been used as effective templates for the controlled growth of calcium carbonate crystals, focussing on the control of the polymorph and crystal orientation. Crosslinked gelatin films [5], polymer substrates [6], and crystal-imprinted polymer surfaces [7] have also recently been used to direct the
* Corresponding author. Tel.: +86 10 62782770; fax: +86 10 62771160. E-mail address: [email protected] (Q. Feng). 0928-4931/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.msec.2005.08.038
controlled growth of calcium carbonate crystals. Calcium carbonate films have been successfully prepared in the presence of both organic substrates and soluble polymeric additives [8,9]. Interestingly, a designed peptide has been synthesized and used for the conformation-dependent control of the calcite morphology [10]. Proteins isolated from mollusk shells [11,12] and intracrystalline macromolecules from sea urchin spines [13] have shown distinct control on the polymorph of calcium carbonate crystals. The modulation of collagen on crystal morphology of calcium carbonate has been studied in vitro [14]. Manoli induced calcium carbonate crystals by chitin and elastin and obtained calcite crystals in both cases [15,16]. Investigations show that proteins can modulate calcium carbonate morphology and also the polymorph [17 – 19]. Magnesium is known to exert a significant effect on calcium carbonate precipitation and, when present in sufficient concentration, generally results in the precipitation of aragonite rather than the thermodynamically favoured phase, calcite [20 – 24]. Recently, many groups have studied the role of magnesium in calcium carbonate precipitation. Meldrum studied the role of magnesium in stabilising
Y. Jiao et al. / Materials Science and Engineering C 26 (2006) 648 – 652
amorphous calcium carbonate and controlling calcite morphologies [25]. Dawe studied the influence of magnesium on the kinetics of calcite precipitation and calcite crystal morphology [26]. Many studies on the combined effect of organic additives and magnesium ions on calcium carbonate crystal growth have been reported recently [27 – 29], but little research has been carried out on the cooperative influence of protein and magnesium ions on calcium carbonate precipitation. This paper mainly discusses the cooperative influence of collagen and magnesium ions on calcium carbonate precipitation. The aim of these experiments is to find the effect on the polymorph and morphology of calcium carbonate by combined collagen and magnesium ions.
2.1. Preparation of collagen Acid-soluble collagen type I was obtained from rat-tail tendon using a protocol identical to that described by Pins et al. [30]. The rat-tail tendon was dissolved in 10mM HCl at room temperature for 7 h. The solution was centrifuged at 15,000 rpm at 4 -C for 45 min, and then filtered. 0.7 M NaCl was added to induce precipitation, then the precipitate was collected by centrifugation (15,000 rpm, 4 -C, 30 min) and redissolved in 10 mM HCl. Acid-soluble mixture was gathered by dialysis against an aqueous phosphate buffer (20 mM disodium hydrogen phosphate, pH 7.4) at 4 -C for 12 h. The precipitated collagen was collected by centrifugation (15,000 rpm, 4 -C, 45 min), and then dissolved into a concentrated solution by dialyzing the pellets against a large volume of 10 mM HCl at room temperature for 12 h. The collagen solution was stored at 4 -C. 2.2. Method of synthesizing crystals In our experiment, the method of synthesizing crystals was according to the report of Aizenberg et al. [31]. The crystals were grown by slow diffusion (about 7 days) of NH4HCO3 vapor into cell-culture dishes containing magnesium and calcium chloride solutions mixed with collagen solutions at room temperature in a closed desiccator. The amounts of Ca and Mg and the Mg:Ca ratio in the initial solutions are given in Table 1. The solutions were combined in a beaker and were stirred for 2 min before placing glass cover slips on the bottom of the cell-culture dishes. The stirring was stopped and the solution put in a closed dessiccator for crystal precipitation. After crystalli-
Table 1 Composition of precipitation solutions 2
CaCl (mol) MgCl2 (mol)
zation the cover glass-slips were dried in an oven at 40 -C for 12 h. 2.3. Characterization of synthesized crystals X-ray diffraction (XRD),using a D/max-RB X-ray diffractometer with 40 keV CuKa radiation, was employed to analyze the crystal structure and orientation. The morphologies of the crystals were observed under a Hitachi S-450 scanning electron microscope (SEM) after the crystal had grown for 7 days and deposited with gold by sputtering. 3. Results and discussion 3.1. The influence of collagen and magnesium on polymorph of CaCO3
2. Experimental procedure
Mg : Ca
649
0:1
1:1
2:1
3:1
4:1
5:1
0.02 0
0.02 0.02
0.02 0.04
0.02 0.06
0.02 0.08
0.02 0.1
From XRD it can be seen that the polymorph of CaCO3 changed with increasing the concentration of magnesium in the solution containing collagen. When the ratio of Mg/Ca is 0 or 1 only calcite crystals are formed (Fig. 1a), and when the ratio of Mg/Ca is 2 some vaterite and aragonite crystals are precipitated besides calcite (Fig. 1b). The proportion of vaterite and aragonite crystals increased with the increased concentration of magnesium, and when the ratio of Mg/Ca attain 4 most crystals are aragonite, only a few are calcite crystals (Fig. 1c.). While nearly all crystals are aragonite when the ratio of Mg/Ca is 5 (Fig. 1d.). When there is no collagen present, only at ratio of Mg/Ca of 4 or above that aragonite can be seen (Fig. 1e), in other instance only calcite can be seen, which is in agreement with conventional theoretical results. Magnesium is known to induce aragonite formation from sea water and in vitro at ratio of Mg / Ca equal to or greater than 4, while at lower Mg / Ca ratio mostly calcite and magnesian calcite are formed [32]. When collagen is present alone the growth of CaCO3 crystals is greatly inhibited, but there is no obvious change in CaCO3 polymorph and only calcite crystals are formed [14]. The above results show that when collagen and magnesium are both present, aragonite crystals precipitated at a lower Mg / Ca ratio of 2. This indicates that collagen has a promotional effect on magnesium ions in controlling the polymorph of CaCO3 crystals. The reason of this kinetic phenomenon is considered as contribute to two aspects: on the one hand, magnesium ions inhibit the growth of calcite. The partially dehydrated magnesium ions attach to the surface of the nascent calcite nucleus, the strongly bound residual hydration sphere poisons the surface [33], and inhibits subsequent growth. At the same time, the collagen acts with magnesium and increase the magnesium hydrate absorbed in calcite nucleus. On the other hand, owing to the polymorph of calcium carbonate is relate to its energy state [34], magnesium is likely also to react with collagen and to change the stereochemical structure of collagen molecules, and thus induces the aragonite or vaterite with higher energy in high energy state structure.
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Fig. 1. XRD data for calcium carbonate crystals modulated by collagen and magnesium ions (C: calcite, V: vaterite, A: aragonite the concentration of collagen is 0.1 g/l, calcium ion is 0.02 mol/l). (a) Calcite crystals at Mg/Ca:0 or 1. (b) Calcite crystals with some vaterite and aragonite at Mg/Ca:2. (c) Aragonite crystals with a few calcite at Mg/Ca:4. (d) Aragonite crystals at Mg/Ca:5 (e) Aragonite crystals form besides calcite at Mg/Ca:5 when no collagen present.
3.2. The influence of collagen and magnesium on morphology of CaCO3 A much wide range of calcium carbonate morphologies is generated in the presence of both collagen and magnesium ions. When the concentration of protein is 0.1 g/l. The crystals grow in absence of magnesium are irregular rhombohedral calcite with little distortion, about 120 um in size (Fig. 2a). At low magnesium concentration (Mg / Ca ratio is 1), irregular lumpish calcite crystals are formed with lamellar growth structure (Fig.
2b). By increasing the concentration of magnesium, discoid and dumbbell crystals are obtained (Fig. 2c, d, e). When the concentration of magnesium is high enough (Mg / Ca ratio attains to 5), spherical aragonite crystals are precipitated, about 150 um in size (Fig. 2f). Keeping magnesium concentration unchanged but increasing the concentration of collagen (from 0.1 g/l to 0.4 g/l) spherical aragonite crystals with more regular shape can be obtained(Fig. 2g). In contrast to this, when there is no collagen in the solution we get clusters of aragonite crystals with needle shape at higher magnesium concentration (Fig. 2h).
Y. Jiao et al. / Materials Science and Engineering C 26 (2006) 648 – 652
651
Fig. 2. SEM morphologies of the calcium carbonate crystals precipitated in the solution, collagen concentration: 0.1 g/l. (a) Irregular rhombohedral calcite crystal grown in the solution without magnesium. (b) Irregular lumpish crystals with lamellar growth structure (Mg/Ca:1). (c, d, e) Discoid and dumbbell calcium carbonate crystals. (f) Spherical aragonite crystals at higher magnesium concentration (Mg/Ca:5). (g) Spherical aragonite crystals with more regular shape (Mg/Ca:5, collagen concentration:0.4 g/l). (h) Aragonite crystals with needle shape without collagen (Mg/Ca:5).
Above results showed that in the presence of collagen irregular rhombohedral calcite with lamellar structure can be obtained. When magnesium is added, the morphology of crystals changed greatly, from lumpish to dumbbell, and spherical. The morphological changes of calcium carbonate crystals reveal that in the presence of both collagen and magnesium ions, we tend to obtain spherical aragnite spherulites instead of other forms, especially when the concentration of magnesium is high. Through controlling the concentration of collagen and magnesium, spherical crystals with more regular shapes could be obtained.
itation. The results indicated that collagen has a promotional effect on magnesium ions in controlling the polymorph of CaCO3 crystals. By cooperation of collagen and magnesium ions in different concentration, crystals showed a sequence of morphology changes and especially, aragonite crystals with regular spherical morphology were precipitated at high Mg/Ca ion concentration ratio. The experiments suggested that collagen acts in combination with magnesium ions to inhibit calcite crystal growth, while favor the formation of aragonite crystals.
4. Conclusion
Acknowledgements
The present paper mainly studied the co-effect of collagen and magnesium ions on calcium carbonate precip-
This work was supported by the National Natural Science Foundation of China, Grant No.50272035.
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