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Homogeneous Precipitation by Enzyme-Catalyzed Reactions. 2. Strontium and Barium Carbonates† Ivan Sondi‡ and Egon Matijevic´* Center for Advanced Materials Processing, Clarkson University, Potsdam, New York 13699-5814 Received August 20, 2002. Revised Manuscript Received December 6, 2002
Catalytic decomposition of urea by enzyme urease in aqueous strontium and barium chloride solutions was used to rapidly precipitate witherite (BaCO3) and strontianite (SrCO3) particles at room temperature. At the early stages of the process, uniform spheroidal particles were generated, which were shown to consist of nanosized subunits. On continuous aging of the same systems the additionally precipitated alkali earth carbonates grow as whiskers onto original core particles, which eventually fully transform into crystalline rodlike clusters.
Introduction Carbonates occur widely as the main mineral components in rocks and sediments and as inorganic components in exoskeletons and tissues of many mineralizing organisms. Therefore, processes leading to the formation of these compounds have been of considerable interest in geo- and biosciences, as well as in materials. In particular, the chemical and biological precipitations of calcium carbonate polymorphs have been studied in some detail.1-7 However, fewer investigations have dealt with other alkaline earth carbonates.8-12 Recently, it was described how different calcium carbonate precipitates could be obtained in solutions of calcium salts by catalytic decomposition of urea at 90 °C13 and by enzyme-catalyzed decomposition of urea by urease at lower temperatures.14,15 Our previous study has shown that this enzyme can yield various calcium carbonate polymorphs and exert significant influence * To whom correspondence should be addressed. † Part 1. Reference 14. ‡ On leave of absence from the Rudjer Bos ˇ kovic´ Institute, Zagreb, Croatia. (1) Weiner, S.; Addadi, L. J. Mater. Chem. 1977, 7, 689. (2) Albeck, S.; Addadi, L.; Weiner, S. Connect. Tissue Res. 1996, 35, 419. (3) Xu, G.; Yao, N.; Aksay, I. A.; Groves, J. T. J. Am. Chem. Soc. 1998, 120, 11977. (4) Mann, S.; Heywood, B. R.; Rajam, S.; Birchall, J. D. Nature 1988, 334, 692. (5) D’Souza, S. M.; Alexander, C.; Carr, S. W.; Waller, A. M.; Whitcombe, M. J.; Vulfson, E. N. Nature 1999, 398, 312. (6) Kralj, D.; Brecˇevic´, Lj.; Nielsen, E. J. Cryst. Growth 1990, 104, 793. (7) Brecˇevic´, Lj. J. Chem. Soc., Faraday Trans. 1996, 92, 1017. (8) Lange, R.; Bergbauer, M.; Szewzyk, U.; Reitner, J. Facies 2001, 45, 195. (9) Owusu, G.; Litz, J. E. Hydrometallurgy 2000, 57, 23. (10) Ferris, F. G.; Fratton, C. M.; Gertis, J. P.; Schultzelam, S.; Lollar, B. S. Geomicrobiol. J. 1995, 13, 57. (11) Chen, J. F.; Wang, Y. H.; Guo, F.; Wang, X. M.; Zheng, C. Ind. Eng. Chem. Res. 2000, 39, 948. (12) Sizov, P. V.; Sizova, S. A.; Rovnyi, S. I. Radiochemistry 1998, 40, 265. (13) Wang, L.; Sondi, I.; Matijevic´, E. J. Colloid Interface Sci. 1999, 218, 545. (14) Sondi, I.; Matijevic´, E. J. Colloid Interface Sci. 2001, 238, 208. (15) Bachmeier, K. L.; Williams, A. M.; Warmington, J. R.; Bang, S. S. J. Biotechnol. 2002, 93, 171.
on the development of calcite in different habits.14 Catalytic decomposition of urea by urease has also been used in the preparation of aluminum hydroxide, aluminum basic sulfate,16-18 hydrotalcite,19 hydroxyapatite precursors,20 and oxide fuel cell particles.21 It is well-known that in natural environments urea is a common product of biodegradation of organic substances, while urease occurs in a number of plants, in several species of yeasts, and in many bacteria. Thus, the role of urease, generated by the latter, plays a significant role in the formation of intracellular crystals,22-24 in the microbial precipitation of calcite in remediation of the surface and subsurface porous media,25 and in cracks of granite and concrete.26 The present investigation deals with the influence of urease on the precipitation of anhydrous carbonates from the aragonite group, that is, witherite (BaCO3) and strontianite (SrCO3), in solutions containing strontium and barium chloride and urea at room temperature. The aim of the study was to determine the effects of this enzyme on the morphological and structural characteristics of the resulting solids, with special emphasis on different phases in the growth process. Experimental Section Materials. Stock solutions of reagent-grade SrCl2, BaCl2, and urea were freshly prepared and filtered through 0.22-µm Millipore membranes before use. (16) Unuma, H.; Kato, S.; Ota, T.; Takahashi, M. Adv. Powder Technol. 1988, 9, 181. (17) Kara, F.; Sahin, G. J. Eur. Ceram. Soc. 2000, 20, 689. (18) Simpson, R. E.; Habeger, C.; Rabinovich, A.; Adair, J. H. J. Am. Ceram. Soc. 1998, 81, 1377. (19) Ogawa, M.; Kaiho, H. Langmuir 2002, 18, 4240. (20) Bayraktar, D.; Tas, A. C. J. Mater. Sci. Lett. 2001, 20, 401. (21) Tas, A. C.; Majewski, P. J.; Aldinger, F. J. Am. Ceram. Soc. 2002, 85, 1414. (22) Hesse, A.; Heimbach, D. World J. Urol. 1999, 17, 308. (23) Edinliljegren, A.; Grenabo, L.; Hedelin, H.; Pettersson, S.; Wang, Y. H. J. Urol. 1994, 152, 208. (24) Grover, J. E.; Rope, A. F.; Kaneshiro, E. S. J. Eukaryot. Microbiol. 1997, 44, 366. (25) Stocks-Fischer, S.; Galinat, J. K.; Bang, S. S. Soil Biol. Biochem. 1999, 31, 1563. (26) Gollapudi, U. K.; Knutson, C. L.; Bang, S. S.; Islam, M. R. Chemosphere 1995, 30, 695.
10.1021/cm020852t CCC: $25.00 © 2003 American Chemical Society Published on Web 02/28/2003
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Figure 1. Scanning electron micrographs (SEM) of strontium (a) and barium (b) carbonate particles obtained by enzyme-catalyzed decomposition of urea in aqueous solutions of strontium and barium salts at room temperature after 2 min of reaction time (Table 1, samples 3 and 7), and of strontium (c) and barium (d) carbonates, obtained by aging the same solutions at 90 °C for 2 h in the absence of urease (Table 1, samples 1 and 5). Table 1. Composition of Solutions for the Precipitation of Strontium and Barium Carbonates sample 1a 2 3 4 5a 6 7 8 a
[SrCl2] mol dm-3
[BaCl2] mol dm-3
[urea] mol dm-3
0.2 0.2 0.2 0.2
0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5
0.2 0.2 0.2 0.2
[urease] mg cm-3 0.1 0.5 1 0.1 0.5 1
Systems aged at 90 °C; all others aged at 25 °C.
Enzyme urease (lot 31D4773, molecular weight of 480 000 Da, Worthington Biochemical Corporation) was fractionated from crude Jack Beans (Canavlia ensiformis) meal extract, with activity of 71 units/mg of dry weight. Preparation of the Particles. All experiments were carried out at room temperature under conditions given in Table 1. The reacting solutions containing strontium and barium salts and urea were degassed by nitrogen bubbling before urease was injected and then kept at room temperature from 2 min to 2 h under gentle stirring. The pH of the solutions changed from 6.3 at the beginning to 7.9 after 2 h of the precipitation process. Characterization of the Particles. The size and the morphology of the particles were examined by scanning
electron microscopy (SEM) and by the Voyager X-ray Microanalysis and Digital Imaging System, while their structure was evaluated by X-ray diffraction (XRD). The crystalline phase of witherite (BaCO3) and strontianite (SrCO3) type particles were identified according to the Powder Diffraction File PDF-2 (International Center for Diffraction Data). The average crystallite size was calculated from the line broadening of the XRD patterns.27
Results Scanning electron micrographs in Figure 1 demonstrate the effect of different methods used in the precipitation of finely dispersed strontianite and witherite. Under otherwise the same experimental conditions, reasonably uniform, spheroidal SrCO3 and BaCO3 particles were rapidly formed by the urease-catalyzed decomposition of urea at room temperature in only 2 min (Figure 1a,b; Table 1, samples 3 and 7), while needlelike crystals were obtained by thermal decomposition of urea at 90 °C after 2 h of aging (Figure 1c,d; Table 1, samples 1 and 5). In the latter case no solid phase appears at shorter reaction times. It is evident that at much higher magnification the precipitates of SrCO3, prepared by the enzymatic reac(27) Cullity, B. D. Elements of X-ray Diffraction; Addison-Wesley: Reading, MA, 1978; p 102.
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Figure 4. XRD patterns (a) of the strontium carbonate precipitate obtained by aging a solution containing 0.5 mol dm-3 urea and 0.2 mol dm-3 SrCl2 at 90 °C for 2 h (Table 1, sample 1) and (b) of barium carbonate particles obtained by aging a solution containing 0.5 mol dm-3 urea and 0.2 mol dm-3 BaCl2 at 90 °C for 2 h (Table 1, sample 5).
Figure 2. SEM of the same strontium carbonate particles shown in Figure 1a at higher magnifications to display their internally composite nature.
istic of strontianite. The size of the nanosized subunits evaluated by the Scherrer equation is ∼20 nm. In contrast, needlelike particles of both carbonates (Figure 1c,d) displayed no substructure, and the XRD patterns indicate their high crystallinity (Figure 4). Particles shown in Figure 1, removed from the mother liquor, remained indefinitely stable and could be redispersed in water without undergoing any changes. However, if the reaction proceeded for longer periods of time, their natures changed in a profound manner. Figure 5 demonstrates the evolution of the SrCO3 particles over 2 h of aging in the same system, the composition of which is that of sample 3 in Table 1. This process is accompanied by the additional formation of the solid phase, as shown in Figure 6. The final rodlike particles exhibit enhanced crystallinity, indicated by the XRD pattern in Figure 3c. Since the same effects were observed with BaCO3, there was no need to reproduce essentially the same kind of SEM and XRD data. It is noteworthy that changing the concentration of urease in the reacting systems did not affect the crystallinity, morphology, or the growth pattern of the two carbonates. However, the amount of the added enzyme influenced the rate of the solid phase formation (Figures 6 and 7). However, the yields of precipitated SrCO3 and BaCO3, obtained at each concentration of urease, were reasonably close. Discussion
Figure 3. XRD patterns of strontium carbonate precipitates obtained in a solution containing 0.5 mol dm-3 urea, 0.2 mol dm-3 SrCl2, and 0.5 mg cm-3 urease (Table 1. sample 3) aged at 25 °C for (a) 2 min. (Figures 1a and 2), (b) 60 min, and (c) 120 min (Figure 5b,c).
tion, consist of nanosized subunits (Figure 2a,b). The XRD pattern of this solid (Figure 3a), taken shortly after the separation from the reacting solution, is character-
There are two major effects of urease on the formation of alkaline earth carbonates in the presence of urea. The first refers to the rapid initial rate of the solid phase formation at room temperature and the second to the evolution of different morphologies of the dispersed matter on continued reaction. However, there are distinct differences in the natures of the dispersed products with different reacting cations. Thus, it was previously shown14 that the initially precipitated calcium carbonate appeared as an amorphous gel, which on further aging changed to vaterite and calcite crystals of several micrometers in size and of different shapes.
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Figure 6. Change of the weight and of the yield of strontium carbonate with time, precipitated in solutions containing 0.5 mol dm-3 urea, 0.2 mol dm-3 SrCl2, and (O) 10, (4) 50, and (0) 100 mg cm-3 urease.
Figure 7. Change of the weight and the yield of barium carbonate with time, precipitated in solutions containing 0.5 mol dm-3 urea, 0.2 mol dm-3 SrCl2, and (O) 10, (4) 50, and (0) 100 mg cm-3 urease.
Figure 5. SEM of strontium carbonate precipitates obtained in a solution containing 0.5 mol dm-3 urea, 0.2 mol dm-3 SrCl2, and 0.5 mg cm-3 urease aged at 25 °C (Table 1, sample 3) for (a) 2, (b) 60, and (c) 120 min.
urease. However, on prolonged aging the original spheroids grow into needlelike clusters. While the mechanism of the involved process is not clear, it would seem that initially nanosized crystals of the two carbonates are formed, which rapidly aggregate to uniform primary particles, stabilized by the protein. This kind of aggregation process has now been established as a rather common mechanism in the formation of larger, colloidal particles of different chemical compositions including pure metals28 and numerous inorganic compounds.29-31 A model has been developed which explains the formation of uniform final particles by such aggregation processes.32,33 Of specific interest in the systems described in this study is the second stage in which the original composite particles, on aging in the same mother liquor, convert into needles, typical of these carbonates. Figures 6 and 7 show that the amounts of precipitates continue to increase with time, which means that
In contrast, the first products in solutions of strontium and barium salts consist of rather uniform spheroidal particles of distinct crystallinity, despite the fact that they are aggregates of nanosize subunits. This finding differs from the normal expectations, because as a rule BaCO3 and SrCO3 appear as elongated rods, illustrated in Figure 1c,d, when precipitated in the absence of
(28) Goia, D. V.; Matijevic´, E. Colloids Surf. 1999, 146, 139. (29) Hsu, W. P.; Ronnquist, L.; Matijevic´, E. Langmuir 1988, 4, 31. (30) Ocan˜a, M.; Matijevic´, E. J. Mater. Res. 1990, 5, 1083. (31) Matijevic´, E. Chem. Mater. 1993, 5, 412. (32) Privman, V.; Goia, D. V.; Park, J.; Matijevic´, E. J. Colloid Interface Sci. 1999, 213, 36. (33) Park, J.; Privman, V.; Matijevic´, E. J. Phys. Chem. B 2001, 105, 1163.
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additional carbonates are generated in the respective systems. However, instead of forming new spheroidal particles, these solids grow as whiskers on the existing cores, which simultaneously undergo internal restructuring. It would seem that the free energy of the latter systems is less than what it would be, if more of the nanosized precursors were formed. It is quite possible that the subunits in the primary particles act as nucleating agents for the subsequent growth of the needles.
Sondi and Matijevic´
The results of this study are indicative of the processes involved in the biomineralization of alkaline earth anhydrous carbonates of the aragonite group, commonly taking place in natural environments. Acknowledgment. This research has been supported in part by NSF Grant DMR-0102644. The authors gratefully acknowledge Dr. Branka SalopekSondi for conducting some of the reported experiments. CM020852T
