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The effect of anionic polyelectrolytes on the crystallization of calcium oxalate hydrates
Journal of Crystal Growth 100 (1990) 627—634 North-Holland
627
THE EFFECT OF ANIONIC POLYELECTROLYTES ON THE CRYSTALLIZATION OF CALCIUM OXALATE HYDRATES Joseph S. MANNE Department of Urology, Long Island Jewish Medical Center, New Hyde Park, New York 11042, USA
Naresh BIALA Department of Chemical Engineering and Applied Chemistry. Columbia University, New York, New York 10027, USA
Arthur D. SMITH Department of Urology, Long Island Jewish Medical Center, New Hyde Park, New York, 11042, USA
and Carl C. GRYTE Department of Chemical Engineering and Applied Chemistry, Columbia University, New York, New York 10027, USA
Received 27 April 1989
Calcium oxalate was precipitated from supersaturated calcium oxalate solutions in the presence of different water soluble polymers: poly(acrylic acid), poly(methacrylic acid), poly(styrene sulfonate-alt-maleic anhydride), poly(maleic anhydride), and 3M and poly(vinyl sulfonate). The supersaturated solutions were made with initial calcium and oxalate concentrations of 4 x 10 1.5 x 10 3M respectively. In the presence of anionic polyelectrolytes there was a change in the hydrate form of calcium oxalate which precipitated. With low concentrations of polyelectrolyte (up to 1 ppm) only the monohydrate was formed, at intermediate concentration (1—50 ppm) trihydrate was the predominant product, and at high concentrations (above 50 ppm) the dihydráte form was the dominant product. Nonpolyelectrolytes had no effect on the crystallization and thus only calcium oxalate monohydrate was formed. This effect was found to occur with all polyelectrolytes used regardless of molecular weight or charge. The change from one hydrate from to another occurred at a lower polymer concentration with the low molecular weight polymers.
1. Introduction There is extensive evidence of the ability of polymers to influence calcium salt crystallization and growth. McCartney and Alexander [1] have reported that calcium sulfate crystal growth is inhibited by certain polymers including poly(acrylic acid), poly(methacrylic acid), carboxymethylcellulose, and poly(ethylene oxide). Crawford et al. [2] have reported that the effectiveness of calcium crystallization inhibition by a polymer is inversely proportional to its molecular weight. Similar results have been found with the inhibition of calcium carbonate crystals by poly(methacry0022-0248/90/$03.50 © Elsevier Science Publishers B.V. (North-Holland)
lates) [3]. In an study by Crawford and Smith [4] the inhibition of calcium sulfate crystal nucleation and growth was accomplished with poly(acrylic acid). It is believed by the authors that this is mediated by adsorption of the polyelectrolyte on the surface. The crystals produced in the presence of the poly(acrylic acid) were smaller and more numerous than those produced without the polymer present. Polyelectrolytes are used to inhibit and sometimes to remove phosphate, sulfate and carbonate scales from heat transfer surfaces during power generation. The formation of bone, teeth and marine exoskeletons involve deposition of calcium hydroxyapatite (phosphate) in the pres-
628
J. S. Manne et at.
/
Effect ofanionic polyelectrolytes on crystallization ofcalcium oxalate hydrates
ence of collagen like macromolecules. Dental plaque, a biological but similarly unwanted scale, involves calcium precipitation in the presence of colloidal materials in the dental cavity. In each case crystallization occurs on a surface on which significant concentrations of polymeric macromolecules have adsorbed. Urine is frequently supersaturated with calcium oxalate. Both stone-formers and normal individuals have been noted to have calcium crystals in their urine by Smith and co-workers [5]. There is extensive experimental evidence that the precipitation of calcium oxalate is affected by polymeric and nonpolymeric compounds [6]. These polymers are believed to contribute to the formation of a matrix for deposition of calcium oxalate crystals [7—11]. They can also alter the growth of the crystals. RNA, hepann, pyrophosphates, polyphosphates and Tamm—Horsfall mucoproteins are examples of such molecules [12,13]. In a recent study by Smith and co-workers [14], supersaturated calcium oxalate was precipitated in the presence of certain natural polymers including chondroitin sulphate, pyrophosphate, RNA, and heparin. The percentage of calcium oxalate dihydrate increased with increasing concentration of polymer. In this work we demonstrate that this change in hydrate form with polymer concentration is general property of a wide range of anionic polyelectrolytes. 2. Experimental 2.1. Polymeric materials
Poly(acrylic acid) was obtained in a low molecular weight (PAAL, K759, Goodrich, Mn 5000) and high molecular weight (PMAAH, synthesis in benzene using AIBN initiator at 60°C, Mn, 250,000) form. Poly(methacrylic acid) was also obtained as a low (PMAA, LP 30 Dearborn Chemical, Mn 5000) and a high (PMAAH, synthesis similar to PAAH, Mn 550,000) molecular weight polymer. An alternating copolymer of styrene sulfonate and maleic anhydride (PSS-MA) was obtained from ARCO chemical. Poly(vinyl sulfonate) (PVS, Air Products, Mn 2000) was used as received. =
=
2.2. Preparation of calcium oxalate crystals
Calcium oxalate crystals were formed from supersaturated solutions at 210 C. Stock solutions of ammonium oxalate (0.15M), calcium chloride (0.4M) and various polymers (1000 ppm) were prepared in doubly distilled water. The polymer solution was mixed with 10 cm3 of 0.15M ammonium oxalate. This was placed in a thermostat equipped with a magnetic stirrer and diluted with distilled water to 990 cm3. To this solution 10 cm3 of 0.4M calcium chloride was added with constant mixing. The initial concentrations of calcium and oxalate were 4 x 103M and 1.5 X 103M, respectively. A white precipitate of calcium oxalate was observed to form and slowly settle out of the solution. Crystals were recovered by pressure filtration using a 0.45 ~tm microporous filter. The pH measured before and after the crystallization was 6.5 ±0.3. The samples were then air-dried and stored in vials for further study. 2.3. Preparation of calcium oxalate hydrates
In order to calibrate the X-ray method of analysis of the different calcium oxalate hydrate forms, it was necessary to prepare separately each of the forms. Calcium oxalate monohydrate or whewellite (COM) was easily prepared under the conditions in section provided that theoxalate polymeric given component was 2.2, excluded. Calcium dihydrate or weddelite (COD) forms spontaneously in urine and Gardner [15] showed how it could be prepared from synthetic urine. Following their methods, we added to 500 cm3 of freshly voided urine, 25 cm3 of ammonium oxalate (0.25M) with constant stirring. The precipitate formed was filtered and dried and found by X-rays to be exclusively dihydrate. There are different ways to make COT crystals [16]. In this work, COT was exclusively formed when calcium oxalate precipitated as described in section 2.2 in the presence of 5 ppm PAAL.
=
2.4. X-ray diffraction standards for calcium oxalate hydrates
Mechanical mixtures of COM—COT and COD—COT were prepared and their powder X-ray
J.S. Manne et at.
/ Effect of anionic polyelectrolytes on
crystallization ofcalcium oxalate hydrates
629
therms are presented in terms of the oxalate to calcium ratio as a function of the oxalate con-
1.20
respect to calcium oxalate. Crawford, Crematy and Alexander [22] report average urinary concentrations of calcium and oxalate to be respeccentration. and 1.78 xsupersaturated 104M. Thus, with the lively 4.0 x Urine 103Mis ordinarily concentration product in urine is about 280 times larges than that anticipated from the solubility product of COM. In this same source they give the value of calcium oxalate saturation as 5.12 X iO~. Typical supersaturation shown in fig. 2.by sis, the urine oxalate concentrationis was increased In order to obtain sufficient crystals for analy-
0 0.80
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0.40
/X~
0.00 0.00
0.20
I
I
0.40
0.60
0.80
1
1.00
MASS FRACTION Fig. 1. Calibration of X-ray powder patterns. (A) Mass fraction
of COM, in a mixture of COM (29 = 14.9) and COT (29 16.1), versus the area fraction of the COM peak of the total peak areas. (•) Mass fraction versus peak area fraction for COD (20=14.3)inaCOD—COTmixture.
patterns were measured using CaKct X-rays. COM, COD and COT were identified using the published data from the Joint Committee on Powder Diffraction Standards 1971 (COT and COM [17]; COD [18]). Analysis of the mixtures was determined from peak areas for COM at 29 14.93 and 15.29, for COD at 29 14.32 and for COT at 29 13.22 and 16.1. The calibrations of peak area fraction as a function of mixture mass fraction given in fig. I were used to determine the fraction of the various hydrate forms in the different samples of precipitated calcium oxalate.
about one order of magnitude over that commonly found in urine. In fig. 2, the initial concentration, and the trajectory of solution concentration change during crystallization are given. Although most of the crystal mass is formed at oxalate concentrations greater than those found in urine, the composition trajectory of the present system passes close and parallel to that applicable to ordinary urine. The hydrate form of calcium oxalate which precipitated from solution was a strong function
—
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2.5. Scanning electron microscopy
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Dry calcium oxalate crystals were mounted on conductive adhesive, shadowed with a thin coating of gold and observed by SEM.
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3. Results and discussion
OXALATE Fig. 2. Oxalate to calcium ratio versus oxalate concentration.
The solubility products for COM, COT and COD at 37°C are reported to be respectively (in moles2/liter2): 2.47 X iO~, 10.82 x i0~ and 6.09 x IO~[19—21].In fig. 2, these solubility iso-
~
for COM (A), COT (B) and COD (C) is 2.47 x i0~, 10.82x iO~ and 6.09x iO~ moles2/liter2, respectively, at
37°C [19—211.Initial solution conditions (D) and (— the solution trajectory during crystallization and typical concentrations for urine (E) as shown.
J.S. Manne et at. / Effect of anionic polyelectrolytes on crystallization of calcium oxalate hydrates
630
0
20
15
25
2e Fig. 3. X-ray diffraction powder pattern for calcium oxalate crystals recovered from precipitation from a solution containing indicated concentrations of PAAL (ppm).
of the concentration of polyelectrolyte in solution. This is well illustrated by the case of PAA(L). The calcium oxalate crystallization in the presence of low concentrations of low molecular weight poly
Table 1 Effects of various polymer types and concentrations Polymer
(acrylic) acid (PAAL) occurred in less than 10 s while at the higher polymer concentrations the precipitation occurred over a period of 5 mm. The crystals were easily filtered and dried. The X-ray powder patterns for oxalate formed in increasing concentrations of PAAL are given in fig. 3. In the absence of PAAL, the peak is characteristic of COM. As concentration of PAAL increases, the X-ray data indicates a mixture of COM and COT while at concentrations above 50 ppm only COD is found. In all cases the X-ray peaks are distinct well separated and suitable for quantitative analysis. In table I, the effects of the various polymer types and concentrations are summarized. In general, regardless of the character of the polyelectrolyte (carboxylate or sulfonate), as polymer concentration increases there is a shift from monohydrate to trihydrate and finally to dihydrate. In fig. 4a the fraction of each hydrate formed for both PAAL and PAAH is given as a function of polymer concentration. Consider the data for PAAL. At concentration below 0.5 ppm PAAL only COM is found. When the concentration is increased to about I ppm polymer there is a sharp shift from COM to COT formation and COT is observed exclusively in the concentration range between 1 and 5 ppm. At concentrations above 50 ppm only COD is observed. These data suggest that the polyacrylic acid exerts a very strong influence on the crystallization process. Although carboxylate polymers do bind calcium in solution (and thus lower calcium activity), the molar concentrations of polymeric carboxylate are generally much lower than those of the calcium. Even at 100
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Concentrations of polymer (in parts per million) 0.1
0.5
1.0
5.0
10
50
100
500
PAAL PAAH
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M M
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T M,T
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D D
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M M
M, T M
M, T M
T, D T
T, D T. D
D T, D
D T, D
D D
M, T M,D,T M,T calcium oxalate
D D T.D trihydrate.
T, D D D
D D D
PMAL M, T M, T PVS M M,T PSSMA M M a) M = calcium oxalate monohydrate; D
T M,T M,T calcium oxalate
T M,T M,T dihydrate; T =
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Effect ofanionic polyelectrolytes on crystallization ofcalcium oxalate hydrates
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Fig. 4. (a) Mass fraction of hydrate forms of calcium oxalate as a function of polymer concentration: for PAAL, 0 is COM, ~D is COT and• is COD; for PAAH, G is COM, 4 is COT and • is COD. (b) Mass fraction of calcium oxalates as a function of the polymer concentration of: PMAAL, ~ is COM, 11 is COT and A 15 COD; for PMAAH, v is COM, ~ is COT and v is COD. (c) Mass fraction of hydrate types as a function of polymer concentration: for PVS, 0 is for COM, W is for COT and• is for COD; for PSSMA, ~ is for COM, II is for COT and A is for COD.
J.S. Manne et at / Effect of anionic polyelectrolytes on crystallization of calcium oxalate hydrates
632
ppm PAAL, the calcium concentration is 3 times higher than the polyelectrolyte fixed charge. In fig. 4a data are also given for hydrate formed in the presence of the high molecular weight poly(acrylic) acid, PAAH. Although the trend is the same as concentration of polymer increases, it appears that
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PAAL shifts the oxalate from COM to COT at a lower concentration than does PAAH. High molecular weight polyelectrolyte should have a stronger effect on the solution activity of divalent ions. If the hydrate control were the result of changes in the solution activity of calcium in the
_______
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I Fig. 5. SEM (Cambridge: Model 200) photographs of sample calcium oxalate crystals produced at different conditions. (a) ~OM crystals, formed in 0.5 ppm PAAL; bar indicates 4 ~m. (b) COT (5.0 ppm PAAL). (C) COD (100 ppm PAAL). (d) COD and COT (50 ppm PAAH). (e) COD (100 ppm PMAA). (1) COD (500 ppm PSSMA).
J. S. Manne et at.
/
Effect of anionic polyelectrolytes on crystallization ofcalcium oxalatehydrates
presence of polyelectrolyte, then at the same mass concentration, the high molecular weight polymer should be more effective than the ow molecular weight material. However the opposite is observed. It appears that polymer adsorption is the most reasonable mechanism for hydrate control. A given mass of strongly adsorbed low molecular weight poly(acrylic acid) contains many more polymer molecules and can more effectively control crystal growth over a crystal surface than can a similarly strongly adsorbed high molecular weight material. The low molecular weight material covers more of the crystal surface. In fig. 4b, similar results are shown for high and low molecular weight poly(methacrylic acid). It appears that PMAAL is more effective than PAAL since the change in the calcium oxalate yield from COM to COT occurs at a lower concentration for PMAAL. The appearance of COD occurs at a polymer concentration that is less dependent on polymer type and molecular weight. In fig. 4c, data are given for two sulfonate containing polyelectrolytes, poly(vinyl sulfonate) and poly(styrene sulfonate-alt-maleic anhydride). Surprisingly, the hydrate formed at the different polymer concentrations is very similar to that observed with the carboxylate containing polymers. Since sulfonates are expected to have less effect on the solutions activity of calcium ions than carboxylates, ion binding in solution seems less important than polymer adsorption on the crystal surface. In figs. 5a—5f, SEM micrographs are presented of the various crystal forms observed in this work. The COM and COT in figs. 5a and Sb are faceted single (sometimes clumped) crystals. They appeared similar in shape regardless of the type of solution from which they were formed. In fig. 5c, the typical bipyramidal form of COD is shown in crystals formed in the presence of 50 ppm PAAH. At 100 ppm PAAL the faceted is seen. In fig. 5e, 100 ppm of PMAAL gives large populations of nonfaceted spherullites (dumbbells). Such shapes are common when viscous polymeric fluid control the crystallization process [23]. In the extreme, fig. 5f clearly shows a residual polymer layer around the spheroidal COD particles. Crystalluria commonly contains both the bipyramidal and the
633
spherullitic form of the COD suggesting that it is urine polyelectrolytes that control crystalluria formation. The mechanism for the phenomena described in this report is intriguing but not well understood. However we can make some strong hypotheses based on our results and of others. This phenomena may be due to kinetic effects. It is known from the work of Nancollas and Tomazic [24] that during the crystallization of calcium oxalate hydrates in the presence of polymer that there is a greater adsorption of polyelectrolyte by COM than COD or COT. They also showed that this greater adsorption of polymer is associated with greater inhibition of both COM nucleation and growth than for COD and COT. In polymerfree solution the rate of growth of COM seeds is greater than COT or COD. However, in the presence of polymer the growth rate of COM is slowed enough so that its relative rate is now less than COD or COT. It is also known that COT seeds grow faster than COD. Hence in the medium polymer concentration ranges it is expected that more COT forms than COD. At higher polymer concentrations (order of magnitude 10—6) the growth rate of COD stabilizes, whereas that of COT continues to decrease. Extrapolation from published data shows that the rate constant will finally drop below that for COT. This would help explain the predominance of COD at high polymer concentrations. It is known that COT and COD are less stable than COM and that at equilibrium the COT and COD will convert to COM. However it appears that in the nonequilibrium state that kinetic factors may predominate and lead to the formation of COT and COD in the presence of polymer.
Acknowledgements This work was supported in part by a grant from Dearborn Chemical: Lake Zurich, Illinois. J.S. Manne gratefully acknowledges the support of the Department of Urology, Long Island Jewish Medical Center, and the NIH (research fellowship F32 DK08280-01), Columbia University.
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Effect ofanionic polyelectrolytes on crystallization of calcium oxalate hydrates
References
[12] W.G. Robertson, Urol. Intern. 41(1986) 329. [13] ME. Sheehan and G.H. Nancollas, J. Urol. 132 (1984)
[1] E.R. McCartney and A.E. Alexander, J. Colloid Sci. 13
(1958) 383. [2] J.E.
[3] [4] [5] [6] [7]
[8] [9) [10]
[11]
Crawford,
E.P.
Crematy and
A.E.
Alexander,
Australian J. Chem. 21(1968)1067. F.V. Williams and R.A. Ruehrwein, J. Am. Chem. Soc. 79 (1957) 4898. J.E. Crawford and BR. SMith, J. Colloid Interface Sci. 21 (1966) 623. P.G. Werness, J.H. Bergert and L.H. Smith, J. Crystal Growth 53 (1966) 1981. N. Garti, S. Sang and F. Tibika, Invest. Urol. 18(2) 149. S.R. Khan, B. Finlayson and R.L. Hackett, in: Scanning Electron Microscopy/1983, Vol. I, Ed. 0. Johari (SEM, AMF O’Hare, IL, 1983) p. 379. B. van ‘t Riet, in: Urolithiasis: Physical Aspects (NatI. Acad. Sci., Washington, DC, 1972). MV. Smith, J. Urol. 107 (1972) 164. S.R. Khan and R.L. Hackett, in: Scanning Electron Microscopy/1984, Vol. I, Ed. 0. Johari (SEM, AMF O’Hare IL, 1984) p. 935. H. Iwata, A. Nishio, A. Wakatsuki, K. Ochi and M. Takeuchi, J. Urol. 133 (1985) 334.
158. [14] X. Martin, L.H. Smith and PG. Werness, Kidney Intern. 25 (1984) 984. [15] G.L. Gardner, J. Phys. Chem. 82(8) (1978) 864. [16] B. Tomazic and OH. Nancollas, J. Crystal Growth 46 (1979) 355. [17] L. Walter-Levy, Compt. Rend. (Paris) 254 (1962) 1073. [18] EL. Prien and C. Frondel, J. Urol. 57 (1947) 949. [19] G.L. Gardner and RH. Doremus, Invest. Urol. 15 (1978) 458. [20] B. Tomazic and G.H. Nancollas, J. Colloid Interface Sci. 75 (1980) 149. [21] G.L. Gardner, J. Colloid Interface Sci. 54 (1976) 298. [22] J.E. Crawford, E.P. Crematy and A.E. Alexander, Australian J. Chem. 21(1968) 406. [23] H.D. Keith and F.J. Padden, J. AppI. Phys. 34 (1963) 2409. [24] B. Tomazic and G.H. Nancollas, J. Colloid Interface Sci. 75 (1980) 149.
627
THE EFFECT OF ANIONIC POLYELECTROLYTES ON THE CRYSTALLIZATION OF CALCIUM OXALATE HYDRATES Joseph S. MANNE Department of Urology, Long Island Jewish Medical Center, New Hyde Park, New York 11042, USA
Naresh BIALA Department of Chemical Engineering and Applied Chemistry. Columbia University, New York, New York 10027, USA
Arthur D. SMITH Department of Urology, Long Island Jewish Medical Center, New Hyde Park, New York, 11042, USA
and Carl C. GRYTE Department of Chemical Engineering and Applied Chemistry, Columbia University, New York, New York 10027, USA
Received 27 April 1989
Calcium oxalate was precipitated from supersaturated calcium oxalate solutions in the presence of different water soluble polymers: poly(acrylic acid), poly(methacrylic acid), poly(styrene sulfonate-alt-maleic anhydride), poly(maleic anhydride), and 3M and poly(vinyl sulfonate). The supersaturated solutions were made with initial calcium and oxalate concentrations of 4 x 10 1.5 x 10 3M respectively. In the presence of anionic polyelectrolytes there was a change in the hydrate form of calcium oxalate which precipitated. With low concentrations of polyelectrolyte (up to 1 ppm) only the monohydrate was formed, at intermediate concentration (1—50 ppm) trihydrate was the predominant product, and at high concentrations (above 50 ppm) the dihydráte form was the dominant product. Nonpolyelectrolytes had no effect on the crystallization and thus only calcium oxalate monohydrate was formed. This effect was found to occur with all polyelectrolytes used regardless of molecular weight or charge. The change from one hydrate from to another occurred at a lower polymer concentration with the low molecular weight polymers.
1. Introduction There is extensive evidence of the ability of polymers to influence calcium salt crystallization and growth. McCartney and Alexander [1] have reported that calcium sulfate crystal growth is inhibited by certain polymers including poly(acrylic acid), poly(methacrylic acid), carboxymethylcellulose, and poly(ethylene oxide). Crawford et al. [2] have reported that the effectiveness of calcium crystallization inhibition by a polymer is inversely proportional to its molecular weight. Similar results have been found with the inhibition of calcium carbonate crystals by poly(methacry0022-0248/90/$03.50 © Elsevier Science Publishers B.V. (North-Holland)
lates) [3]. In an study by Crawford and Smith [4] the inhibition of calcium sulfate crystal nucleation and growth was accomplished with poly(acrylic acid). It is believed by the authors that this is mediated by adsorption of the polyelectrolyte on the surface. The crystals produced in the presence of the poly(acrylic acid) were smaller and more numerous than those produced without the polymer present. Polyelectrolytes are used to inhibit and sometimes to remove phosphate, sulfate and carbonate scales from heat transfer surfaces during power generation. The formation of bone, teeth and marine exoskeletons involve deposition of calcium hydroxyapatite (phosphate) in the pres-
628
J. S. Manne et at.
/
Effect ofanionic polyelectrolytes on crystallization ofcalcium oxalate hydrates
ence of collagen like macromolecules. Dental plaque, a biological but similarly unwanted scale, involves calcium precipitation in the presence of colloidal materials in the dental cavity. In each case crystallization occurs on a surface on which significant concentrations of polymeric macromolecules have adsorbed. Urine is frequently supersaturated with calcium oxalate. Both stone-formers and normal individuals have been noted to have calcium crystals in their urine by Smith and co-workers [5]. There is extensive experimental evidence that the precipitation of calcium oxalate is affected by polymeric and nonpolymeric compounds [6]. These polymers are believed to contribute to the formation of a matrix for deposition of calcium oxalate crystals [7—11]. They can also alter the growth of the crystals. RNA, hepann, pyrophosphates, polyphosphates and Tamm—Horsfall mucoproteins are examples of such molecules [12,13]. In a recent study by Smith and co-workers [14], supersaturated calcium oxalate was precipitated in the presence of certain natural polymers including chondroitin sulphate, pyrophosphate, RNA, and heparin. The percentage of calcium oxalate dihydrate increased with increasing concentration of polymer. In this work we demonstrate that this change in hydrate form with polymer concentration is general property of a wide range of anionic polyelectrolytes. 2. Experimental 2.1. Polymeric materials
Poly(acrylic acid) was obtained in a low molecular weight (PAAL, K759, Goodrich, Mn 5000) and high molecular weight (PMAAH, synthesis in benzene using AIBN initiator at 60°C, Mn, 250,000) form. Poly(methacrylic acid) was also obtained as a low (PMAA, LP 30 Dearborn Chemical, Mn 5000) and a high (PMAAH, synthesis similar to PAAH, Mn 550,000) molecular weight polymer. An alternating copolymer of styrene sulfonate and maleic anhydride (PSS-MA) was obtained from ARCO chemical. Poly(vinyl sulfonate) (PVS, Air Products, Mn 2000) was used as received. =
=
2.2. Preparation of calcium oxalate crystals
Calcium oxalate crystals were formed from supersaturated solutions at 210 C. Stock solutions of ammonium oxalate (0.15M), calcium chloride (0.4M) and various polymers (1000 ppm) were prepared in doubly distilled water. The polymer solution was mixed with 10 cm3 of 0.15M ammonium oxalate. This was placed in a thermostat equipped with a magnetic stirrer and diluted with distilled water to 990 cm3. To this solution 10 cm3 of 0.4M calcium chloride was added with constant mixing. The initial concentrations of calcium and oxalate were 4 x 103M and 1.5 X 103M, respectively. A white precipitate of calcium oxalate was observed to form and slowly settle out of the solution. Crystals were recovered by pressure filtration using a 0.45 ~tm microporous filter. The pH measured before and after the crystallization was 6.5 ±0.3. The samples were then air-dried and stored in vials for further study. 2.3. Preparation of calcium oxalate hydrates
In order to calibrate the X-ray method of analysis of the different calcium oxalate hydrate forms, it was necessary to prepare separately each of the forms. Calcium oxalate monohydrate or whewellite (COM) was easily prepared under the conditions in section provided that theoxalate polymeric given component was 2.2, excluded. Calcium dihydrate or weddelite (COD) forms spontaneously in urine and Gardner [15] showed how it could be prepared from synthetic urine. Following their methods, we added to 500 cm3 of freshly voided urine, 25 cm3 of ammonium oxalate (0.25M) with constant stirring. The precipitate formed was filtered and dried and found by X-rays to be exclusively dihydrate. There are different ways to make COT crystals [16]. In this work, COT was exclusively formed when calcium oxalate precipitated as described in section 2.2 in the presence of 5 ppm PAAL.
=
2.4. X-ray diffraction standards for calcium oxalate hydrates
Mechanical mixtures of COM—COT and COD—COT were prepared and their powder X-ray
J.S. Manne et at.
/ Effect of anionic polyelectrolytes on
crystallization ofcalcium oxalate hydrates
629
therms are presented in terms of the oxalate to calcium ratio as a function of the oxalate con-
1.20
respect to calcium oxalate. Crawford, Crematy and Alexander [22] report average urinary concentrations of calcium and oxalate to be respeccentration. and 1.78 xsupersaturated 104M. Thus, with the lively 4.0 x Urine 103Mis ordinarily concentration product in urine is about 280 times larges than that anticipated from the solubility product of COM. In this same source they give the value of calcium oxalate saturation as 5.12 X iO~. Typical supersaturation shown in fig. 2.by sis, the urine oxalate concentrationis was increased In order to obtain sufficient crystals for analy-
0 0.80
P Li_
0.40
/X~
0.00 0.00
0.20
I
I
0.40
0.60
0.80
1
1.00
MASS FRACTION Fig. 1. Calibration of X-ray powder patterns. (A) Mass fraction
of COM, in a mixture of COM (29 = 14.9) and COT (29 16.1), versus the area fraction of the COM peak of the total peak areas. (•) Mass fraction versus peak area fraction for COD (20=14.3)inaCOD—COTmixture.
patterns were measured using CaKct X-rays. COM, COD and COT were identified using the published data from the Joint Committee on Powder Diffraction Standards 1971 (COT and COM [17]; COD [18]). Analysis of the mixtures was determined from peak areas for COM at 29 14.93 and 15.29, for COD at 29 14.32 and for COT at 29 13.22 and 16.1. The calibrations of peak area fraction as a function of mixture mass fraction given in fig. I were used to determine the fraction of the various hydrate forms in the different samples of precipitated calcium oxalate.
about one order of magnitude over that commonly found in urine. In fig. 2, the initial concentration, and the trajectory of solution concentration change during crystallization are given. Although most of the crystal mass is formed at oxalate concentrations greater than those found in urine, the composition trajectory of the present system passes close and parallel to that applicable to ordinary urine. The hydrate form of calcium oxalate which precipitated from solution was a strong function
—
10
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2.5. Scanning electron microscopy
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Dry calcium oxalate crystals were mounted on conductive adhesive, shadowed with a thin coating of gold and observed by SEM.
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3. Results and discussion
OXALATE Fig. 2. Oxalate to calcium ratio versus oxalate concentration.
The solubility products for COM, COT and COD at 37°C are reported to be respectively (in moles2/liter2): 2.47 X iO~, 10.82 x i0~ and 6.09 x IO~[19—21].In fig. 2, these solubility iso-
~
for COM (A), COT (B) and COD (C) is 2.47 x i0~, 10.82x iO~ and 6.09x iO~ moles2/liter2, respectively, at
37°C [19—211.Initial solution conditions (D) and (— the solution trajectory during crystallization and typical concentrations for urine (E) as shown.
J.S. Manne et at. / Effect of anionic polyelectrolytes on crystallization of calcium oxalate hydrates
630
0
20
15
25
2e Fig. 3. X-ray diffraction powder pattern for calcium oxalate crystals recovered from precipitation from a solution containing indicated concentrations of PAAL (ppm).
of the concentration of polyelectrolyte in solution. This is well illustrated by the case of PAA(L). The calcium oxalate crystallization in the presence of low concentrations of low molecular weight poly
Table 1 Effects of various polymer types and concentrations Polymer
(acrylic) acid (PAAL) occurred in less than 10 s while at the higher polymer concentrations the precipitation occurred over a period of 5 mm. The crystals were easily filtered and dried. The X-ray powder patterns for oxalate formed in increasing concentrations of PAAL are given in fig. 3. In the absence of PAAL, the peak is characteristic of COM. As concentration of PAAL increases, the X-ray data indicates a mixture of COM and COT while at concentrations above 50 ppm only COD is found. In all cases the X-ray peaks are distinct well separated and suitable for quantitative analysis. In table I, the effects of the various polymer types and concentrations are summarized. In general, regardless of the character of the polyelectrolyte (carboxylate or sulfonate), as polymer concentration increases there is a shift from monohydrate to trihydrate and finally to dihydrate. In fig. 4a the fraction of each hydrate formed for both PAAL and PAAH is given as a function of polymer concentration. Consider the data for PAAL. At concentration below 0.5 ppm PAAL only COM is found. When the concentration is increased to about I ppm polymer there is a sharp shift from COM to COT formation and COT is observed exclusively in the concentration range between 1 and 5 ppm. At concentrations above 50 ppm only COD is observed. These data suggest that the polyacrylic acid exerts a very strong influence on the crystallization process. Although carboxylate polymers do bind calcium in solution (and thus lower calcium activity), the molar concentrations of polymeric carboxylate are generally much lower than those of the calcium. Even at 100
©
Concentrations of polymer (in parts per million) 0.1
0.5
1.0
5.0
10
50
100
500
PAAL PAAH
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D T, D
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D D T.D trihydrate.
T, D D D
D D D
PMAL M, T M, T PVS M M,T PSSMA M M a) M = calcium oxalate monohydrate; D
T M,T M,T calcium oxalate
T M,T M,T dihydrate; T =
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Effect ofanionic polyelectrolytes on crystallization ofcalcium oxalate hydrates
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Fig. 4. (a) Mass fraction of hydrate forms of calcium oxalate as a function of polymer concentration: for PAAL, 0 is COM, ~D is COT and• is COD; for PAAH, G is COM, 4 is COT and • is COD. (b) Mass fraction of calcium oxalates as a function of the polymer concentration of: PMAAL, ~ is COM, 11 is COT and A 15 COD; for PMAAH, v is COM, ~ is COT and v is COD. (c) Mass fraction of hydrate types as a function of polymer concentration: for PVS, 0 is for COM, W is for COT and• is for COD; for PSSMA, ~ is for COM, II is for COT and A is for COD.
J.S. Manne et at / Effect of anionic polyelectrolytes on crystallization of calcium oxalate hydrates
632
ppm PAAL, the calcium concentration is 3 times higher than the polyelectrolyte fixed charge. In fig. 4a data are also given for hydrate formed in the presence of the high molecular weight poly(acrylic) acid, PAAH. Although the trend is the same as concentration of polymer increases, it appears that
~
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PAAL shifts the oxalate from COM to COT at a lower concentration than does PAAH. High molecular weight polyelectrolyte should have a stronger effect on the solution activity of divalent ions. If the hydrate control were the result of changes in the solution activity of calcium in the
_______
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I Fig. 5. SEM (Cambridge: Model 200) photographs of sample calcium oxalate crystals produced at different conditions. (a) ~OM crystals, formed in 0.5 ppm PAAL; bar indicates 4 ~m. (b) COT (5.0 ppm PAAL). (C) COD (100 ppm PAAL). (d) COD and COT (50 ppm PAAH). (e) COD (100 ppm PMAA). (1) COD (500 ppm PSSMA).
J. S. Manne et at.
/
Effect of anionic polyelectrolytes on crystallization ofcalcium oxalatehydrates
presence of polyelectrolyte, then at the same mass concentration, the high molecular weight polymer should be more effective than the ow molecular weight material. However the opposite is observed. It appears that polymer adsorption is the most reasonable mechanism for hydrate control. A given mass of strongly adsorbed low molecular weight poly(acrylic acid) contains many more polymer molecules and can more effectively control crystal growth over a crystal surface than can a similarly strongly adsorbed high molecular weight material. The low molecular weight material covers more of the crystal surface. In fig. 4b, similar results are shown for high and low molecular weight poly(methacrylic acid). It appears that PMAAL is more effective than PAAL since the change in the calcium oxalate yield from COM to COT occurs at a lower concentration for PMAAL. The appearance of COD occurs at a polymer concentration that is less dependent on polymer type and molecular weight. In fig. 4c, data are given for two sulfonate containing polyelectrolytes, poly(vinyl sulfonate) and poly(styrene sulfonate-alt-maleic anhydride). Surprisingly, the hydrate formed at the different polymer concentrations is very similar to that observed with the carboxylate containing polymers. Since sulfonates are expected to have less effect on the solutions activity of calcium ions than carboxylates, ion binding in solution seems less important than polymer adsorption on the crystal surface. In figs. 5a—5f, SEM micrographs are presented of the various crystal forms observed in this work. The COM and COT in figs. 5a and Sb are faceted single (sometimes clumped) crystals. They appeared similar in shape regardless of the type of solution from which they were formed. In fig. 5c, the typical bipyramidal form of COD is shown in crystals formed in the presence of 50 ppm PAAH. At 100 ppm PAAL the faceted is seen. In fig. 5e, 100 ppm of PMAAL gives large populations of nonfaceted spherullites (dumbbells). Such shapes are common when viscous polymeric fluid control the crystallization process [23]. In the extreme, fig. 5f clearly shows a residual polymer layer around the spheroidal COD particles. Crystalluria commonly contains both the bipyramidal and the
633
spherullitic form of the COD suggesting that it is urine polyelectrolytes that control crystalluria formation. The mechanism for the phenomena described in this report is intriguing but not well understood. However we can make some strong hypotheses based on our results and of others. This phenomena may be due to kinetic effects. It is known from the work of Nancollas and Tomazic [24] that during the crystallization of calcium oxalate hydrates in the presence of polymer that there is a greater adsorption of polyelectrolyte by COM than COD or COT. They also showed that this greater adsorption of polymer is associated with greater inhibition of both COM nucleation and growth than for COD and COT. In polymerfree solution the rate of growth of COM seeds is greater than COT or COD. However, in the presence of polymer the growth rate of COM is slowed enough so that its relative rate is now less than COD or COT. It is also known that COT seeds grow faster than COD. Hence in the medium polymer concentration ranges it is expected that more COT forms than COD. At higher polymer concentrations (order of magnitude 10—6) the growth rate of COD stabilizes, whereas that of COT continues to decrease. Extrapolation from published data shows that the rate constant will finally drop below that for COT. This would help explain the predominance of COD at high polymer concentrations. It is known that COT and COD are less stable than COM and that at equilibrium the COT and COD will convert to COM. However it appears that in the nonequilibrium state that kinetic factors may predominate and lead to the formation of COT and COD in the presence of polymer.
Acknowledgements This work was supported in part by a grant from Dearborn Chemical: Lake Zurich, Illinois. J.S. Manne gratefully acknowledges the support of the Department of Urology, Long Island Jewish Medical Center, and the NIH (research fellowship F32 DK08280-01), Columbia University.
634
J.S. Manne et at
/
Effect ofanionic polyelectrolytes on crystallization of calcium oxalate hydrates
References
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