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Effect of bacteria and salinity on calcium sulfate precipitation
Desalination 287 (2012) 301–309
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Desalination j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / d e s a l
Effect of bacteria and salinity on calcium sulfate precipitation R. Sheikholeslami ⁎, G.T. Lau School of Engineering, University of Edinburgh, Edinburgh, EH9 3JL, UK
a r t i c l e
i n f o
Article history: Received 23 April 2010 Received in revised form 25 January 2011 Accepted 26 January 2011 Available online 15 March 2011 Keywords: Composite Fouling Inorganic and biological fouling Calcium sulfate Pseudomonas Fluorescence Scaling Membrane fouling
a b s t r a c t Membrane and thermal desalination are plagued by composite fouling because of inorganic, biological and organic constituents. Interactive effects in composite fouling, though of paramount practical importance, are mostly ignored. This paper presents interactive effects of biological matter, Pseudomonas Fluorescence (PF), on precipitation of calcium sulfate, a sparingly soluble salt and an inorganic foulant. Experiments were conducted in isothermal (25 °C) batch tests. The effect of PF and its concentration on CaSO4 precipitation and deposit structure at varying salinities were determined. The Pitzer model, suitable for high salinities, is used in calculation of instantaneous CaSO4's ion activity products. The Scanning Electron Microscopy (SEM) and X-Ray Diffraction (XRD) show that salinity and concentration levels alter the crystal structure but not the crystal phase (gypsum). Under the influence of PF, crystallization of CaSO4 did not occur and sulfur levels significantly increased; it can be surmised that microbial activity of PF reduced the sulfate ions to elemental sulfur and altered precipitation of CaSO4. This does not suggest that biological fouling or presence of bacteria generally inhibits precipitation fouling of calcium sulfate; it underlines that the generally ignored interactive effects are of significant importance and should be considered as corner stone in advancement of fouling research. © 2011 Elsevier B.V. All rights reserved.
1. Introduction The scarcity of fresh water and declining quality of water resources are major cause of problem in many countries around the world. Freshwater available to humans and not in glaciers, constitute less that 0.14% of Earth's surface and as such it should be considered an endangered species. Since more than 70% of Earth is occupied by seawater, desalination of seawater and also other saline waters have great potential to contribute to having alternative sources of fresh water supply. Fouling hampers the operation of both thermal and membrane desalination units and has operational, economical and environmental penalties. System and feed water characterization and design of appropriate pretreatment process are paramount to prevention of fouling in desalination units and in particular proper operation of Reverse Osmosis (RO) units. Despite the multitude of research on fouling control, pretreatment technologies and anti-fouling membranes and modules, membrane fouling is still a limiting factor which continues to exist persistently and hampers the operation of membrane separation technologies. Fouling results in permeate flux decline, poor solute rejection and even eventual blockage of flow channel; it shortens the lifespan of membranes and reduces its overall efficiency as well as increasing its maintenance and operational costs. The extent of membrane fouling is
⁎ Corresponding author. E-mail address: [email protected] (R. Sheikholeslami). 0011-9164/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.desal.2011.01.075
often associated to the properties of feed system. There are many foulants naturally present in seawater and they can be classified into soluble inorganic salts, colloids, biological matters and dissolved organic compounds. There are several types of fouling occurring simultaneously in practical industrial water systems but due to its complexity, most of the research focus on each type of fouling in isolation. The traditional fouling research usually looks at the fouling behavior of a given foulant and as such translation of laboratory results for these studies to actual operational condition is usually unsatisfactory. There have been numerous separate studies on biological fouling [1–8] and on inorganic fouling of various process equipments [9–14]. However, little attention has been paid to simultaneous inorganic and biological fouling of aqueous systems. The interactive effects in composite fouling which are affected by the changes in concentration, temperature, pH, nutrients, ionic and organic and biological species, colloids, O2 content, CO2 content and flow velocity were discussed in two review papers by Sheikholeslami [15,16]. The major fouling contributors to RO fouling are soluble inorganic salts, dissolved organic matter, and colloidal size biological and particulate matter. It is of importance to look at the interactive effects of biological and inorganic fouling. Some of the major contributors to inorganic fouling are CaSO4 and CaCO3. Biological matters have a wide range and certainly differ from location to location greatly. It is important to look at the interactive effects of biological and inorganic fouling; a more fundamental and systemic assessment would be to choose a well-studied biological matter and investigate its presence on precipitation kinetics, thermodynamics,
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and behavior of inorganic soluble salts that contribute to fouling in RO units. As such in this work PF which is a well studied [17–23] biological foulant is used to investigate its effects on CaSO4 precipitation. The objective of this study is to investigate and explore the interactive effects of inorganic and biological fouling in seawater desalination using CaSO4 and PF at salinities and concentrations encountered in seawater RO. 2. Materials and methods The tests were carried out in constant volume batch reactors for solution matrices made of model simulated solutions with water salinities representing seawater recovery levels of 35% and 50% and Concentration Polarization (CP) of 1 and 2 as shown in Table 1. The feed seawater composition in a RO desalination plant contains approximately 0.5 M NaCl and 0.03 M CaSO4. A commercialized seawater RO (SWRO) desalination plant can operate to a maximum of 50% recovery. Therefore the NaCl and CaSO4 concentrations in the exit concentrate stream are, respectively, about 1 M and 0.06 M; for a CP of 2, the estimated concentrations next to the membrane surface are thus up to 2 M for NaCl and 0.12 M for CaSO4 at the end of the RO module. In this work, experiments were carried out to investigate the interactive effect of CaSO4 precipitation in the range of 0.06 M to 0.10 M, in NaCl solution ranging between 0.5 M to 1.5 M and with various concentrations of PF ranging between 1 × 106 cfu/ml and 20 × 106 cfu/ml. The experimental conditions are summarized in Table 2. The experiments were carried out in batch tests at a constant temperature of 25 °C. The thermodynamics and kinetics assessments were carried out for each batch test. All chemicals and reagents used were of analytical grade. All the test tubes used in the experiment had been scrubbed, rinsed with dilute hydrochloric acid, washed thoroughly with distilled water, and dried before use. The supersaturating solutions of NaCl and CaSO4 were prepared in the laboratory by mixing solutions of Na2SO4 and CaCl2 respectively with the NaCl solution. All solutions were prepared with micro-filtered (with 0.22 μm Millipore filter) distilled water and further filtered after preparation to remove any impurities and undissolved salts that were present. The concentration and pH of each individual salt solution were measured before mixing. The individual salt solutions were prepared and then carefully mixed and transferred into a series of 30 ml plastic test tubes. The 30 ml plastic test tubes were used as constant volume batch reactors for kinetic analysis as well as determination of thermodynamics and water quality. A few coupons of reverse osmosis membrane were cut into equal size and shape of dimension 1 cm × 1 cm and inserted into the test tubes. They were used to observe the adherence of precipitates and extracellular polymeric substances (EPS) to the membranes. Nutrient broth was composed of Lab-Lemco powder (1 g/L), Yeast extract (2 g/L), peptone (5 g/L) and sodium chloride (5 g/L) and PF cultures were prepared by and obtained from the microbiology laboratory. There were 20 experimental batches of which 9 batches contained only salt solutions while the other 11 batches contained salt solutions, nutrient and bacteria (PF). Each batch was monitored for more than 3 weeks to ensure that equilibrium had been reached. At a given time, one tube of sample from each batch of mixed solution was removed for water quality analysis and pH measurement; the solution was discarded after the analytical tests were conducted. The water quality was determined by measurement of Table 1 Salt concentrations of CaSO4 and NaCl at different water recoveries and CP values. Recovery
0%
35% (CP = 1)
50% (CP = 1)
35% (CP = 2)
50% (CP = 2)
NaCl (M) CaSO4 (M)
0.5 0.03
0.75 0.05
1.0 0.06
1.5 0.1
2.0 0.12
Table 2 Experimental conditions at 25 °C for the tests. Batch no.
NaCl (M)
CaSO4 (M)
Nutrient (ppm)
PF (× 106 cfu/ml)
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
0.5 0.5 0.5 1.0 1.0 1.0 1.5 1.5 1.5 1.0 1.0 1.0 1.0 1.0 0.5 0.5 0.5 1.5 1.5 1.5
0.065 0.080 0.100 0.065 0.080 0.100 0.065 0.080 0.100 0.060 0.060 0.060 0.080 0.100 0.060 0.080 0.100 0.060 0.080 0.100
– – – – – – – – – 5 5 5 5 5 5 5 5 5 5 5
– – – – – – – – – 1 10 20 10 10 10 10 10 10 10 10
sodium (Na+), calcium (Ca+ 2) and sulfate (SO− 4 ) ions using ICP-AES (Inductively Coupled Plasma-Atomic Emission Spectroscopy). Each sample was filtered with a 0.22 μm filter and further diluted with micro-filtered water before ICP analysis so that the salts were below the solubility limit as to avoid precipitation. The precipitates and extracellular polymeric substance that adhered on the surface of membrane at the end of each experimental batch were analyzed using Scanning Electron Microscopy (SEM) to examine their surface structure and porosity. Electron Dispersion X-ray (EDX) Spectroscopy was used to determine chemical constituents in the precipitates and extra-cellular polymeric substance for their elemental composition identification and quantification. The crystallographic structure of precipitates was characterized and identified using X-Ray Diffraction (XRD). The analysis of precipitate formed in the runs containing biological matter needed specific preparation as discussed here. Solution of 2% by volume of Osmium Tetraoxide (OsO4) was prepared and the sample was soaked in it for an hour. The soaking was for fixation of the structure of sample. Following that the sample was rinsed with distilled water 3 times to remove any remains of Osmium Tetraoxide. The sample then was dehydrated with stagewise concentrations of ethanol (starting from 30%v/v, followed by 50%v/v, 75%v/v, 95%v/v and gradually increased to 100% v/v). Each dehydration stage took 15 min except for the last two stages that required 30 min for each. Then samples underwent critical point drying (CPD) using Bal-tec 030 Critical Point Dryer and liquid CO2 to replace the ethanol content. 3. Results and discussion The accuracy and reproducibility of results were assessed before any attempt in data reduction. Experimental error analysis was conducted for preparation of samples and all sample taking and measurements. Fig. 1 represents an example of repeated tests showing a very good reproducibility; the maximum uncertainty for the repeated tests was 7%. 3.1. Thermodynamics analysis The activity coefficients of ionic species in complex seawater solution are calculated by the Pitzer model which is applicable to ionic strength above 0.01 M and up to 6 M. Details are provided elsewhere [24]. General thermodynamic relations are available elsewhere [4]. The instantaneous Ion Activity Product (IAP) of CaSO4 in the experimental run was calculated from the measurement of Na+,
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Fig. 1. A Sample of reproducibility test.
Ca2+ and SO2− concentration in the solution using the Pitzer model 4 and plotted as a function of time. All samples in the experimental run were measured until equilibrium was reached. The IAP value obtained at equilibrium is the Thermodynamic Solubility Constant (Ksp) of the salts in solution. Summary of the thermodynamic and kinetic calculations are listed in Table 3. Presence of PF had affected, as will be discussed later, the precipitation thermodynamics and kinetics and had rendered the calculation of their constants not possible and as such only the results for the runs without bacteria are reported in Table 3 where n is order of the reaction and S.S. is the Super Saturation. Results from Table 3 show that the average Ksp value for CaSO4 was 2.49 ± 0.4 × 10− 5 (mol/l)2 which was comparable with the published data for CaSO4 (2.52 × 10− 5 (mol/l)2) at 25 °C [25] and the average of 2.43 ± 0.7 × 10− 5 (mol/l)2 [24]. Fig. 2 shows that for 0.5 M NaCl solution the initial drop in instantaneous IAP values of CaSO4 versus time was the same for the three solutions with varying CaSO4 concentrations. However, as the concentration of the NaCl increased to 1 M and 1.5 M, there was a clear difference in the initial instantaneous IAP drop for various CaSO4 concentrations. These results indicated that the dependence on the actual degree of supersaturation, which is salinity dependent, becomes important at salinity levels higher than 1 M. The initial supersaturation of solution decreases as NaCl concentration increases. For a given calcium sulfate concentration, the instantaneous IAP as a function of time is plotted in Fig. 3 for various salinity levels. At higher salinity level of 1 M, there was less difference in the instantaneous IAP decline versus time for various calcium sulfate concentrations. The effect of salinity on instantaneous IAP was more pronounced for lower calcium sulfate concentrations. In general, the variations were more pronounced at lower solution supersaturation levels, being higher salinities and lower calcium sulfate concentrations. At high supersaturation levels spontaneous and homogeneous nucleation occurs rendering the effect of other factors much less pronounced.
3.2. Induction periods Supersaturation is a pre-requisite for nucleation to occur which would further lead to crystallization. A supersaturated solution is
Table 3 Summary of results for pure CaSO4 at various salinities. Batch no.
NaCl (M)
CaSO4 (M)
n
1 2 3 4 5 6 7 8 9
0.5
0.065 0.080 0.100 0.065 0.080 0.100 0.065 0.080 0.100
4.03
1
1.5
Krxn (mol/l∙h) 0.0021
6.18 0.0062 7.29 0.0067
Ksp (CaSO4) (mol/l)2
S.S. (initial)
Induction time (h)
2.13E-05 2.21E-05 2.88E-05 2.49E-05 2.94E-05 2.63E-05 2.53E-05 2.49E-05 2.12E-05
4.4 5.7 7.6 2.6 2.9 6.0 2.0 3.6 5.8
5.00 0.17 0.00 24.00 1.03 0.00 48.00 4.00 0.00
Fig. 2. Instantaneous IAP for a given salinity at various CaSO4 concentrations.
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Fig. 4. a. Precipitation of CaSO4 in presence of NaCl as monitored by reduction of Ca2+ concentration. b. Precipitation of CaSO4 in presence of NaCl as monitored by reduction concentration. of SO2− 4
concentrations increase to 0.08 M and 0.1 M. At the highest calcium sulfate concentration, the nucleation is spontaneous and induction time is independent of salinity level. These results are in good agreements with those previously reported by Sheikholeslami and Ong [24]. At lower concentration levels needle-liked star-shaped crystals were formed. As the calcium sulfate concentration increased, and solution had shorter induction periods, small and flaky crystals were formed; this was due to spontaneous precipitation which usually results in smaller crystal sizes in greater numbers. Fig. 3. Instantaneous IAP for a given CaSO4 concentration at various Salinities.
called a metastable solution when no nucleation occurs and solution remains supersaturated. The period of time in which the supersaturated solution remains in a metastable state before nucleation and crystallization occurs is called induction period. The induction period of salt precipitation is dependent upon the type of precipitating salt and its concentration. The experimental results have shown that presence of other soluble ionic species has an effect on the induction period of precipitating salt. The induction period of CaSO4 was examined by monitoring the ionic species in a non-precipitating reduction of both Ca2+ and SO2− 4 NaCl solution (Figs. (4a) and (4b)). Time and observations for crystallization were also recorded throughout the whole experimentation. Table 3 shows that the induction period for pure CaSO4 (0.065 M) increases significantly from 5 h to 48 h as concentration of NaCl increases from 0.5 M to1.5 M resulting in decreases in the degree of supersaturation due to the existence of non-precipitating NaCl salt and increases in the ionic strength of solution. The presence of nonprecipitating soluble salt changes the Gibbs free energy of nucleation reaction and hence affecting the surface energy of a crystal. The effect of ionic strength on induction time reduces as the calcium sulfate
3.3. Effect of PF The experimental results in Fig. 5 show a representative run in presence of PF; the concentration of S2− increased over a period of time in the presence of the bacteria. The same trends were obtained for runs at other calcium sulfate concentrations and salinities in presence of PF. The concentrations of Ca2+ and Na+, Cl− were monitored throughout the runs and the analytical results showed that they remained constant. The reason for the increase in S2− concentration was due to microbial activities. The results suggested there was generation of hydrogen sulfide (H2S) by the bacteria during microbial activity where reduction of sulfate (SO2− 4 ) occurred. Some samples were tested by ion chromatography to confirm this hypothesis. The results from ion chromatography analysis showed a 2− over a period of reduction of SO2− 4 concentration with increase in S time; the results for a representative run are plotted in Fig. 5 for the 0.1 M CaSO4 solution in 1 M NaCl with 10 × 106 cfu/ml PF. The same trends were obtained for other runs in presence of PF. The analytical results confirmed the hypothesis that the bacteria consumed sulfate as nutrient for their microbial activities and the reduction of sulfate further affects the metastability of the calcium sulfate in the solution. The reduction of sulfate to sulfide changed the
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6 Fig. 5. Concentration of S2− and SO2− 4 of 0.1 M CaSO4 in 1 M NaCl with 10 × 10 cfu/ml PF.
thermodynamic solubility of solution and prevented crystallization of CaSO4. Therefore no induction period, kinetic or thermodynamic analysis could be conducted for solutions that contained PF. Brownian diffusion, aggregation and sedimentation are responsible for the formation of biofilm in this work as the experiments were conducted in a batch system where there was no fluid flow. A period of time was required for the bacteria to arrive at the surface of a membrane, attach and colonize. The formation of extracelluar polymeric substance (EPS) was mostly observed after 96 h at low salinities. At higher salinities, the time period for formation of EPS decreased to 48 h. These results showed that high salinities promote faster formation of biofilm as suggested previously [15] and that higher ionic strength reduces the stability of bacteria in suspension and hence promotes faster biofouling. As stated above, no CaSO4 crystals were formed for all the experiments conducted in presence of PF irrespective of calcium sulfate concentrations or salinities in the solution. 3.4. Kinetics analysis Kinetic rates and order of reaction for pure CaSO4 precipitation at various salinities were analyzed using nth order initial rate method. In this study the solution salinities were at 0.5, 1, and 1.5 M. Therefore the kinetic analysis showing the effect of salinity is not comprehensive and the results are not conclusive. However, in light of our previous experimental studies also showing the effect of salinity on kinetics [24,26,27], the kinetic analysis is reported here. The kinetic rate order, n and rate constant, Krxn for CaSO4 at various salinities are summarized in Table 3. The results show that the order of reaction for CaSO4 precipitation was very much affected by the presence of NaCl. The presence of non-precipitating species increased the order of reaction rate (n); it was 4.03, 6.18 and 7.29, respectively, for 0.5 M, 1 M and 1.5 M NaCl solutions. Results shown in Fig. 6 suggest that CaSO4 precipitation in NaCl solution was a poly-nuclear reaction. Extrapolation of the order reaction (n) curve to 0 M NaCl concentration results in a value of approximately 2 for the order of reaction (n).
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Thus, it is in good agreement with the literature value obtained by Liu and Nancollas [28] indicating that CaSO4 dihydrate (gypsum) precipitation in stable supersaturated solution at a temperature between 15 °C and 45 °C followed a 2nd order rate equation with respect to depletion of total Ca2+ [28]. However, the Krxn values obtained in this work cannot be used for comparison with theirs [28] as this work was conducted without the aid of seeds for crystal growth and surface area was unknown whereas their experiments were conducted using a seeded crystal growth technique and known surface area of seeds. Primary homogeneous nucleation and spontaneous precipitation was involved in this work and hence the Gibbs free energy of reaction would have been more than that required for the work of Liu and Nancollas where a secondary heterogeneous nucleation occurred. Table 3 shows that the reaction rate constant, Krxn, values increased from 0.021 to 0.067 mol/l∙h as concentrations of NaCl increased from 0.5 M to 1.5 M. Although the reaction rate increased with concentrations of NaCl, it could not be used for direct comparison with the results of Liu and Nancollas [28] since the growth area and number of nucleus to initiate precipitation are unknown in this work. The overall rate of reaction is considered to be faster at higher salinities. This was in good agreement with previous results of Sheikholeslami and Ong [24] where the same trend was observed and the order of reaction rate increased with increases in salinity values. As stated before, kinetic analysis could not be conducted for the runs in presence of PF due to absence of CaSO4 precipitation. 3.5. pH analysis The pH for seawater is around 8 due to contribution of major bicarbonate species. The pH of all the experimental batch tests was monitored and measured to be within the range of 7 to 8 throughout the experiment. Comparison of pH change over time for the tests with pure CaSO4 (Fig. 7a) and those in presence of PF (Fig. 7b) at 1 M NaCl depicts the effect of bacteria on solution pH. The pH for 0.065 M CaSO4 in 1 M NaCl during the run remained within the pH range of 7 to 7.4 while the pH for CaSO4 solutions in the presence of PF in1 M NaCl decreased from about 7.4 to 6.3; there was a significant change of pH under the influence of bacteria and it further depicts influence of biological fouling on inorganic fouling as most inorganic salt crystallizations are pH dependent. The drop in pH over a period of time was due to microbial activities by production of CO2 as a result of respiration and by reduction of sulfate to H2S. The formation of hydrogen sulfide could be due to anaerobic condition created, after depletion of limited dissolved oxygen in the solution, in the experiment as there was no air/gap in the constant volume batch reactors. After depletion of the dissolved oxygen, bacteria could reduce sulfate to sulfide by using the sulfate ions as a terminal electron acceptor in respiration and so lead to the reduction of sulfate. The hydrogen sulfide (H2S) produced would dissociate into HS− ions; dissociation of HS− ions into elemental sulfur and H+ ions in the solution contributed to the increase of S2− concentration and lowering the pH. A look at Fig. 7b shows that the final solution pH was lowest for the lowest microorganism concentration; however, this does not suggest that the maximum pH drop was for this run. Fig. 7b shows that the initial pH values were about 7.45, 7.25, and 7.08 for, respectively, runs with highest to lowest microorganism concentrations. Therefore, the actual pH drop was highest for the run containing highest microorganism concentration and was lowest for the run containing lowest microorganism concentration. 3.6. Scale structure and morphology
Fig. 6. Rate constant and order of reaction for pure CaSO4 at various salinities.
Figs. 8–10 show the effect of calcium sulfate concentration on the scale morphology. The results shown are for tests conducted at 1 M
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Fig. 9. SEM images of 0.080 M pure CaSO4 in 1 M NaCl.
salinity; similar trends were observed for other salinities. Fig. 8 shows that at low concentration of CaSO4 (0.065 M), the crystals exhibited long and fine needle-like structure which is typical of calcium sulfate dihydrate (CaSO4.2H2O) or gypsum. As concentration of CaSO4 increased to 0.1 M (Fig. 10), the crystal structure was altered; however, the phase did not change and remained as gypsum based on the XRD results. At a higher concentration of CaSO4 (Fig. 10), the crystals appeared to be fine white powder that looked ‘flaky and flowery’; this was due to spontaneous precipitation in the highly supersaturated solution where crystals were formed instantaneously with no induction time for them to develop and crystallization was more of a result of nucleation than as a result of crystal growth. These findings were in good agreement with the scale morphology studies by Sheikholeslami and Ong [24]. It was observed that as salinity increased from 0.5 M to1.5 M, the size of CaSO4 crystals became larger. The formation of larger sized
crystals was a result of the higher ionic strength solution and the longer induction period where crystallization was more due to crystal growth than nucleation. Therefore, in general the crystal size was affected by the degree of supersaturation of solution. As supersaturation decreases, the crystal sizes become larger and vice versa; overall, a longer induction period favors crystal growth and larger crystals while a shorter induction period favors nucleation and smaller crystal sizes. During the experiment, it was observed that some crystals were loosely adhered to the surface of the test-tubes while others were floating freely in the solution. The CaSO4 crystals that had adhered on the test-tube surface were not tenacious and could be easily removed by scrubbing with water. Also the crystals that had adhered on the membrane fell off easily during coating for SEM analysis and hence the crystals were analyzed without membranes. Figs. 11–13 show the SEM images of EPS (Extracellular Polymeric Substance) formed by PF in presence of CaSO4 in NaCl solutions. The images do not show any differences in the morphologies of EPS formed in terms of bacteria concentration. The structure of biofilms formed by PF exhibited cross-link cellular fibers interlinking with one another. The SEM images show that PF of approximately 1.4 μm size was firmly attached onto the EPS fibers. At higher magnifications, the bacteria appeared to be elongated and rod shaped. The EDX analysis showed that there were no salt crystals embedded within the EPS. During the experiment, it was observed that the amount of EPS formed was dependent on the concentration of bacteria present; as expected, the thickness of biofilms increased when bacteria concentrations increased from 1 × 106 cfu/ml to 20 × 106 cfu/ml due to enhanced microbial activity. It was also observed that Ca2+ presence
Fig. 8. SEM images of 0.065 M pure CaSO4 in 1 M NaCl.
Fig. 10. SEM images of 0.1 M pure CaSO4 in 1 M NaCl.
Fig. 7. a. pH versus time for pure 0.065 M CaSO4 at 1 M NaCl. b. pH versus time for pure 0.06 M CaSO4 in presence of PF at 1 M NaCl.
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Fig. 11. 0.06 M CaSO4 in 1 M NaCl with 1 × 106 cfu/ml PF at 400 and 4000 magnification.
Fig. 12. 0.06 M CaSO4 in 1 M NaCl with 10 × 106 cfu/ml PF at 400 and 4000 magnifications.
had some effect on the adhesiveness of biofilm; as the Ca2+ concentration increased, the biofilm adhered to the membrane surface and bottom surface of the test-tube. They later broke off from the
main substratum and formed individual colonies within the solution and also adhered on the wall surface of test tubes after a period of time.
Fig. 13. 0.06 M CaSO4 in 1 M NaCl with 20 × 106 cfu/ml PF at 400 and 4000 magnification.
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Fig. 14. EDX of crystals from 0.065 M CaSO4 in 0.5 M NaCl.
Fig. 16. EDX of crystals from 0.08 M CaSO4 in 1 M NaCl.
3.7. Electron Dispersion X-ray analysis
4. Conclusions
The CaSO4 crystals were further analyzed on their elemental composition using Electron Dispersion X-ray (EDX) spectroscopy. The EDX results showed that there was no difference in the elemental composition of precipitates; the major components present in the crystals were calcium, sulfur and oxygen at low salinities. Higher quantities of calcium, sulfur and oxygen were detected as concentration of CaSO4 increased. The results showed that relatively high purity of CaSO4 crystals were obtained at low salinities. Figs. 14 and 15 show the EDX spectrums of 0.06 M and 0.1 M CaSO4 in 0.5 M NaCl, respectively. As salinities increased, relatively high quantities of sodium and chloride were detected by the EDX analysis revealing that during precipitation some sodium and chloride ions were trapped within calcium sulfate thus lowering the purity of CaSO4 crystals formed. This suggested that salinities not only affect the induction period of CaSO4 precipitation but also the purity of CaSO4 crystals formed. Figs. 16 and 17 show the EDX spectrums of 0.08 M CaSO4 in 1 M NaCl and 1.5 M NaCl, respectively.
Batch tests were carried out to study the thermodynamics and kinetics of calcium sulfate under the influence of various concentrations of PF at various salinities ranging from 0.5 M to 1.5 M at isothermal conditions of 25 °C. The Pitzer model was used to determine the ion activity coefficients as discussed elsewhere [24]. The increase in the concentration of non-precipitating NaCl salt did not affect the thermodynamic solubility product (Ksp, ion activity product at equilibrium) of CaSO4. Initial rate method was used to determine the kinetics for CaSO4 precipitation reaction. It was found that kinetics of CaSO4 was strongly affected by the salinity levels. As concentration of NaCl increased, the rate of reaction increased from 0.0021 to 0.0067 mol/l∙h. With increases in salinity the order of reaction increased from 4.03 to 7.29 indicating a poly-nuclear reaction; extrapolating the trend to zero salinity level resulted in the order of reaction of 2 which was in good agreement with values of Liu and Nancollas [28]. The increase in concentration of CaSO4 affected the scale morphology causing the crystals to become flaky and fine in comparison to crystals formed at lower concentration of CaSO4 which exhibited a needle-shaped structure. However, the crystal phase of CaSO4 remained as gypsum (calcium sulfate dihydrate) irrespective of salinity and concentrations levels. There was no crystallization of CaSO4 in the presence of PF. However, there was a significant decline in the sulfate concentration in the solution with time. The solutions were tested for sulfide concentration and the results showed a progressively increasing sulfide concentration as the sulfate concentrations dropped over period of time. This reveals the reduction of sulfate by the bacteria. In presence of bacteria, there was also a significant drop in the pH of solutions from a range of pH 7.4 to pH 6.3 over time. The drop in pH is expected to have been caused by contribution of H+ ions from dissociation of H2S and also production of CO2 due to bacterial respiration.
3.8. XRD analysis Precipitates were examined by XRD analysis where angle of the incident beam is diffracted according to the crystal lattice spacing and hence identifying the crystal phase. The diffraction angle has the unit of Degree 2-Theta and is calculated using Bragg's Law (d = λ = sinðθÞ) where θ is the diffraction angle (Degree 2-Theta), λ is X-ray wavelength (1.5405 A for Cu tube), and d is lattice spacing. Different crystal phases have different lattice spacing of atoms and hence the angle of diffraction differs from one crystal phase to another. The XRD analysis was used to identify crystal phases and the results showed CaSO4∙2H2O (gypsum) to be the crystal phase for all precipitates. Neither salinity nor the calcium sulfate concentrations affected the crystal phase; though they did affect the crystal size and shape.
Fig. 15. EDX of crystals from 0.1 M CaSO4 in 0.5 M NaCl.
Fig. 17. EDX of crystals from 0.08 M CaSO4 in 1.5 M NaCl.
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The results further highlight and reinforce that the interactive effects in fouling have significant effect and cannot and should not be ignored. Acknowledgment The authors gratefully acknowledge supply of membranes by Hydranautics and financial support of Middle Eastern Desalination Research Centre (MEDRC) in this work. References [1] W.G. Characklis, Microbial fouling, in: W.G.a.M., K.C. Characklis (Eds.), Biofilms, Wiley, New York, 1990. [2] W.G. Characklis, Biofouling: effects and control, in: W.G. Characklis (Ed.), Biofouling and Biocorrosion in Industrial Water Systems, 1991, pp. 7–28. [3] H.C. Flemming, Biofouling in water treatment, Proceedings of the International Workshop on Industrial Biofouling and Biocorrosion, 1990. [4] W.E. Garret, T.J. Rabas, Biofouling mitigation of plain and enhanced condenser tubes, Fouling Mitigation of Industrial Heat Exchangers, 19958, Eng. Found. [5] C.B. Panchal, P.K. Takahashi, W. Avery, Biofouling control using ultrasonic and ultraviolet treatments, Fouling Mitigation of Industrial Heat Exchangers, 1995, p. 118, San Luis Ebispo, CA, Argonne National Lab and Department of Energy. Report No ANL/ES/CP–92228; CONF-9506406-2. [6] E.S. Beardwood, Modeling and performance monitoring of biofilms, Fouling Mitigation of Industrial Heat Exchangers, Engineering Foundation, 1995. [7] T.R. Bott, Aspects of biofilm formation and destruction, Corrosion Reviews 11 (1–2) (1993) 1–24. [8] J.W. Costerton, Z. Lewandowski, Microbial biofilms, Annual Review of Microbiology 49 (1995) 711–745. [9] M. Brusilovsky, J. Borden, D. Hasson, Flux decline due to gypsum precipitation on RO membranes, Desalination 86 (1992) 187–222. [10] D. Hasson, Perl, Scale deposition in a laminar falling-film system, Desalination 37 (1981) 279–292. [11] D. Hasson, Rate of decrease of heat transfer due to scale deposition, Dechema Monographien 47 (1962) 233. [12] D. Hasson, D. Bramson, B. Limoni-Relis, R. Semiat, Influence of the flow system on the inhibitory action of CaCO3 scale prevention additives, Desalination and Environment, European Desalination Society, 1996.
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[13] R. Sheikholeslami, Tube Material and Augmented Surface Effects in Heat Exchanger Scaling, University of British Columbia, Vancouver, Canada, 1984. [14] R. Sheikholeslami, A.P. Watkinson, Scaling of plain and externally finned heat exchanger tubes, American Society of Mechanical Engineers, Journal of Heat Transfer 108 (1) (1986) 147–152. [15] R. Sheikholeslami, Composite fouling — inorganic and biological — a review, Evironmental Progress 18 (2) (1999) 113–122. [16] R. Sheikholeslami, Composite fouling of heat transfer equipment in aqueous media — a review, Special Issue of Heat Transfer Engineering J. 21 (2) (2000) 34–42. [17] D.E. Caldwell, J.R. Lawrence, Growth kinetics of Pseudomonas Fluorescens microcolonies within the hydrodynamic boundary layers of surface microenvironments, Microbial Ecology 12 (1986) 299–312. [18] J.E. Duddridge, C.A. Kent, J.F. Laws, Effect of surface shear stress on attachment of pseudomonas fluorescens to S.S. under defined flow conditions, Biotechnology and Bioengineering 26 (1982) 153–164. [19] I.T. Paulsen, C.M. Press, J. Ravel, Complete genome sequence of the plant commensal Pseudomonas fluorescens Pf-5, Nature Biotechnology 23 (2005) 873–878. [20] D. Saravanakumar, R. Samiyappan, ACC deaminase from Pseudomonas fluorescens mediated saline resistance in groundnut (Arachis hypogea) plants, Journal of Applied Microbiology 102 (2007) 1283–1292. [21] X.X. Zhang, P.B. Rainey, The role of a P1-type ATPase from Pseudomonas fluorescens SBW25 in copper homeostasis and plant colonization, Molecular Plant-Microbe Interactions 20 (2007) 581–588. [22] P. Hseuh, L. Teng, H. Pan, Y. Chen, C. Sun, S. Ho, K. Luh, Outbreak of Pseudomonas fluorescens bacteremia among oncology patients, Journal of Clinical Microbiology 36 (1998) 2914–2917. [23] C.K. Stover, X.Q. Pham, A.L. Erwin, Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen, Nature 406 (2000) 959–964. [24] R. Sheikholeslami, H.W.K. Ong, Kinetics and thermodynamics of calcium carbonate and calcium sulfate at salinities up to 1.5 m, Desalination 157 (2003) 217–234. [25] P.W. Atkins, Physical Chemistry, 6th ed.Oxford University Press, Oxford, 1998. [26] R. Sheikholeslami, Scaling of process equipment by saline streams — challenges ahead, Water Science and Technology: Water Supply — IWA Journal 49 (2) (2004) 201–211. [27] R. Sheikholeslami, Nucleation and kinetics of mixed salts in scaling, AIChE 49 (1) (2003) 194–202. [28] B. Liu, G.H. Nancollas, The kinetics of crystal growth of calcium sulfate dihydrate, Journal of Crystal Growth 6 (1970) 281–289.
Contents lists available at ScienceDirect
Desalination j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / d e s a l
Effect of bacteria and salinity on calcium sulfate precipitation R. Sheikholeslami ⁎, G.T. Lau School of Engineering, University of Edinburgh, Edinburgh, EH9 3JL, UK
a r t i c l e
i n f o
Article history: Received 23 April 2010 Received in revised form 25 January 2011 Accepted 26 January 2011 Available online 15 March 2011 Keywords: Composite Fouling Inorganic and biological fouling Calcium sulfate Pseudomonas Fluorescence Scaling Membrane fouling
a b s t r a c t Membrane and thermal desalination are plagued by composite fouling because of inorganic, biological and organic constituents. Interactive effects in composite fouling, though of paramount practical importance, are mostly ignored. This paper presents interactive effects of biological matter, Pseudomonas Fluorescence (PF), on precipitation of calcium sulfate, a sparingly soluble salt and an inorganic foulant. Experiments were conducted in isothermal (25 °C) batch tests. The effect of PF and its concentration on CaSO4 precipitation and deposit structure at varying salinities were determined. The Pitzer model, suitable for high salinities, is used in calculation of instantaneous CaSO4's ion activity products. The Scanning Electron Microscopy (SEM) and X-Ray Diffraction (XRD) show that salinity and concentration levels alter the crystal structure but not the crystal phase (gypsum). Under the influence of PF, crystallization of CaSO4 did not occur and sulfur levels significantly increased; it can be surmised that microbial activity of PF reduced the sulfate ions to elemental sulfur and altered precipitation of CaSO4. This does not suggest that biological fouling or presence of bacteria generally inhibits precipitation fouling of calcium sulfate; it underlines that the generally ignored interactive effects are of significant importance and should be considered as corner stone in advancement of fouling research. © 2011 Elsevier B.V. All rights reserved.
1. Introduction The scarcity of fresh water and declining quality of water resources are major cause of problem in many countries around the world. Freshwater available to humans and not in glaciers, constitute less that 0.14% of Earth's surface and as such it should be considered an endangered species. Since more than 70% of Earth is occupied by seawater, desalination of seawater and also other saline waters have great potential to contribute to having alternative sources of fresh water supply. Fouling hampers the operation of both thermal and membrane desalination units and has operational, economical and environmental penalties. System and feed water characterization and design of appropriate pretreatment process are paramount to prevention of fouling in desalination units and in particular proper operation of Reverse Osmosis (RO) units. Despite the multitude of research on fouling control, pretreatment technologies and anti-fouling membranes and modules, membrane fouling is still a limiting factor which continues to exist persistently and hampers the operation of membrane separation technologies. Fouling results in permeate flux decline, poor solute rejection and even eventual blockage of flow channel; it shortens the lifespan of membranes and reduces its overall efficiency as well as increasing its maintenance and operational costs. The extent of membrane fouling is
⁎ Corresponding author. E-mail address: [email protected] (R. Sheikholeslami). 0011-9164/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.desal.2011.01.075
often associated to the properties of feed system. There are many foulants naturally present in seawater and they can be classified into soluble inorganic salts, colloids, biological matters and dissolved organic compounds. There are several types of fouling occurring simultaneously in practical industrial water systems but due to its complexity, most of the research focus on each type of fouling in isolation. The traditional fouling research usually looks at the fouling behavior of a given foulant and as such translation of laboratory results for these studies to actual operational condition is usually unsatisfactory. There have been numerous separate studies on biological fouling [1–8] and on inorganic fouling of various process equipments [9–14]. However, little attention has been paid to simultaneous inorganic and biological fouling of aqueous systems. The interactive effects in composite fouling which are affected by the changes in concentration, temperature, pH, nutrients, ionic and organic and biological species, colloids, O2 content, CO2 content and flow velocity were discussed in two review papers by Sheikholeslami [15,16]. The major fouling contributors to RO fouling are soluble inorganic salts, dissolved organic matter, and colloidal size biological and particulate matter. It is of importance to look at the interactive effects of biological and inorganic fouling. Some of the major contributors to inorganic fouling are CaSO4 and CaCO3. Biological matters have a wide range and certainly differ from location to location greatly. It is important to look at the interactive effects of biological and inorganic fouling; a more fundamental and systemic assessment would be to choose a well-studied biological matter and investigate its presence on precipitation kinetics, thermodynamics,
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and behavior of inorganic soluble salts that contribute to fouling in RO units. As such in this work PF which is a well studied [17–23] biological foulant is used to investigate its effects on CaSO4 precipitation. The objective of this study is to investigate and explore the interactive effects of inorganic and biological fouling in seawater desalination using CaSO4 and PF at salinities and concentrations encountered in seawater RO. 2. Materials and methods The tests were carried out in constant volume batch reactors for solution matrices made of model simulated solutions with water salinities representing seawater recovery levels of 35% and 50% and Concentration Polarization (CP) of 1 and 2 as shown in Table 1. The feed seawater composition in a RO desalination plant contains approximately 0.5 M NaCl and 0.03 M CaSO4. A commercialized seawater RO (SWRO) desalination plant can operate to a maximum of 50% recovery. Therefore the NaCl and CaSO4 concentrations in the exit concentrate stream are, respectively, about 1 M and 0.06 M; for a CP of 2, the estimated concentrations next to the membrane surface are thus up to 2 M for NaCl and 0.12 M for CaSO4 at the end of the RO module. In this work, experiments were carried out to investigate the interactive effect of CaSO4 precipitation in the range of 0.06 M to 0.10 M, in NaCl solution ranging between 0.5 M to 1.5 M and with various concentrations of PF ranging between 1 × 106 cfu/ml and 20 × 106 cfu/ml. The experimental conditions are summarized in Table 2. The experiments were carried out in batch tests at a constant temperature of 25 °C. The thermodynamics and kinetics assessments were carried out for each batch test. All chemicals and reagents used were of analytical grade. All the test tubes used in the experiment had been scrubbed, rinsed with dilute hydrochloric acid, washed thoroughly with distilled water, and dried before use. The supersaturating solutions of NaCl and CaSO4 were prepared in the laboratory by mixing solutions of Na2SO4 and CaCl2 respectively with the NaCl solution. All solutions were prepared with micro-filtered (with 0.22 μm Millipore filter) distilled water and further filtered after preparation to remove any impurities and undissolved salts that were present. The concentration and pH of each individual salt solution were measured before mixing. The individual salt solutions were prepared and then carefully mixed and transferred into a series of 30 ml plastic test tubes. The 30 ml plastic test tubes were used as constant volume batch reactors for kinetic analysis as well as determination of thermodynamics and water quality. A few coupons of reverse osmosis membrane were cut into equal size and shape of dimension 1 cm × 1 cm and inserted into the test tubes. They were used to observe the adherence of precipitates and extracellular polymeric substances (EPS) to the membranes. Nutrient broth was composed of Lab-Lemco powder (1 g/L), Yeast extract (2 g/L), peptone (5 g/L) and sodium chloride (5 g/L) and PF cultures were prepared by and obtained from the microbiology laboratory. There were 20 experimental batches of which 9 batches contained only salt solutions while the other 11 batches contained salt solutions, nutrient and bacteria (PF). Each batch was monitored for more than 3 weeks to ensure that equilibrium had been reached. At a given time, one tube of sample from each batch of mixed solution was removed for water quality analysis and pH measurement; the solution was discarded after the analytical tests were conducted. The water quality was determined by measurement of Table 1 Salt concentrations of CaSO4 and NaCl at different water recoveries and CP values. Recovery
0%
35% (CP = 1)
50% (CP = 1)
35% (CP = 2)
50% (CP = 2)
NaCl (M) CaSO4 (M)
0.5 0.03
0.75 0.05
1.0 0.06
1.5 0.1
2.0 0.12
Table 2 Experimental conditions at 25 °C for the tests. Batch no.
NaCl (M)
CaSO4 (M)
Nutrient (ppm)
PF (× 106 cfu/ml)
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
0.5 0.5 0.5 1.0 1.0 1.0 1.5 1.5 1.5 1.0 1.0 1.0 1.0 1.0 0.5 0.5 0.5 1.5 1.5 1.5
0.065 0.080 0.100 0.065 0.080 0.100 0.065 0.080 0.100 0.060 0.060 0.060 0.080 0.100 0.060 0.080 0.100 0.060 0.080 0.100
– – – – – – – – – 5 5 5 5 5 5 5 5 5 5 5
– – – – – – – – – 1 10 20 10 10 10 10 10 10 10 10
sodium (Na+), calcium (Ca+ 2) and sulfate (SO− 4 ) ions using ICP-AES (Inductively Coupled Plasma-Atomic Emission Spectroscopy). Each sample was filtered with a 0.22 μm filter and further diluted with micro-filtered water before ICP analysis so that the salts were below the solubility limit as to avoid precipitation. The precipitates and extracellular polymeric substance that adhered on the surface of membrane at the end of each experimental batch were analyzed using Scanning Electron Microscopy (SEM) to examine their surface structure and porosity. Electron Dispersion X-ray (EDX) Spectroscopy was used to determine chemical constituents in the precipitates and extra-cellular polymeric substance for their elemental composition identification and quantification. The crystallographic structure of precipitates was characterized and identified using X-Ray Diffraction (XRD). The analysis of precipitate formed in the runs containing biological matter needed specific preparation as discussed here. Solution of 2% by volume of Osmium Tetraoxide (OsO4) was prepared and the sample was soaked in it for an hour. The soaking was for fixation of the structure of sample. Following that the sample was rinsed with distilled water 3 times to remove any remains of Osmium Tetraoxide. The sample then was dehydrated with stagewise concentrations of ethanol (starting from 30%v/v, followed by 50%v/v, 75%v/v, 95%v/v and gradually increased to 100% v/v). Each dehydration stage took 15 min except for the last two stages that required 30 min for each. Then samples underwent critical point drying (CPD) using Bal-tec 030 Critical Point Dryer and liquid CO2 to replace the ethanol content. 3. Results and discussion The accuracy and reproducibility of results were assessed before any attempt in data reduction. Experimental error analysis was conducted for preparation of samples and all sample taking and measurements. Fig. 1 represents an example of repeated tests showing a very good reproducibility; the maximum uncertainty for the repeated tests was 7%. 3.1. Thermodynamics analysis The activity coefficients of ionic species in complex seawater solution are calculated by the Pitzer model which is applicable to ionic strength above 0.01 M and up to 6 M. Details are provided elsewhere [24]. General thermodynamic relations are available elsewhere [4]. The instantaneous Ion Activity Product (IAP) of CaSO4 in the experimental run was calculated from the measurement of Na+,
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Fig. 1. A Sample of reproducibility test.
Ca2+ and SO2− concentration in the solution using the Pitzer model 4 and plotted as a function of time. All samples in the experimental run were measured until equilibrium was reached. The IAP value obtained at equilibrium is the Thermodynamic Solubility Constant (Ksp) of the salts in solution. Summary of the thermodynamic and kinetic calculations are listed in Table 3. Presence of PF had affected, as will be discussed later, the precipitation thermodynamics and kinetics and had rendered the calculation of their constants not possible and as such only the results for the runs without bacteria are reported in Table 3 where n is order of the reaction and S.S. is the Super Saturation. Results from Table 3 show that the average Ksp value for CaSO4 was 2.49 ± 0.4 × 10− 5 (mol/l)2 which was comparable with the published data for CaSO4 (2.52 × 10− 5 (mol/l)2) at 25 °C [25] and the average of 2.43 ± 0.7 × 10− 5 (mol/l)2 [24]. Fig. 2 shows that for 0.5 M NaCl solution the initial drop in instantaneous IAP values of CaSO4 versus time was the same for the three solutions with varying CaSO4 concentrations. However, as the concentration of the NaCl increased to 1 M and 1.5 M, there was a clear difference in the initial instantaneous IAP drop for various CaSO4 concentrations. These results indicated that the dependence on the actual degree of supersaturation, which is salinity dependent, becomes important at salinity levels higher than 1 M. The initial supersaturation of solution decreases as NaCl concentration increases. For a given calcium sulfate concentration, the instantaneous IAP as a function of time is plotted in Fig. 3 for various salinity levels. At higher salinity level of 1 M, there was less difference in the instantaneous IAP decline versus time for various calcium sulfate concentrations. The effect of salinity on instantaneous IAP was more pronounced for lower calcium sulfate concentrations. In general, the variations were more pronounced at lower solution supersaturation levels, being higher salinities and lower calcium sulfate concentrations. At high supersaturation levels spontaneous and homogeneous nucleation occurs rendering the effect of other factors much less pronounced.
3.2. Induction periods Supersaturation is a pre-requisite for nucleation to occur which would further lead to crystallization. A supersaturated solution is
Table 3 Summary of results for pure CaSO4 at various salinities. Batch no.
NaCl (M)
CaSO4 (M)
n
1 2 3 4 5 6 7 8 9
0.5
0.065 0.080 0.100 0.065 0.080 0.100 0.065 0.080 0.100
4.03
1
1.5
Krxn (mol/l∙h) 0.0021
6.18 0.0062 7.29 0.0067
Ksp (CaSO4) (mol/l)2
S.S. (initial)
Induction time (h)
2.13E-05 2.21E-05 2.88E-05 2.49E-05 2.94E-05 2.63E-05 2.53E-05 2.49E-05 2.12E-05
4.4 5.7 7.6 2.6 2.9 6.0 2.0 3.6 5.8
5.00 0.17 0.00 24.00 1.03 0.00 48.00 4.00 0.00
Fig. 2. Instantaneous IAP for a given salinity at various CaSO4 concentrations.
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Fig. 4. a. Precipitation of CaSO4 in presence of NaCl as monitored by reduction of Ca2+ concentration. b. Precipitation of CaSO4 in presence of NaCl as monitored by reduction concentration. of SO2− 4
concentrations increase to 0.08 M and 0.1 M. At the highest calcium sulfate concentration, the nucleation is spontaneous and induction time is independent of salinity level. These results are in good agreements with those previously reported by Sheikholeslami and Ong [24]. At lower concentration levels needle-liked star-shaped crystals were formed. As the calcium sulfate concentration increased, and solution had shorter induction periods, small and flaky crystals were formed; this was due to spontaneous precipitation which usually results in smaller crystal sizes in greater numbers. Fig. 3. Instantaneous IAP for a given CaSO4 concentration at various Salinities.
called a metastable solution when no nucleation occurs and solution remains supersaturated. The period of time in which the supersaturated solution remains in a metastable state before nucleation and crystallization occurs is called induction period. The induction period of salt precipitation is dependent upon the type of precipitating salt and its concentration. The experimental results have shown that presence of other soluble ionic species has an effect on the induction period of precipitating salt. The induction period of CaSO4 was examined by monitoring the ionic species in a non-precipitating reduction of both Ca2+ and SO2− 4 NaCl solution (Figs. (4a) and (4b)). Time and observations for crystallization were also recorded throughout the whole experimentation. Table 3 shows that the induction period for pure CaSO4 (0.065 M) increases significantly from 5 h to 48 h as concentration of NaCl increases from 0.5 M to1.5 M resulting in decreases in the degree of supersaturation due to the existence of non-precipitating NaCl salt and increases in the ionic strength of solution. The presence of nonprecipitating soluble salt changes the Gibbs free energy of nucleation reaction and hence affecting the surface energy of a crystal. The effect of ionic strength on induction time reduces as the calcium sulfate
3.3. Effect of PF The experimental results in Fig. 5 show a representative run in presence of PF; the concentration of S2− increased over a period of time in the presence of the bacteria. The same trends were obtained for runs at other calcium sulfate concentrations and salinities in presence of PF. The concentrations of Ca2+ and Na+, Cl− were monitored throughout the runs and the analytical results showed that they remained constant. The reason for the increase in S2− concentration was due to microbial activities. The results suggested there was generation of hydrogen sulfide (H2S) by the bacteria during microbial activity where reduction of sulfate (SO2− 4 ) occurred. Some samples were tested by ion chromatography to confirm this hypothesis. The results from ion chromatography analysis showed a 2− over a period of reduction of SO2− 4 concentration with increase in S time; the results for a representative run are plotted in Fig. 5 for the 0.1 M CaSO4 solution in 1 M NaCl with 10 × 106 cfu/ml PF. The same trends were obtained for other runs in presence of PF. The analytical results confirmed the hypothesis that the bacteria consumed sulfate as nutrient for their microbial activities and the reduction of sulfate further affects the metastability of the calcium sulfate in the solution. The reduction of sulfate to sulfide changed the
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6 Fig. 5. Concentration of S2− and SO2− 4 of 0.1 M CaSO4 in 1 M NaCl with 10 × 10 cfu/ml PF.
thermodynamic solubility of solution and prevented crystallization of CaSO4. Therefore no induction period, kinetic or thermodynamic analysis could be conducted for solutions that contained PF. Brownian diffusion, aggregation and sedimentation are responsible for the formation of biofilm in this work as the experiments were conducted in a batch system where there was no fluid flow. A period of time was required for the bacteria to arrive at the surface of a membrane, attach and colonize. The formation of extracelluar polymeric substance (EPS) was mostly observed after 96 h at low salinities. At higher salinities, the time period for formation of EPS decreased to 48 h. These results showed that high salinities promote faster formation of biofilm as suggested previously [15] and that higher ionic strength reduces the stability of bacteria in suspension and hence promotes faster biofouling. As stated above, no CaSO4 crystals were formed for all the experiments conducted in presence of PF irrespective of calcium sulfate concentrations or salinities in the solution. 3.4. Kinetics analysis Kinetic rates and order of reaction for pure CaSO4 precipitation at various salinities were analyzed using nth order initial rate method. In this study the solution salinities were at 0.5, 1, and 1.5 M. Therefore the kinetic analysis showing the effect of salinity is not comprehensive and the results are not conclusive. However, in light of our previous experimental studies also showing the effect of salinity on kinetics [24,26,27], the kinetic analysis is reported here. The kinetic rate order, n and rate constant, Krxn for CaSO4 at various salinities are summarized in Table 3. The results show that the order of reaction for CaSO4 precipitation was very much affected by the presence of NaCl. The presence of non-precipitating species increased the order of reaction rate (n); it was 4.03, 6.18 and 7.29, respectively, for 0.5 M, 1 M and 1.5 M NaCl solutions. Results shown in Fig. 6 suggest that CaSO4 precipitation in NaCl solution was a poly-nuclear reaction. Extrapolation of the order reaction (n) curve to 0 M NaCl concentration results in a value of approximately 2 for the order of reaction (n).
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Thus, it is in good agreement with the literature value obtained by Liu and Nancollas [28] indicating that CaSO4 dihydrate (gypsum) precipitation in stable supersaturated solution at a temperature between 15 °C and 45 °C followed a 2nd order rate equation with respect to depletion of total Ca2+ [28]. However, the Krxn values obtained in this work cannot be used for comparison with theirs [28] as this work was conducted without the aid of seeds for crystal growth and surface area was unknown whereas their experiments were conducted using a seeded crystal growth technique and known surface area of seeds. Primary homogeneous nucleation and spontaneous precipitation was involved in this work and hence the Gibbs free energy of reaction would have been more than that required for the work of Liu and Nancollas where a secondary heterogeneous nucleation occurred. Table 3 shows that the reaction rate constant, Krxn, values increased from 0.021 to 0.067 mol/l∙h as concentrations of NaCl increased from 0.5 M to 1.5 M. Although the reaction rate increased with concentrations of NaCl, it could not be used for direct comparison with the results of Liu and Nancollas [28] since the growth area and number of nucleus to initiate precipitation are unknown in this work. The overall rate of reaction is considered to be faster at higher salinities. This was in good agreement with previous results of Sheikholeslami and Ong [24] where the same trend was observed and the order of reaction rate increased with increases in salinity values. As stated before, kinetic analysis could not be conducted for the runs in presence of PF due to absence of CaSO4 precipitation. 3.5. pH analysis The pH for seawater is around 8 due to contribution of major bicarbonate species. The pH of all the experimental batch tests was monitored and measured to be within the range of 7 to 8 throughout the experiment. Comparison of pH change over time for the tests with pure CaSO4 (Fig. 7a) and those in presence of PF (Fig. 7b) at 1 M NaCl depicts the effect of bacteria on solution pH. The pH for 0.065 M CaSO4 in 1 M NaCl during the run remained within the pH range of 7 to 7.4 while the pH for CaSO4 solutions in the presence of PF in1 M NaCl decreased from about 7.4 to 6.3; there was a significant change of pH under the influence of bacteria and it further depicts influence of biological fouling on inorganic fouling as most inorganic salt crystallizations are pH dependent. The drop in pH over a period of time was due to microbial activities by production of CO2 as a result of respiration and by reduction of sulfate to H2S. The formation of hydrogen sulfide could be due to anaerobic condition created, after depletion of limited dissolved oxygen in the solution, in the experiment as there was no air/gap in the constant volume batch reactors. After depletion of the dissolved oxygen, bacteria could reduce sulfate to sulfide by using the sulfate ions as a terminal electron acceptor in respiration and so lead to the reduction of sulfate. The hydrogen sulfide (H2S) produced would dissociate into HS− ions; dissociation of HS− ions into elemental sulfur and H+ ions in the solution contributed to the increase of S2− concentration and lowering the pH. A look at Fig. 7b shows that the final solution pH was lowest for the lowest microorganism concentration; however, this does not suggest that the maximum pH drop was for this run. Fig. 7b shows that the initial pH values were about 7.45, 7.25, and 7.08 for, respectively, runs with highest to lowest microorganism concentrations. Therefore, the actual pH drop was highest for the run containing highest microorganism concentration and was lowest for the run containing lowest microorganism concentration. 3.6. Scale structure and morphology
Fig. 6. Rate constant and order of reaction for pure CaSO4 at various salinities.
Figs. 8–10 show the effect of calcium sulfate concentration on the scale morphology. The results shown are for tests conducted at 1 M
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Fig. 9. SEM images of 0.080 M pure CaSO4 in 1 M NaCl.
salinity; similar trends were observed for other salinities. Fig. 8 shows that at low concentration of CaSO4 (0.065 M), the crystals exhibited long and fine needle-like structure which is typical of calcium sulfate dihydrate (CaSO4.2H2O) or gypsum. As concentration of CaSO4 increased to 0.1 M (Fig. 10), the crystal structure was altered; however, the phase did not change and remained as gypsum based on the XRD results. At a higher concentration of CaSO4 (Fig. 10), the crystals appeared to be fine white powder that looked ‘flaky and flowery’; this was due to spontaneous precipitation in the highly supersaturated solution where crystals were formed instantaneously with no induction time for them to develop and crystallization was more of a result of nucleation than as a result of crystal growth. These findings were in good agreement with the scale morphology studies by Sheikholeslami and Ong [24]. It was observed that as salinity increased from 0.5 M to1.5 M, the size of CaSO4 crystals became larger. The formation of larger sized
crystals was a result of the higher ionic strength solution and the longer induction period where crystallization was more due to crystal growth than nucleation. Therefore, in general the crystal size was affected by the degree of supersaturation of solution. As supersaturation decreases, the crystal sizes become larger and vice versa; overall, a longer induction period favors crystal growth and larger crystals while a shorter induction period favors nucleation and smaller crystal sizes. During the experiment, it was observed that some crystals were loosely adhered to the surface of the test-tubes while others were floating freely in the solution. The CaSO4 crystals that had adhered on the test-tube surface were not tenacious and could be easily removed by scrubbing with water. Also the crystals that had adhered on the membrane fell off easily during coating for SEM analysis and hence the crystals were analyzed without membranes. Figs. 11–13 show the SEM images of EPS (Extracellular Polymeric Substance) formed by PF in presence of CaSO4 in NaCl solutions. The images do not show any differences in the morphologies of EPS formed in terms of bacteria concentration. The structure of biofilms formed by PF exhibited cross-link cellular fibers interlinking with one another. The SEM images show that PF of approximately 1.4 μm size was firmly attached onto the EPS fibers. At higher magnifications, the bacteria appeared to be elongated and rod shaped. The EDX analysis showed that there were no salt crystals embedded within the EPS. During the experiment, it was observed that the amount of EPS formed was dependent on the concentration of bacteria present; as expected, the thickness of biofilms increased when bacteria concentrations increased from 1 × 106 cfu/ml to 20 × 106 cfu/ml due to enhanced microbial activity. It was also observed that Ca2+ presence
Fig. 8. SEM images of 0.065 M pure CaSO4 in 1 M NaCl.
Fig. 10. SEM images of 0.1 M pure CaSO4 in 1 M NaCl.
Fig. 7. a. pH versus time for pure 0.065 M CaSO4 at 1 M NaCl. b. pH versus time for pure 0.06 M CaSO4 in presence of PF at 1 M NaCl.
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Fig. 11. 0.06 M CaSO4 in 1 M NaCl with 1 × 106 cfu/ml PF at 400 and 4000 magnification.
Fig. 12. 0.06 M CaSO4 in 1 M NaCl with 10 × 106 cfu/ml PF at 400 and 4000 magnifications.
had some effect on the adhesiveness of biofilm; as the Ca2+ concentration increased, the biofilm adhered to the membrane surface and bottom surface of the test-tube. They later broke off from the
main substratum and formed individual colonies within the solution and also adhered on the wall surface of test tubes after a period of time.
Fig. 13. 0.06 M CaSO4 in 1 M NaCl with 20 × 106 cfu/ml PF at 400 and 4000 magnification.
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Fig. 14. EDX of crystals from 0.065 M CaSO4 in 0.5 M NaCl.
Fig. 16. EDX of crystals from 0.08 M CaSO4 in 1 M NaCl.
3.7. Electron Dispersion X-ray analysis
4. Conclusions
The CaSO4 crystals were further analyzed on their elemental composition using Electron Dispersion X-ray (EDX) spectroscopy. The EDX results showed that there was no difference in the elemental composition of precipitates; the major components present in the crystals were calcium, sulfur and oxygen at low salinities. Higher quantities of calcium, sulfur and oxygen were detected as concentration of CaSO4 increased. The results showed that relatively high purity of CaSO4 crystals were obtained at low salinities. Figs. 14 and 15 show the EDX spectrums of 0.06 M and 0.1 M CaSO4 in 0.5 M NaCl, respectively. As salinities increased, relatively high quantities of sodium and chloride were detected by the EDX analysis revealing that during precipitation some sodium and chloride ions were trapped within calcium sulfate thus lowering the purity of CaSO4 crystals formed. This suggested that salinities not only affect the induction period of CaSO4 precipitation but also the purity of CaSO4 crystals formed. Figs. 16 and 17 show the EDX spectrums of 0.08 M CaSO4 in 1 M NaCl and 1.5 M NaCl, respectively.
Batch tests were carried out to study the thermodynamics and kinetics of calcium sulfate under the influence of various concentrations of PF at various salinities ranging from 0.5 M to 1.5 M at isothermal conditions of 25 °C. The Pitzer model was used to determine the ion activity coefficients as discussed elsewhere [24]. The increase in the concentration of non-precipitating NaCl salt did not affect the thermodynamic solubility product (Ksp, ion activity product at equilibrium) of CaSO4. Initial rate method was used to determine the kinetics for CaSO4 precipitation reaction. It was found that kinetics of CaSO4 was strongly affected by the salinity levels. As concentration of NaCl increased, the rate of reaction increased from 0.0021 to 0.0067 mol/l∙h. With increases in salinity the order of reaction increased from 4.03 to 7.29 indicating a poly-nuclear reaction; extrapolating the trend to zero salinity level resulted in the order of reaction of 2 which was in good agreement with values of Liu and Nancollas [28]. The increase in concentration of CaSO4 affected the scale morphology causing the crystals to become flaky and fine in comparison to crystals formed at lower concentration of CaSO4 which exhibited a needle-shaped structure. However, the crystal phase of CaSO4 remained as gypsum (calcium sulfate dihydrate) irrespective of salinity and concentrations levels. There was no crystallization of CaSO4 in the presence of PF. However, there was a significant decline in the sulfate concentration in the solution with time. The solutions were tested for sulfide concentration and the results showed a progressively increasing sulfide concentration as the sulfate concentrations dropped over period of time. This reveals the reduction of sulfate by the bacteria. In presence of bacteria, there was also a significant drop in the pH of solutions from a range of pH 7.4 to pH 6.3 over time. The drop in pH is expected to have been caused by contribution of H+ ions from dissociation of H2S and also production of CO2 due to bacterial respiration.
3.8. XRD analysis Precipitates were examined by XRD analysis where angle of the incident beam is diffracted according to the crystal lattice spacing and hence identifying the crystal phase. The diffraction angle has the unit of Degree 2-Theta and is calculated using Bragg's Law (d = λ = sinðθÞ) where θ is the diffraction angle (Degree 2-Theta), λ is X-ray wavelength (1.5405 A for Cu tube), and d is lattice spacing. Different crystal phases have different lattice spacing of atoms and hence the angle of diffraction differs from one crystal phase to another. The XRD analysis was used to identify crystal phases and the results showed CaSO4∙2H2O (gypsum) to be the crystal phase for all precipitates. Neither salinity nor the calcium sulfate concentrations affected the crystal phase; though they did affect the crystal size and shape.
Fig. 15. EDX of crystals from 0.1 M CaSO4 in 0.5 M NaCl.
Fig. 17. EDX of crystals from 0.08 M CaSO4 in 1.5 M NaCl.
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