Environ. Sci. Techno/. 1995,29, 2280-2285
Removal of Cadmium, Lead, and Zinc from Soil by a Rhamnolipid DAVID C. H E R M A N , J A N I C K F . ARTIOLA, A N D RAINA M. MILLER* Department of Soil and Water Science, University of Arizona, Tucson, Arizona 85721
Complexation of cadmium, lead, and zinc (singly and in a mixture) by a monorhamnolipid biosurfactant produced by Pseudomonas aeruginosa ATCC 9027 was studied in batch solution and soil experiments. Conditional stability constants (log KL) for metalrhamnolipid complexation in a buffered medium (0.1 M Pipes, pH 6.8) were determined in duplicate using an ion-exchange technique and averaged 6.5 (Cd2+), 6.6 (Pb2"), and 5.4 (Zn2+); these values are similar or slightly higher than literature values for Cd2+ and Pb2+ complexation with fulvic acid and activated sludge solids. To determine the ability of rhamnolipid to desorb soil-bound metals, rhamnolipid solutions (12.5, 25,50, and 80 m M ) were added to soil containing sorbed Cd2+ (1 -46 mmol kg-I), Pb2+ (1 -96 mmol kg-'), or a mixture of Pb2+-Cd2+-Zn2+ (3.4 mmol kg-'). At 12.5 and 25 m M rhamnolipid, rhamnolipid sorption to soil exceeded 78%, and less than 11% of soilbound Cd2+ and Zn2+ was desorbed. However, ion exchange of bound metals with K' present in the rhamnolipid matrix could account for the removal of between 16 and 48% of the sorbed Cd2+ and Zn2+. At 50 and 80 m M rhamnolipid, rhamnolipid sorption to soil decreased to between 20 and 77%, and the removal of Cd2+ and Zn2' could exceed the removal by ion exchange by as much as 3-fold. The behavior of Pb2+ was quite different. Less than 2% of soil-bound Pb2+ was desorbed due to ion exchange, although up to 43% was desorbed by 80 m M rhamnolipid.
Introduction Previous work demonstrated efficient cadmium complexation in solution by an anionic biosurfactant, rhamnolipid, that is produced by Pseudomonas aeruginosa ATCC 9027 (1). Although the use of bacteria or bacterial exopolymers for complexation of metals from waste streams has been extensively studied and reviewed (2- 7),the application of smaller biomolecules for metal complexation is of special interest for application to in situ soil washing and pump* Corresponding author: Phone: (520) 621-7231; Fax: (520) 6211647; E-mail address: RMMiller@CCIT. ARIZONA. EDU.
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ENVIRONMENTAL SCIENCE & TECHNOLOGY / VOL. 29, NO. 9,1995
and-treat remediation technologies (8). In these technologies, the size of the metal-ligand complexis crucial to avoid filtration of metal complexes by small soil pores. Ideally, such complexes should be less than 50 nm in diameter since filtration has been shown to have little effect on particles of this size,such as viral particles and microspheres (9). As reported by Champion et al. (101, rhamnolipid aggregates are predominantly small vesicles, < 50 nm in diameter, and micelles (5 nm in diameter) at pH > 6.0. Addition of cadmium at pH 6.8 was not found to alter rhamnolipid aggregate sizes appreciably, suggesting that rhamnolipid-cadmium complexes would not be removed by filtration. Biosurfactants, such as rhamnolipids, are of particular interest for use in remediation technologies for several reasons: (a) they are naturally occurring products and thus can be considered as "biodegradable" additives, (b) they are produced in great structural variety by many different microorganisms and may have unique metal binding capacities and selectivities in comparison to available synthetic chelates or surfactants, and (c) there is potential for in situ production of biosurfactants. Little is yet known about conditions that stimulate in situ production of surfactants in soils; however, Oberbremer et al. (11) have reported a spontaneous induction of surfactant production during biodegradation of hydrocarbons in soil columns suggesting that in situ biosurfactant production can occur. The objective of this study was to determine biosurfactant selectivity for metals, both singly and in mixtures, as well as to investigate biosurfactant-facilitated desorption of metals from soil. The biosurfactant used in this study was a monorhamnolipid acid produced by Pseudomonas aeruginosa ATCC 9027 (12, 13). The monorhamnolipid used (Figure 1) is characterized by a critical micelle concentration (cmc) of 0.1 mM (15) and a pK, of 5.6 (16). The metals used in this study were cadmium, lead, and zinc. A sandy loam soil with 0.11% total organic carbon was used for all metal desorption studies.
Materials and Methods Soil. The soil used in this study was a Hayhook sandy loam collected at 25-50 cm depth from the C horizon of a Hayhook soil, a coarse-loamy, mixed, non-acid thermic typic torriorthent. The soil was air-dried and sieved through a no. 10 standard sieve (2 mm openings). Selected characteristics of this soil were as follows: particle size distribution, 86% sand, 4.7% silt, 8.8% clay; pH 7.5, as determined after extraction with an equal volume of water; total organic carbon, 0.11%; cation-exchange capacity, 7.04 mequiv 1OOg-I; iron oxides (Fe20d,0.19%. Metals. Cd(N03)2,Pb(NO&, and Zn(SO& used in this studywere obtained fromadrich (Milwaukee,W). Atomic absorption (AA)standard solutions for each metal were obtained from Fisher (Pittsburgh, PA). Concentrated metal stock solutions (usually 10 mM) were prepared in 1%"03 and then diluted to the working concentration, as described below. Biosurfactant Production. Pseudomonas aeruginosa ATCC 9027 was obtained from the American Type Culture Collection (Rockville,MD). Monorhamnolipid production (12)and purification using Silica Gel 60 column chroma-
0013-936X/95/0929-2280$09.00/0
D 1995 American Chemical Society
HO -0 W O C H - CH2COO-CH-CH2COOH I
Ho OH
I
(pm(C?"4)n CH3
CH3
FIGURE 1. Structure of monorhamnolipid produced by Pseudomonas aeruginosa ATCC 9027. The predominant form of monorhamnolipid was m and n = 6.
tography have been described previously (13). Monorhamnolipid concentration was estimated using two techniques, surface tension measurement and L-rhamnose measurement. Surface tension was measured using a Sensadyne 6000 tensiometer (Chem-dyneResearch Corp., Milwaukee, WI). An alternative surface tension measure was made using a surface tensiomat, Model 21 (Fisher Scientific), which employs the Du Nouy ring method of quantifying surface tension. In each case, a calibration curve relating surface tension (dynlcm)to rhamnolipid concentration (mg L-I) was prepared. Rhamnolipid concentration was also quantified by measuring L-rhamnose using the 6-deoxyhexose method (13). Determination of Biosurfactant-Metal Complexation in Solution. All metal complexation experiments were performed in 0.1 M Pipes [piperazine-N,N-bis(2ethanesulfonic acid)]buffer solutions (Sigma, St. Louis, MO) with pH adjusted to 6.8 using 1 M NaOH. A cation-exchange technique described by Cheng et al. (17) and Sterritt (18) was adapted for use in the determination of conditional stability constants to describe the rhamnolipid-metal complexation for cadmium, lead, and zinc. A cationexchange resin (SP Sephadex (2-25) was obtained from Pharmacia Biotech AB (Uppsala, Sweden),washed several times in distilled water, and then allowed to expand overnight in distilled water. The resin was washed several times in 0.1 M Pipes buffer and then diluted to 9 mg mL-l using Pipes buffer solution containing varying concentrations of rhamnolipid (0-4.2 mM). For each experiment, resin-rhamnolipid mixtures (14.5mL) were transferred into triplicate 20-mL scintillation vials, and a 0.5-mL aliquot of a concentrated metal solution (1.5 mM cadmium, lead, or zinc in 0.1 M Pipes buffer) was added to give a final metal concentration of 0.05 mM. The vials were then sealed and shaken at 100 rpm for a minimum of 2 h at room temperature on a rotary shaker. The ion-exchange resin was allowed to settle for at least 10 min, and a 5-mL supernatant sample was removed for atomic absorption analysis to determine metal concentration in the aqueous phase. Atomic absorption (AA) analyses were performed on an Instrument Laboratory Video 12 aalae spectrophotometer (Allied Analytical Systems, Waltham, MA). A calibration curve for each metal was constructed using AA standard solutions that were diluted into 1%HN03. Sorption of Rhamnolipidby Hayhook Soil. To measure rhamnolipid sorption, 2.5 g of Hayhook soil was placed into 40-mL polypropylene centrifuge bottles and conditioned in 5 mL of 0.1 M KNOB(pH 6.8). After conditioning for 2 days at room temperature on a shaker (100 rpm), the bottles were centrifuged (40000g, 20 min), and the supernatant was removed. The remaining soil pellets were autoclaved once to inhibit biodegradation of rhamnolipid, and then soil pellets from triplicate bottles were suspended in 5 mL of 0.1 M KN03 containing 0, 12.5, 25, 50, and 80 mM rhamnolipid. Control bottles contained the same rhamnolipid solutions without added soil. The bottles were
incubated for 3 days at room temperature at 100 rpm and then centrifuged; rhamnolipid concentration in the supernatant was determined using surface tension analysis. Purified rhamnolipid is a weak acid, and when dissolved in water the pH was adjusted to 6.8 using 2 M KOH. The amount of K+ added as KOH and as K N 0 3 was used to estimate the ionic strength of the rhamnolipid solution. In the 12.5-80 mM rhamnolipid range, the K+ concentration present after pH adjustment was found to vary from 0.11 to 0.18 M, depending on the rhamnolipid concentration. Pipes buffer was not used in sorption experiments for two reasons: (a) use of a buffer in remediation would be prohibitively costly and (b) use of a buffer might alter the interaction between rhamnolipid and soil. Biosurfactant-Facilitated Desorption of Metals. For each experiment, soils were first loaded with metals. To do this, 2.5 g of Hayhook soil was placed into 40-mL centrifuge bottles, conditioned in 0.1 M K N 0 3 , and autoclaved as described above. The supernatant was then replaced with 5 mL of 0.1 M KNOBcontaining 1 mM either of Cd2+,Pb2+,or a metal mixture containing 1 mM each of Cd2+,Zn2+,and Pb2+. After 3 days of shaking, the tubes were centrifuged, and the supernatant was diluted into 1% "03 for AA analysis. Sorbed metal concentration was calculated from the difference between initial metal concentration and metal concentration in the supematant after sorption. The metal-containing soil was suspended in 5 mL of 0.1 M KNO3 containing 0, 12.5,25,50, or 80 mM rhamnolipid or in 5 mL of a reference (or control) solution, in triplicate. The reference solution was used to determine the potential for metal removal due to ion exchangewith cations present in the rhamnolipid solution matrix. This matrix was found to have a high ionic strength due to the need to neutralize (pH = 6.8)the purified rhamnolipid solution using a solution of 2 M KOH. For example, pH adjustment of the 80 mM rhamnolipid solution required the addition of 0.08 M of K+ (as KOH) to the rhamnolipid matrix. The resulting rhamnolipid matrixK+concentrations ranged between 0.11 and 0.18 M depending on the amount of rhamnolipid present. Therefore, each reference solution contained K+ (as KN03) at the same molar concentration of K+ (as KOH) present in each rhamnolipid treatment level. The centrifuge bottles containing soil mixed with rhamnolipid solution or the corresponding reference solution were shaken for 3 days and then centrifuged, and the supernatant was analyzed for metal concentration by AA analysis and for rhamnolipid concentration by surface tension measurement. The supernatant samples used for AA analysis were acidified to a pH < 2 using 0.1 mL of concentrated "03 and centrifuged to pellet the rhamnolipid, and then the supernatant was diluted in 1%HN03 for AA analysis. To ensure that all the metal was recovered from the rhamnolipid pellet, the pellet was washed a second time with 1%"03 and the supernatant was analyzed by A4 for metals.
Results Rhamnolipid-Metal Complexation Analysis. Rhamnolipid complexation of metals was investigated using an ionexchange technique, as described by Cheng et al. (17) and Sterritt (18). This technique is based on the equilibrium complexation of a metal with an organic ligand and of a metal with a cation-exchange resin, as described by the following equilibrium reactions: VOL. 29, NO. 9, 1995 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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where Mis the aqueous concentration of metal (mol L-I), L is the organic ligand (mol L-*), x is the number of ligands per complexmolecule (mol mol-'), MLx is the metal-ligand complex in solution (mol L-'1, R is the resin concentration (kg L-I), and MR is the metal bound to resin (mol kg-I). Using these equilibrium relationships, distribution coefficients between a metal and a resin in the absence and the presence of an organic ligand can be written as ,lo = MRlM
A=
MR M MLx
+
(? 1
(4)
ENVIRONMENTAL SCIENCE &TECHNOLOGY / VOL. 29. NO. 9, 1995
I
, 1
0
2 3 4 rhamnolipid (mM)
5
FIGURE 2. Partitioning of CdZ+between ion-exchange resin and the fluid phase as rhamnolipid concentration is increased from 0 to 4.2 mM. Each line represents a separate experiment, and each point represents the mean and standard deviation of triplicate samples. TABLE 1
Conditional Stability Constants for Rhamnolipid Complexation of Heavy Metals experiment 1
(5)
The values of x and KLcan be determined by graphing the linear function log (&/,I - 1) versus log L where L is rhamnolipid concentration. An example of the data required to derive KL and x is shown in Figure 2 for cadmium. This figure shows two separate experiments used to determine cadmium complexation with rhamnolipid. In the absence of rhamnolipid (0 mM), approximately 71% of the Cd2+was bound to the resin, leaving 29% in the aqueous phase. Addition of increasing concentrations of rhamnolipid caused increasing amounts of Cd2+to be present in the aqueous phase until at the highest concentration tested, 4.2 mM rhamnolipid, between 88%and 97%of the Cd2"was found in the aqueous phase. The aqueous phase was also analyzed using a cadmium ion-specific electrode (1)to distinguish between free Cdz+and rhamnolipid-complexed Cd2+. At 4.2 mM rhamnolipid, less than 2% of Cdz- was detected as free metal, indicating that almost all the Cd2- determined by AA analysis was complexed with rhamnolipid. The partitioning of rhamnolipid between the resin and the aqueous phase in the absence of metals was also determined. In this case, aqueous phase rhamnolipid concentrations were determined by surface tension and rhamnose assay. Data (not shown) indicated that there was no binding between rhamnolipid and the ion-exchange resin. Using the data shown for cadmium in Figure 2 and similar data for lead and zinc (not shown),KL and x values for rhamnolipid complexation were determined and are compared in Table 1. These data indicated that rhamnolipid had a similar capacity to complex the three metals tested with a slight preference for binding lead and cadmium over zinc. KL values appeared to be sensitive to slight variations in observed complexation. Using cadmium as an example, rhamnolipid complexation behavior was almost identical between the first and second experiments (Figure2);however, the calculated stabilityconstant varied 10-fold. The value of the rhamnolipid-metal complexation 2282
I
(3)
The distribution coefficients(eqs3 and41 can be determined experimentally, and a conditional stability constant (KL) can be obtained using the relationship
log - - 1 = l o g K , + x l o g L
1
metal
log
Cd2+
ZnZt Pb2+
Kt
6.0 5.0 6.5
experiment 2 X
log Kt
x
1.9 1.5 1.7
7.0 5.8 6.7
2.2 1.7 1.7
TABLE 2
Rhamnolipid Complexation of Individual Metals or Metals Present in a Mixturea % of metal in aqueous phase YO of metal in aqueous phase in the absence of rhamnolipid in the presence of rhamnolipidC
metal
individual metal*
CdZt 2 9 . 6 + 2.0
Zn2+ 34.9 f 0.5 PbZt
15.3 & 1.2
metal in mixture
individual metal
metal in mixture
32.3 f 3.7 36.6 i- 0.4 14.9 d~ 2.1
69.3i- 1.4 73.9 i 0 . 5 95.3 i- 0.6
57.6 It 1.4 5 9 . 4 f 0.9 73.2 i- 4.2
Rharnnolipid complexation is indicated by the transfer of resinbound metals into the aqueous phase in the presence of rharnnolipid. Mean (standard deviation) of triplicate samples. 2 rnM rharnnolipid. a
ratio (x)was also calculated and found to be approximately 2 mol of rhamnolipid/mol of metal in all cases (Table 1). Rhamnolipid Complexation of a Metal Mixture. The ion-exchange resin was also used to compare rhamnolipid complexation of a single metal with metals in a mixture. In the first stage of this experiment, Cd2+,Zn2-, and Pb2were exposed to ion-exchange resin either separately (0.1 mM concentration) or in a mixture (0.1mM of each metal). Results in Table 2 show that binding of each metal to the ion-exchange resin in the absence of rhamnolipid was the same whether metals were present separately or in a mixture. In the next stage of the experiment, 2 mM rhamnolipid was added, and the increase in metal concentration in the aqueous phase was determined. Rhamnolipid complexation of single metals followed the order Pb2+ (95.3% complexed) > Zn2+(73.9%)2 Cd2+(69.3%). Rhamnolipid complexation of metals in a mixture followed the same order of preference as for metals alone; however, the total amount of each metal complexed was reduced by 17-2396. This may be a result of competition for rhamnolipid binding sites since for single metals the metal
I
40000 1
35000
4
15000
i
10000
I log c u 0
12.5 25.0 50.0 80.0
-0
1 2 3 4 5 1 1
10000
20000
S
30000
c mg L-1 FIGURE 3. Sorption isotherm of rhamnolipid in Hayhook soil. S is the amount of rhamnolipid sorbed to soil (mg kg-'1, and C is the rharnnolipid remaining in the fluid phase (mg 1-l). The insert shows the isotherm fitted to the Freundlich equation. Each point is the mean of triplicate samples.
concentration was 0.1 mM and for metal mixtures the total metal concentration was 0.3 mM. Rhamnolipid Sorption to Hayhook Soil. The sorption of rhamnolipid to Hayhook soil was determined over a rhamnolipid concentration range of 12.5-80 mM. A nonlinear sorption isotherm was obtained (Figure31, which could be fitted with the Freundlich equation (eq 6): S = KfC'
60 40
1-
2o0
12.5 25.0 50.0 80.0
rhamnolipid mM FIGURE 4. Comparison of metal desorption from Hayhook soil by rhamnolipid (open bar) and a reference solution (shaded bar) containing between 0.11 and 0.18 M KNO3 depending on the treatment level. Soil was exposed to each metal separately. Each bar represents the mean and standard deviation of triplicate samples.
60
(6)
where S is the amount of rhamnolipid associated with the soil (mg kg-'1, C is the amount of rhamnolipid remaining in solution at equilibrium (mg L-l), Kf is the Freundlich constant, and n is a measure of nonlinearity. The fitted constants were Kf = 6761, n = 0.17, and 6 = 0.96. Rhamnolipid Desorption of Metals from Soil. The first stage of this experiment was to load the Hayhook soil with metal. After exposure of the soil to a 1 mM concentration of two single metals, Cd2+and Pb2+,the amount of sorbed metal was determined. Results showed that 73% of the Cd2+ (1.46 mmol kg-l) and 98% of the Pb2+ (1.96 mmol kg-') was sorbed by the soil. The second stage of this experiment was to determine desorption of soil-bound metals byrhamnolipid solutions. Desorptionof both metals was found to be dependent on the amount of rhamnolipid added (Figure 4). For example, only 2.2% of the Cd2+was removed by a 12.5 mM rhamnolipid solution, but 55.9%of the Cd2+was removed by a 80 mM rhamnolipid solution. Removal of metals by the reference solution, which contained the same molar concentration of K+ as the rhamnolipid solution, showed that in all cases between 15.6% and 18.8% of soil-bound Cd2+was desorbed. In comparison, it was found that distilled water alone removed only 2% of soil-bound Cd2+ (data not shown). Cd2+ desorption from soil at the lower rhamnolipid concentrations (12.5and25 mM) was 2-10-foldless than the reference solutions. At the higher rhamnolipid concentrations (50 and 80 mM), Cd2+desorption was approximately 3-fold greater than the corresponding reference solutions. In contrast to Cd2-, less than 1% of soil-bound Pb2+ was desorbed by the reference solution. The 50 and 80 mM rhamnolipid treatments removed 27.5%and 41.6%of soilbound Pb2-, respectively. Rhamnolipid mobilizationof soil-bound metals was also examined using a mixture of Cd2+,ZnZc,and Pb2+(1 mM each metal) (Figure 5). When applied as a mixture,
12.5 25.0 50.0 80.0
U
d 20 12.5 25.0 50.0 80.0 100
60
40 20 '
0
1 12.5 25.0 50.0 80.0
rhamnolipid mM FIGURE 5. Comparison of metal desorption from Hayhook soil by rhamnolipid (open bar) and a reference solution (shaded bar). Soil was exposed to metals in a mixture. Each bar represents the mean and standard deviation of triplicate samples.
approximately 94% of Pb2+,44% of Cd2-, and 32% of Zn2+ (a total of 3.4 mmol kg-l) was bound to the soil. Exposure of the metal-laden soil to rhamnolipid solutions resulted in a concentration-dependent removal of the metals. Similar to results for single metals, ion-exchange processes accounted for all or a portion of the Cd2+and Zn2' desorbed from soil. Only at 80 mM rhamnolipid did Cd2+or Zn2' desorption exceed that of the reference solution. Ion exchange accounted for only 2% of the desorption of soilbound Pb2+, although up to 43% was desorbed with rhamnolipid treatment (Figure 5). To determine the rhamnolipid availability for metal complexation, rhamnolipid concentrations in sample supernatants were determined by surface tension analysis. As expected, supernatant rhamnolipid concentration inVOL. 29,NO. 9, 1995 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 3
Influence of Metals on Rharnnolipid Partitioning between Solid and Aqueous Phase in Hayhook Soil rhamnolipid (mMIa 12.5 25 50 80
YO rhamnolipid in aqueous phaseb no metal
CdZ+
PbZ+
metal mix
0.9 f 0.1 0.0f 0 0.9 i 0.1 0.0f 0 3 . 0 i 0.8 13.5 f 3 . 5 2.4 i 1.0 21.5 f 8 . 1 36.6 i 7.9 63.3 f 2.3 23.0 i 1.9 71.7 f 6.3 61.4 f 4.1 68.9 f 2.9 53.4 i 3.9 79.8 f 2.9
a Concentration of rhamnolipid added to soil. Mean & standard deviation of triplicate samples.
creased as the amount of rhamnolipid added was increased (Table 3). However, the amount of rhamnolipid sorbed was affected by the presence of metals in the soil. Sorption data were similar when either no metal or just Pb2' was present in the soil. However, when either Cd2+or a mixture of Cd2+,Pb2+,andZn2+was present, sorption of rhamnolipid was decreased, resulting in higher aqueous phase concentrations of rhamnolipid. Further, the amount of rhamnolipid sorption seemed to depend on the amount of metal in the soil. While sorption of rhamnolipid was decreased in the presence of Cd2- (1.46mmol kg-l), it was decreased even further in the presence of the metal mixture (3.4mmol kg-I) with a larger amount of metal sorbed. In most cases, sorption of rhamnolipid by the metal-treated soil was less than sorption by the untreated soil.
Discussion Previously, rhamnolipid complexation of cadmium in solution was demonstrated using an ion-selective electrode to measure Cd2+concentrations ( I ) . In this study, an ionexchange technique was used to demonstrate rhamnolipid complexation of other cationic metals alone and in mixtures. Conditional stability constants (KL) for rhamnolipid-metal complexation were determined in two separate experiments, and the results for Cd2+(6.5) and Pb2+ (6.6) were similar or slightly higher than conditional complexation constants reported for Cd2' and Pb2- binding to fulvic acid (19)and to activated sludge solids (20). Stability constants indicated a binding selectivity for the single metals tested in the order Pb E Cd > Zn (Table 11, and data from complexation of metal mixtures seemed to show a similar selectivity,Pb > Cd szZn (Table 2). All metals had a similar complexation ratio (x)of approximately 2 mol of rhamnolipid/mol of metal. Since rhamnolipid is a monovalent anion and the metals studied are predominantly divalent cations, this complexation ratio seems reasonable. Rhamnolipid biosurfactants were found to be strongly sorbed to Hayhook soil. At rhamnolipid concentrations less than 50 mM, sorption to soil was greater than 70% of the total rhamnolipid added. There are two possible mechanisms for rhamnolipid sorption. The first is cation bridging between the anionic polar head group and sorbed cations on soil, and the second is hydrophobic interactions between the nonpolar tails and hydrophobic regions in the soil organic matter. Since the total organic carbon content of the Hayhook soil used in this study was low (0.1l%l, we considered cation bridging to be the most likely sorption mechanism. It was therefore of concern that the metaldesorption experiments reported in this study were performed at relatively high concentrations of potassium cation (0.11-0.18 M), which could potentially affect rhamnolipid 2284
ENVIRONMENTAL SCIENCE &TECHNOLOGY / VOL. 29, NO. 9,1995
sorption via cation bridging. An initial experiment was conducted to determine whether K+ played a role in the sorption of rhamnolipid to soil. An 80 mM rhamnolipid solution was dialyzed against distilled water using a 500 MW cutoff membrane. Dialysis was used to reduce the amount of K' and any other cations in the rhamnolipid matrix while retaining rhamnolipid. Monorhamnolipid monomers have a molecular weight of 504 (141, although at concentrations above the critical micelle concentration (cmc),an equilibrium is established favoring the formation of micelles, which would have a greater molecular weight (10)and would be held in the retentate. Following dialysis, it was found that the rhamnolipid concentration in the retentate had decreased by 50%,which may have occurred due to diffusion of rhamnolipid monomers through the dialysis membrane or due to sorption of rhamnolipid by the dialysis membrane. Soil sorption of the dialyzed rhamnolipid solution matrix was found to be much less than nondialyzed rhamnolipid solutions. Greater than 95% of the rhamnolipid in a dialyzed solution matrix was found to remain in the supernatant, while the amount of rhamnolipid in the supernatant for comparable nondialyzed rhamnolipid solutions never exceeded 63%. These results support the conclusion that cation bridging in high ionic strength soil solutions is responsible for rhamnolipid sorption onto soil particles. The presence of metals in the soil prior to addition of the rhamnolipid was also found to affect the sorption of rhamnolipid (Table 3). The presence of soil-bound Cd2(1.46 mmol kg-I) or the mixture of Cd2', Pb2', and Zn2(3.40 mmol kg-') resulted in less rhamnolipid sorption to soil than was evident in the absence of any metal. In contrast, Pb2* (1.96 mmol kg-l) had no affect on rhamnolipid sorption to soil. Due to the strong sorption of rhamnolipid by soil, high levels of rhamnolipid treatment were required in order to successfully mobilize soil-bound metals. At the lower rhamnolipid concentrations tested (12.5 and 25 mM), rhamnolipid sorption to soil was high (between 100 and 78%sorbed) and appeared to limit the ability of rhamnolipid to desorb soil-bound metals. At higher rhamnolipid concentrations (50 and 80 mM), sorption was reduced to between 77 and 20%. But higher rhamnolipid concentrations also meant high K- concentrations within the rhamnolipid matrix and, therefore, the predominance of ionexchange processes. Soil-bound Cd2- and Zn2*were found to be sensitive to K' concentrations within the rhamnolipid matrix, and a large percentage of the metals mobilized from soil could be attributed to ion-exchange processes. In contrast to Cd2- and Zn2+,soil-bound Pb2+appeared to be insensitive to ion-exchange effects, and more than 40% of the soil-bound Pb2- was mobilized by the 80 mM rhamnolipid treatment. Lead is a soft Lewis acid, will sorb more strongly to soils, and will form more stable complexes with ligands than either cadmium or zinc ions. Zinc, which is considered a hard Lewis acid, and cadmium, which is borderline between soft and hard, are more likely to be affected by changes in ionic strength (21). Therefore the stability constants and soil sorptions shown in Tables 1 and 2 follow expected trends. Rhamnolipids may be particularly effective in soils contaminated with metals that are less sensitive to ionexchange processes. Ultimately, in application of rhamnolipid to metal-contaminated soils, the efficiency of rhamnolipid treatment must be improved. To do this,
rhamnolipid sorption must be minimized. Workin progress will examine more closely the mechanism of rhamnolipid sorption and the role of cation bridging in rhamnolipid sorption.
Acknowledgments Our thanks to Dr. David Hendricks for his help with AA analyses. The project described was supported in part by Grant 57-CH-197 from the U. S.-Mexico Foundation for Science and in part by Grant E504940 from the National Institute of Environmental Health Sciences, NIH.
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Received for review December 14, 1994. Revised manuscript received May 17, 1995. Accepted J u n e 5, 1995.@ ES9407570 @Abstractpublished in Advance ACS Abstracts, July 15, 1995.
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