Environ. Sci. Technol. 2001, 35, 2022-2025
Spectrophotometric Assay of POE Nonionic Surfactants and Its Application to Surfactant Sorption Isotherms D E R I C K G . B R O W N A N D P E T E R R . J A F F EÄ * Department of Civil and Environmental Engineering, Princeton University, Princeton, New Jersey 08544
The operational range and suitability toward environmental samples of an iodine-iodide (I-I) assay for nonionic surfactants were assessed. The I-I assay provides a rapid and repeatable method for determining aqueous nonionic surfactant concentrations. Through a systematic examination of surfactant structure, the operational range of the assay was shown to be on the order of 10-6 to 10-3 MEO, where the concentration unit MEO is defined as the molar surfactant concentration multiplied by the number of ethylene oxide units in the surfactant molecule. For environmental samples, it was shown that the I-I assay can be applied to measurement of surfactant sorption isotherms to aquifer sands and bacteria cultures. A potential limitation of the I-I assay is interference with humic acids, with the magnitude of the interference dependent on the concentration of humic acids present. The main benefit of the I-I assay is that its high accuracy and ease of application allows measurement of low levels of surfactant sorption. Surfactant sorption to aquifer sand could be measured down to the range of 10-9 mol/g.
Introduction Surfactant applications have been widely studied by environmental engineers. Over the past decade, attention has been focused on remediation of sites contaminated by hydrophobic organic compounds (HOCs) by increasing the HOC solubility through the addition of surfactants. This has led to some studies involving laboratory and field experiments of HOC solubilization (1-4) and above-ground and in-situ soil washing (5-7). Studies have also shown that the surfactant solubilized HOC is bioavailable, and the use of surfactants are being examined for their capability to enhance the biodegradation of HOCs (4, 8-16). Finally, because of the capability of sorbed surfactants to alter surface properties, the effects of surfactants on bacterial transport through porous media are beginning to receive attention (17-23). For all these applications, simple techniques that allow quantification of surfactant sorption on sediments, colloids, and bacteria are desirable. Methods typically employed to measure aqueous surfactant concentrations include high performance liquid chromatography, surface tension measurements, radio-labeled surfactants, and total organic carbon analysis. Each of these techniques requires specialized laboratory equipment and has its own set of limitations on the systems that can be studied. This paper discusses an * Corresponding author phone: (609)258-4653; fax: (609)258-2799; e-mail:
[email protected]. 2022
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FIGURE 1. The nonionic surfactants used in the study are designated CxEy, where x is the number of carbons in the alkyl chain and y is the number of ethylene oxide units in the polyoxyethylene chain. additional spectrophotometric assay for nonionic surfactants, which utilizes an iodine-iodide reagent. This assay is very accurate and can easily be applied without the requirement for specialized equipment. Aqueous solutions of iodine and iodide have been previously applied to spectrophotometric analysis of nonionic surfactants. These iodine-iodide (I-I) assays have been used in a number of different studies for determination of nonionic surfactant sorption to colloids, including polystyrene latex (24-26) and crystalline atrazine (27). The most widely applied assay was proposed by Baleux (28), where an iodine and potassium iodide reagent is added to an aqueous surfactant solution, and the change in absorbance at 500 nm is used to determine the surfactant concentration. Baleux stated that this assay is valid with surfactant concentrations of 1-20 mg/L. The purpose of this paper it to examine two unexplored areas of the I-I assay. First, the operational range of the assay is clarified. All prior studies using the I-I assay focused on only a few surfactant samples, and as such, surfactant structure/concentration effects have not been delineated. This paper presents a systematic study of the operational range of this assay for a series of nonionic surfactants containing alkyl and polyoxyethylene (POE) chains of various lengths and provides a method for determining the operational range based on the surfactant structure. Second, the applicability of this assay for determination of surfactant sorption isotherms to environmentally relevant media is demonstrated. Specifically, surfactant sorption to aquifer sand and bacteria is explored, as is the effect of humic acids on the assay.
Materials and Methods Materials. The nonionic surfactants used in these experiments consist of an alkyl chain as the hydrophobic moiety and a POE chain as the hydrophilic moiety (Figure 1). Four series of surfactants, with varying alkyl and POE chain lengths, were used in these experiments (Table 1). The surfactants are designated CxEy, where x is the number of carbons in the alkyl chain and y is the number of ethylene oxide (EO) units in the POE chain. These surfactants were used as received without any further processing. The bacteria used in these experiments was a Sphingomonas species that was isolated from a mixed culture capable of degrading polycyclic aromatic hydrocarbons (8, 10). These bacteria are rod-shaped, approximately 2 µm long and 0.5 µm in diameter, and are relatively hydrophobic (23). In preparation for the experiments, the bacteria were grown for 5 days at 20 °C on Petri dishes of R2A agar (Diffco). They were then harvested and suspended in a baseline CaCl2 solution at an ionic strength of 2 × 10-3 M and a pH of 7 (adjusted with NaOH). The bacteria solution was placed on a magnetic stirrer and allowed to mix at room temperature for a minimum of 1 h. The solution was then centrifuged at 1500g for 30 min, decanted, resuspended in the baseline solution, and placed on a magnetic stirrer at 20 °C to mix overnight. The next morning the bacteria were centrifuged and resuspended to a final concentration of approximately 10.1021/es001807u CCC: $20.00
2001 American Chemical Society Published on Web 04/06/2001
TABLE 1. Surfactants Used in This Study Encompassed a Range of Alkyl (x) and POE (y) Chain Lengths (See Figure 1) surfactant CxEy
molecular weight
trade name
manufacturer
C12Ey C12E4 C12E7 C12E9 C12E10 C12E23
362 494 582 626 1198
Brij 30
Brij 35
Aldrich Sigma Sigma Sigma Aldrich
C13Ey C13E6 C13E10 C13E12 C13E18
464 640 728 992
C16E10 C16E20
682 1122
C18E10 C18E20 C18E100
710 1150 4670
Aldrich Sigma Aldrich Aldrich
FIGURE 2. Change in the dA500 measurement versus time for C12E23 at two different concentrations indicates that ∼30 min is required for the assay to reach equilibrium.
C16Ey Brij 56 Brij 58
Sigma Sigma
Brij 76 Brij 78 Brij 700
Sigma Sigma Sigma
C18Ey
4.3 × 109 CFU/mL, determined via absorbance measurements at 220 nm (23). The sand used for the surfactant sorption experiment was #60-#70 sieve F-75 Ottawa sand (U.S. Silica). U.S. Silica F-75 Ottawa sand is a rounded sand with a low organic carbon content ( ∼3. The absorbance of the baseline iodine-iodide solution is approximately 0.5 at 500 nm (see Figure 4), so that a dA500 of 3 requires an absolute absorbance of approximately 3.5. This is the realistic upper limit for absorbance measurements with standard spectrophotometers, and thus represents the upper limit for determining surfactant concentrations with the I-I assay. Examination of Figure 4 shows that the peak value of the difference curve is at approximately 470 nm. In an attempt to maximize the resolution of the I-I assay at low surfactant concentrations, calibration curves were determined for wavelengths around this value. These curves were found to be nonlinear, with the nonlinearity due to shifting of the VOL. 35, NO. 10, 2001 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 4. Iodine-iodide assay is based on the difference between the absorbance of the baseline iodine-iodide solution in water and the iodine-iodide added to the surfactant solution.
TABLE 2. Curve Fits for the I-I Assay of the Form dA500 ) Slope*CSurfa C12Ey
FIGURE 5. Normalized calibration curve for the C12Ey surfactant series. Units of MEO indicates the molar surfactant concentration multiplied by the number of ethylene oxide units in the surfactant molecule.
TABLE 3. Curve Fits for the I-I Assay of the Form dA500 ) Slope*Cnorm-surfa
# OE units
4
7
9
10
23
surfactant series
slope
R2
slope R2
8120 0.992
20013 0.999
33957 0.999
32460 0.995
72818 0.995
# OE units
6
10
12
18
C12Ey C13Ey C16Ey C18Ey
3123 3289 2119 1993
0.971 0.995 0.999 0.952
slope R2
19320 0.997
31780 0.996
39900 0.998
61776 0.994
# OE units
10
20
slope R2
20770 1.000
43240 1.000
# OE units
10
20
100
slope R2
20570 1.000
30720 1.000
238600 1.000
C13Ey
a Where C norm_surf is the normalized molar surfactant concentration (MEO). Curve fits are based on the combined data for each surfactant series.
C16Ey
C18Ey
a
Where CSurf is the surfactant concentration in mol/L.
location of the peak to higher wavelengths with increasing surfactant concentration. Because of this, the 500 nm wavelength suggested by Baleaux was maintained. The slopes of the calibration curves in Table 2 change linearly as a function of the POE chain length, suggesting that the I-I assay interacts with individual EO units of the POE chain. To examine this in more detail, the data for each surfactant was normalized by multiplying the molar surfactant concentration by the number of EO units in the POE chain. This normalized molar concentration is given units of “MEO” in order differentiate it from the molar concentration (M) of the surfactant. The data for each surfactant series was combined and then plotted against dA500. Figure 5 shows the normalized data for the C12Ey surfactants, and the curve fits for the combined data for each surfactant series are presented in Table 3. The linearity of the normalized and combined data indicates that the assay is directly related to the length of the POE chain. It should be noted that the slight drop in R2 values from the individual surfactants in Table 2 to combined normalized data in Table 3 is most likely due to the use of the surfactants without further purification. As received from the manufacturer, the surfactants actually contain a small fraction of monomers with POE chain lengths that are somewhat higher and lower than the specified EO number (29), thus inducing slight errors when normalizing by the mean POE chain length. It has been suggested that the I-I assay works by the I3ion being trapped within the helical structure of the POE chain (27). However, the results presented in Figures 3 and 5 indicate that there is an association occurring between the 2024
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I-I solution and each individual EO unit in the POE chain and that it is independent of any helix formation. This is because of the linearity in the I-I assay down to four ethylene oxide units (i.e., C12E4)swith only four EO units in a chain, the POE portion of the surfactant is too short and inflexible to form a helix. This interaction with individual EO units is supported by an earlier study, where the volume of a 0.05% aqueous solution of iodine required to produce turbididness in 0.5% nonionic surfactant solutions was found to be a function of the POE chain length (30). This effect has been ascribed to complexes formed between both iodine and I3with the oxygen atoms in nonionic surfactants (31). One other point to note with the normalized data presented in Table 3 is that while the C12Ey and C13Ey slopes are similar and the C16Ey and C18Ey slopes are similar, there is a difference between the two groupings. If the I-I solution reacts solely with the POE chain (27), then all the surfactants should have the same normalized calibration curves. While the slopes shown in Table 2 suggest there may be some interference from the longer alkyl chains, this grouping may also be due to broader POE chain length distributions in the surfactants with longer alkyl chains. The data presented in Figure 5 and Table 3 suggests a different operational range of the I-I assay than the 1-20 mg/L stated by Baleux. With a practical maximum dA500 of ∼3, the upper limit of the I-I assay is a normalized surfactant concentration of ∼10-3 MEO. The lower limit is a function of the sensitivity of the spectrophotometer. In these experiments, surfactants were routinely measured down to 10-5 MEO, and C12E4 was measured down to ∼10-6 MEO. Thus, the operational range of the I-I assay is on the order of 10-6 to 10-3 MEO. Application of the I-I Assay for Sorption Isotherms. Figures 6 and 7 show the experimental results for surfactant sorption to the bacteria and sand, respectively. In both cases, the sorption isotherms are Langmuirian, with the more hydrophobic C12E4 surfactant having a higher level of sorption than the more hydrophilic C12E23 surfactant. Figure 7 shows that the I-I assay can be used to measure very low levels of surfactant sorption. This is due to the very high correlation coefficients for the calibration curves (Table 2), allowing
acids. This interaction with humic acids illustrates that the suitability of the I-I assay should be assessed for each specific system under examination.
Acknowledgments The authors would like to acknowledge the support of National Science Foundation grant BES-9710301.
Literature Cited FIGURE 6. Sorption isotherms for C12E4 and C12E23 onto the bacteria culture.
FIGURE 7. Sorption isotherms for C12E4 and C12E23 onto the aquifer sand.
FIGURE 8. Addition of the iodine-iodide reagent to an Aldrich humic acid solution resulted in a 30% drop in the absorbance at 500 nm. The absorbance readings at 0.4 mg/L of humic acids are within the error limit of the spectrophotometer. subtraction of relatively large numbers to calculate the low level of sorption. It is important to note that since there was no observed interference with the I-I assay from either the bacteria culture or the sand, the I-I assay was readily applicable for determination of surfactant sorption isotherms for these systems. There was also no difference between systems with deionized water and systems with CaCl2 as the salt, suggesting that the I-I assay is not affected by divalent cations (data not shown). However, in an experiment with Aldrich humic acid, it was found that addition of the iodine-iodide reagent resulted in a decrease in the absorbance at 500 nm of the humic acid solution by ∼30% (see Figure 8). This drop in the visible range suggests that inner sphere complexes are forming between the iodide and humic acids (32). In order for the I-I assay to be a viable means of determining surfactant concentrations, any interference with the assay should result in an absorbance shift well below the dA500 values being measured. Thus, for the interference shown in Figure 8, the presence of low levels of humic acids may affect the lower detection limit of the I-I assay, and the I-I assay may be inappropriate with high concentrations of humic
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Received for review October 24, 2000. Revised manuscript received February 16, 2001. Accepted February 27, 2001. ES001807U
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