Nature of the Interaction between a Peptidolipid Langmuir Monolayer

May 5, 2007 - Haifu Zheng and Xuezhong Du. The Journal of Physical Chemistry B 2010 114 (1), 577-584. Abstract | Full Text HTML | PDF | PDF w/ Links...
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J. Phys. Chem. C 2007, 111, 7826-7833

Nature of the Interaction between a Peptidolipid Langmuir Monolayer and Paraoxon in the Subphase Chengshan Wang,† Jiayin Zheng,† Osvaldo N. Oliveira, Jr.,‡ and Roger M. Leblanc*,† Department of Chemistry, UniVersity of Miami, Coral Gables, Florida 33146, and Instituto De Fisica De Sa˜ o Carlos, UniVersidade de Sa˜ o Paulo, CP 369, 13560-970, Sa˜ o Carlos/SP, Brazil ReceiVed: February 5, 2007; In Final Form: March 21, 2007

Surface potential and Infrared Reflection-absorption spectroscopy (IRRAS) measurements have been used to investigate the interaction between a peptidolipid (stearoyl-Phe-Trp-Ser-His-Glu) Langmuir monolayer and paraoxon aqueous solution as subphase. The molecular recognition in this system involved π-π interactions between the nitrobenzene group of paraoxon and aromatic groups in the peptidolipid. Of particular importance was the dependence of the surface potential on the concentration of paraoxon. Effects were practically negligible for 1.0 × 10-5 and 1.0 × 10-6 M paraoxon concentrations, but the Langmuir monolayer surface potential dropped due to the interaction with paraoxon at a concentration equal or higher than 1.0 × 10-4 M. At 1.5 × 10-3 M, the surface potential-area (∆V-A) isotherm for the peptidolipid displayed an unusual shape, with an almost constant, near-zero surface potential during the monolayer compression. This was interpreted on the basis of IRRAS results as being due to reorientation of the benzene ring of paraoxon, which changed from parallel to the air-water interface in the absence of a monolayer to a tilted orientation upon interacting with the peptidolipid monolayer.

Introduction The increasing damage to the environment caused by pesticides and other water and soil pollutants has made it essential to seek new strategies to detect and to decontaminate such pollutants. These strategies stand higher chances of success if information is available at the molecular level regarding the interaction between pollutant and possible substrata. For paraoxon, an important environmental pollutant widely used as pesticide,1 efforts are necessary not only to detect but also to dispose it in case of accumulation. Methods to detect paraoxon include gas chromatography,2,3 high-performance liquid chromatography,4,5 and bioassays based on enzymes.6-12 The latter approach allows a fast, sensitive, and specific detection of paraoxon, for which enzymes such as acetylcholinesterase (AChE) and organophosphorus hydrolase (OPH) have been used.6-12 A shortcoming in this approach though lies in the limited availability of enzyme sources, which has motivated the use of peptidolipids composed of amino acid residues from the binding site of the enzyme to mimic the enzyme interaction with the substrate.13-15 The synthesis of two peptidolipids, namely, C18H35O (stearoyl)-Phe-Trp-Ser-His-Glu and C18H35O (stearoyl)Gly-His-Ser-Glu-Glu, was reported in a previous paper, where interaction between paraoxon and AChE was mimicked.15 According to X-ray diffraction data from single crystals, the binding site of AChE contains mainly serine (Ser), histidine (His), glutamic acid (Glu), and an aromatic gorge formed by phenylanaline (Phe), tryptophan (Trp), and aromatic residues that assist paraoxon in reaching the binding site.16 The two peptidolipids differed with regard to the groups present in AChE. * Corresponding author. Fax: +1- 305-284-6367. Tel: +1-305-2842194. E-mail: [email protected]. † University of Miami. ‡ Universidade de Sa ˜ o Paulo.

The first peptidolipid contained amino acids appearing in both the binding site and aromatic gorge, whereas the second one only contained the amino acids present in the binding site. From surface pressure-area (π-A) isotherms of the two peptidolipids in absence and presence of paraoxon in the subphase, we noted that only the first peptidolipid interacted with paraoxon and the molecular recognition was specific. The fluorescence from Trp in the first peptidolipid was quenched completely when the concentration of paraoxon was 1.5 × 10-3 M in the subphase, from which we could infer that Trp was involved in the interaction with paraoxon.15 In this paper, we examine the nature of the interaction between paraoxon and the first peptidolipid, hereafter referred to as the peptidolipid, since it will be the only one investigated. The techniques employed for the analysis of the nature of the interaction were the surface pressure and surface potential-area isotherms and infrared reflection-absorption spectroscopy (IRRAS)17-21 of the peptidolipid Langmuir monolayer on subphases containing paraoxon. Because both surface potential and polarized IRRAS spectra depend on molecular orientation, the results obtained allowed us to suggest a molecular model for the nature of the interaction. Experimental Section Materials. The synthesis and purification of the peptidolipid have been reported in a previous paper,15 and the molecular structure is shown in Figure 1. Paraoxon was purchased from Sigma-Aldrich (St Louis, MO), and the purity was higher than 98%. Other organic chemicals and solvents were of reagent grade and were obtained from VWR Co. (Westchester, PA). The water utilized as subphase (pH 5.8) for surface chemistry study was obtained from a Modulab 2020 water purification system (Continental Water System Corp., San Antonio, TX) with a surface tension of 72.6 mN‚m-1 and a resistivity of 18 MΩ·cm at 20.0 ( 0.5 °C.

10.1021/jp071000p CCC: $37.00 © 2007 American Chemical Society Published on Web 05/05/2007

Peptidolipid Langmuir Monolayer and Paraoxon

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Figure 2. Surface pressure-area (π-A) and surface potential-area (∆V-A) isotherms of the peptidolipid at the air-water interface in the absence and presence of paraoxon in the subphase at the concentration of 1.5 × 10-3 M: (a) π-A isotherm on pure water subphase and (b) on paraoxon subphase. (c) ∆V-A isotherm on pure water subphase and (d) on paraoxon subphase.

paraoxon dry film was made by spreading paraoxon on the germanium (Ge) ATR crystal with 45° incident angle and 25 reflections and the ATR spectrum was recorded by coaddition of 64 scans. Results and Discussions

Figure 1. Chemical structures of the peptidolipid C18H35O-Phe-TrpSer-His-Glu and paraoxon.

General Methods for Surface Chemistry Study. All the isotherm measurements were conducted in a clean room [class1000] where constant temperature (20.0 ( 0.5 °C) and humidity (50 ( 1%) were maintained. The spreading solvent was chloroform/methanol (5:1; v/v). A Kibron minitrough (Kibron Inc., Helsinki, Finland) was utilized for the surface pressure-area isotherm. Surface potential measurements were obtained with the Kibron trough using a Kelvin probe (a vibrating capacitor system). The vibrating plate was set at approximately 1 mm above the water surface, and a gold plated trough was used as a counter electrode. The pure water subphase was taken as the zero potential. Surface tension measurements of paraoxon solutions were also measured by a Kibron minitrough at 25.0 ( 0.5 °C. This temperature was chosen because the reference surface tension values of pure water and other organic solvent, such as chloroform and methanol, which were used for calibration, were reported at 25.0 °C.22 IRRAS and Attenuated Total Reflection (ATR) Spectroscopy. IRRAS measurements at the air-water interface were performed on the EQUINOX 55 Fourier transform infrared (FTIR) spectrometer (Bruker Optics, Billerica, MA) equipped with an XA-511 external reflection accessory suitable for the air-water interface experiments. The IR beam was conducted out of the spectrometer and focused onto the water surface of the Kibron minitrough. It has been reported that the shuttle method can waive the water vapor effect;17-19 here it was proven that the water vapor was waived successfully in the clean room [class1000] because the temperature and humidity were kept constant. The external reflection-absorption spectra of pure water were used as reference. The spectra were acquired with a resolution of 8 cm-1 by coaddition of 1024 scans. The

1. Surface Pressure (π) and Surface Potential (∆V)-Area (A) Isotherms. The π-A and ∆V-A isotherms of the peptidolipid Langmuir monolayer are shown in Figure 2. For a pure water subphase (Figure 2a), the lift-off point of the π-A isotherm occurred at 94.0 Å2‚molecule-1, with collapse at 47.5 Å2‚molecule-1 and surface pressure of 62 mN‚m-1. The limiting molecular area obtained by extrapolating the low compressibility region of the isotherm to nil surface pressure was 69.4 Å2‚molecule-1, which is similar to the value predicted with the CPK model,15 i.e., 68.0 Å2‚molecule-1. The surface potential (Figure 2c) increased monotonically, with the onset at 145.0 Å2‚molecule-1 and reaching a maximum value of 130 mV at 61.3 Å2‚molecule-1. When paraoxon was added to the subphase at the concentration of 1.5 × 10-3 M, the Langmuir monolayer (Figure 2b) became more condensed, with lift-off at 105 Å2‚molecule-1, collapse surface pressure and limiting molecular area decreasing to 50 mN‚m-1 and 62.5 Å2‚molecule-1, respectively. The ∆V-A isotherm (Figure 2d) was unusual in the presence of paraoxon, with a nil surface potential during the compression of the peptidolipid Langmuir monolayer. We hypothesized that this unusual ∆V-A isotherm was related to the concentration of paraoxon in the subphase and decided to investigate the concentration effect. 2. π-A and ∆V-A Isotherms of the Peptidolipid Langmuir Monolayer with Different Concentrations of Paraoxon in the Subphase. The effect of paraoxon on the properties of the peptidolipid Langmuir monolayer depends on the paraoxon concentration, as indicated in Figure 3. For the concentration of 1.0 × 10-3 M in the subphase (Figure 3a), the lift-off point of the π-A isotherm appeared at 114.6 Å2‚molecule-1 with collapse at 44.1 Å2‚molecule-1. The limiting molecular area was 67.6 Å2‚molecule-1, which was similar to the value on pure water. As for the ∆V-A isotherm, the surface potential increased slowly and reached a maximum value of 20 mV at 78.5 Å2‚molecule-1. When paraoxon concentration was decreased to 1.0 × 10-4 M (Figure 3b), the π-A isotherm was similar to the one at 1.0 × 10-3 M, whereas the surface potential reached a maximum value of 86 mV when the molecular area decreased to 75.0 Å2‚molecule-1. With further decrease of the concentra-

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Figure 3. π-A and ∆V-A isotherms of the peptidolipid at the air-water interface in presence of paraoxon at different concentrations: (a) 1.0 × 10-3, (b) 1.0 × 10-4, (c) 1.0 × 10-5, and (d) 1.0 × 10-6 M.

TABLE 1: Surface Tension of Paraoxon Solutions at 25.0 ( 0.5 °C concentration of paraoxon (M)

Nil

surface tension 71.99 (mN‚m-1)

Figure 4. ATR spectrum of pure paraoxon dry film.

tion of paraoxon, 1.0 × 10-5 (Figure 3c) and 1.0 × 10-6 M (Figure 3d), the π-A and ∆V-A isotherms were essentially the same as on pure water. Therefore, upon decreasing the paraoxon concentration, the condensation effect becomes progressively smaller in the Langmuir monolayer. The strongest decrease in surface potential induced by paraoxon at 1.5 × 10-3 M also diminishes as the concentration is decreased, until the same results for pure water are observed at 1.0 × 10-6 M paraoxon concentration. Because paraoxon in the bulk could not have such a large effect on surface potential, we presume that paraoxon must accumulate at the interface. Indeed, paraoxon itself is surface active, which was confirmed by measurements of surface tension of paraoxon solutions with no peptidolipid spread. The results are shown in Table 1, in which the measured surface tension varied from 71.99 mN‚m-1 for pure water to 71.9, 70.9, 66.9, and 60.5 mN‚m-1 at 1.0 × 10-6, 1.0 × 10-5, 1.0 × 10-4, and 1.5 × 10-3 M, respectively. In subsidiary experiments, we

1.0 × 10-6 1.0 × 10-5 1.0 × 10-4 1.5 × 10-3 71.9

70.9

66.9

60.5

measured the surface potential of paraoxon subphase at the concentration of 1.5 × 10-3 M, and noted that the surface potential is negative (result not shown), which has an important consequence on the analysis of the nature of the interaction with the peptidolipid Langmuir monolayer. Since changes in the surface potential of the peptidolipid Langmuir monolayer were related to the interaction between the peptidolipid and paraoxon in the subphase, molecular level information about this interaction is required to explain the data. This motivated us to measure the IRRAS spectra of the peptidolipid Langmuir monolayer spread on paraoxon subphases, with the measured ATR spectrum of the paraoxon dry film being used as reference. 3. ATR Spectrum of Paraoxon Dry Film. The ATR spectrum of paraoxon dry film is shown in Figure 4, with the assignment of the peaks being given in Table 2. The peaks at 1612 and 1593 cm-1 were assigned to the stretching mode of the benzene ring. The nitro group has two strong peaks at 1526 and 1348 cm-1 which were assigned to asymmetric and symmetric stretching modes, respectively. The peak at 1491 cm-1 was assigned to the bending mode of the ethyl group and the peaks at 1296 and 1273 cm-1 were assigned to the stretching mode of PdO. The methyl group has three rocking vibration peaks at 1164, 1107, and 930 cm-1. The peak at 1232 cm-1 was assigned to the stretching mode of P-O-Ar and the stretching mode of P-O-Et appeared at 1025 cm-1. The last peak at 860 cm-1 was assigned to the stretching mode of C-N.

Peptidolipid Langmuir Monolayer and Paraoxon TABLE 2: Assignment of the Peak Positions of the ATR Spectrum of a Dry Film of Paraoxon peak position (cm-1)

assignment

860 930 1025 1107 1164 1232 1296, 1273 1348 1491 1526 1612,1593

ν(C-N) CH3 rock ν(P-O-Et) CH3 rock CH3 rock ν (P-O-Ar) ν (PdO) νs(-NO2) δ(C-H of Et) νas(-NO2) ν(CdC of Ar)

4. s-Polarized and p-Polarized IRRAS Spectra of the Peptidolipid Langmuir Monolayer on Pure Water. Polarized IRRAS is one of the most straightforward methods to probe molecular level interactions between guest and film-forming molecules at the air-water interface, especially because the information on orientation of the molecules may be obtained at various stages of compression. The s-polarized and p-polarized IRRAS spectra for a given monolayer may differ due to distinct selectivity rules. As Scheme 1 illustrates, the electronic field of the s-polarized IR beam is perpendicular to the incident plan (x-z plane in Scheme 1), therefore s-polarized IRRAS is only sensitive to vibrations whose transition moment is in the y direction as shown in Scheme 1. The bands of s-polarized IRRAS are always negative, with decreasing intensity when the incident angle increases. For the p-polarized IR beam, the electronic field is parallel to the x-z plane and perpendicular to the forward direction of the incident beam, therefore it is sensitive to vibrations with transition moment in both x and z directions as shown in Scheme 1. For vibrations with transition moment in the x direction, the bands are initially negative and become more intense with increase of incident angle until the Brewster angle (54.2° for the IR light at 2850 cm-1)18 is reached. Above the Brewster angle the bands become positive and decrease in intensity upon further increase of the incident angle. For a vibration with a transition moment parallel to the z direction as shown in Scheme 1, the sign of the signal is reversed: the peak is positive first, then turns negative as the incident angle is above the Brewster angle. The s-polarized IRRAS spectra of the peptidolipid Langmuir monolayer at a surface pressure of 20 mN‚m-1 are shown in Figure 5a. The peaks at 2923 and 2854 cm-1 were assigned to the asymmetric and symmetric stretching modes of CH2, respectively, which indicated that there were some gauche conformations in the alkyl chain.23,24 The peak at 1630 cm-1 and the shoulder at 1654 cm-1 were assigned to the amide I band, while the peaks at 1557, 1542, and 1525 cm-1 were due to the amide II band. The peak at 1580 cm-1 arose from superposition of contributions from Phe and His and the peak at 1510 cm-1 was assigned to the symmetric stretching mode of the residue group of Glu, namely the carboxylate anion. The peak of Trp at 1590 cm-1 was not observed, probably because it was masked by the stronger peak from the superposition of Phe and His. The p-polarized IRRAS spectra of the peptidolipid Langmuir monolayer on pure water are shown in Figure 5b. Only the peaks of CH2 and amide I band were clearly observed. The difference between p-polarized and s-polarized IRRAS spectra is due to the different selection rules as discussed above (Scheme 1). The negative peak of asymmetric and symmetric stretching mode of CH2 below the Brewster angle was at 2923 and 2854 cm-1,

J. Phys. Chem. C, Vol. 111, No. 21, 2007 7829 SCHEME 1: Schematic Illustration of the Principle of s-Polarized and p-Polarized IRRAS

SCHEME 2: Free Rotation of Benzene Ring of Paraoxon at the Air-Water Interface

respectively, which were at the same positions as in s-polarized IRRAS spectra. However, when the incident angle was above the Brewster angle, the two peaks disappeared. The same result was observed even for high surface pressures of 30 or 40 mN‚m-1 (results not shown). If the orientation of CH2 bisector of the alkyl chains was isotropically distributed in the x-y plane as shown in Scheme 1, the peaks of asymmetric and symmetric stretching modes of CH2 should be positive above the Brewster angle. The disappearance of the two peaks is possibly due to two reasons: (i) the orientations of CH2 bisector of the alkyl chains were anisotropically distributed in the x-y plane as shown in Scheme 1. (ii) The alkyl chains have some twists, which were possibly due to the alkyl chains being in a disordered state. As for the amide I band, two peaks at 1683 and 1630 cm-1 and a shoulder peak at 1654 cm-1 were observed. For the amide II band, two peaks at 1557 and 1542 cm-1 were observed, while the peak of carboxylic anion appeared at 1510 cm-1. All these peaks of amide I and II band were negative at first and became positive after the Brewster angle which indicated that the carbonyl groups were parallel to the airwater interface. The peak at 1580 cm-1 assigned to the superposition band of benzene ring in Phe and imidazole group in His was not observed, which revealed that these groups were tilted to the air-water interface. The s-polarized IRRAS spectra with the incident angle at 35° and p-polarized IRRAS spectra with the incident angle at 45° of the peptidolipid Langmuir monolayer at different surface pressures are shown in Figure 5c and 5d. The incident angle at 35° and 45° were selected because the signal-to-noise ratio has the highest value for these angles. As for s-polarized IRRAS spectra, all the peaks intensity increased with the increase of surface pressure. The peaks at 1654 and 1557 cm-1 increased more than the other peaks. In the p-polarized IRRAS spectra, the intensity of the peaks at 1654, 1630, and 1557 cm-1 increased further than the other peaks. Because the direction of transition moments of amide II band is different compared with

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Figure 5. IRRAS of the peptidolipid Langmuir monolayer on pure water: (a) s-polarized and (b) p-polarized IRRAS of Langmuir monolayer of the peptidolipid at 20 mN‚m-1 against different incident angles, (c) s-polarized IRRAS with incident angle at 35°, and (d) p-polarized IRRAS with incident angle at 45° of Langmuir monolayer of the peptidolipid at different surface pressures.

amide I band, we suggest that the OdC-N plane of the amide groups is parallel to the air-water interface and that the carbonyl and N-H groups become more parallel to the air-water interface with the increase of surface pressure. 5. s-Polarized and p-Polarized IRRAS Spectra of Paraoxon Subphase at the Concentration of 1.5 × 10-3 M without the Peptidolipid Langmuir Monolayer. The s-polarized IRRAS spectra of the paraoxon solution are shown in Figure 6a. Almost all the peaks of ATR spectrum of the paraoxon dry film in the range of 1700 to 1100 cm-1 were observed. However, only the peaks at 1348, 1273, 1232, and 1164 cm-1 were observed in the p-polarized IRRAS spectra (Figure 6b). The peak at 1348 cm-1 was assigned to the symmetric stretching mode of the nitro group in paraoxon and this peak was negative when the incident angle was below the Brewster angle, whereas it was positive when the incident angle was above the Brewster angle. Consequently, the symmetric vibration of the nitro group should be parallel to the air-water interface. Because the transition moment of the symmetric stretching mode of the nitro group was parallel to the bisector of the nitro group and benzene ring, one infers that the benzene ring of paraoxon was also parallel to the air-water interface. The peak at 1526 cm-1 assigned to the asymmetric stretching mode of the nitro group was not observed in p-polarized IRRAS spectra. As we know, the benzene ring can rotate freely around the bisector. For the paraoxon whose benzene ring was parallel to the air-water interface (Scheme 2a), the asymmetric vibration was parallel to the air-water interface, whereas for the benzene ring perpendicular to the air-water interface (Scheme 2b), the asymmetric vibration was perpendicular to the air-water interface. Consequently, this random orientation caused the weak intensity of the peak assigned to the asymmetric stretching. For the same reason, the peaks of CdC stretching mode of benzene

Figure 6. S-polarized (a) and p-polarized (b) IRRAS of paraoxon subphase at the concentration of 1.5 × 10-3 M.

ring (1612 and 1593 cm-1) and the bending mode of ethyl group (1491 cm-1) were not observed either. The peaks at 1273, 1232, and 1164 cm-1, which were assigned to the stretching mode of PdO, P-O-Ar, and the rock mode of CH3, were observed.

Peptidolipid Langmuir Monolayer and Paraoxon

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Figure 7. IRRAS of the peptidolipid Langmuir monolayer on paraoxon subphase at the concentration of 1.5 × 10-3 M: (a) s-polarized and (b) p-polarized IRRAS of Langmuir monolayer of the peptidolipid at 20 mN‚m-1 against different incident angles, (c) s-polarized IRRAS with incident angle at 35°, and (d) p-polarized IRRAS with incident angle at 45° of Langmuir monolayer of the peptidolipid at different surface pressures.

These peaks were positive when the incident angle was above the Brewster angle, however they were still positive when the incident angle was at 45, 40, and 35°. When the incident angle was at 25 and 30°, their direction changed to negative. The reason of this result is not clearly understood. The disappearance of the peak of PdO at 1296 cm-1 was possibly due to the relative weak intensity and consequently covered by the peak at 1273 cm-1. The IRRAS spectra of paraoxon subphase at the concentration of 1.0 × 10-3, 1.0 × 10-4, 1.0 × 10-5, and 1.0 × 10-6 M were also measured. The result at the concentration of 1.0 × 10-3 M was similar to the result at the concentration of 1.5 × 10-3 M. When the concentration of paraoxon was further decreased, no peak of paraoxon was observed due to the weak signal. 6. s-Polarized and p-Polarized IRRAS Spectra of the Peptidolipid Langmuir Monolayer in Presence of Paraoxon in the Subphase at the Concentration of 1.5 × 10-3 M. The s-polarized IRRAS spectra of the peptidolipid Langmuir monolayer in presence of paraoxon at the concentration of 1.5 × 10-3 M are shown in Figure 7a. The peaks of asymmetric and symmetric stretching modes of CH2 were observed at 2920 and 2850 cm-1, respectively, which indicated that the alkyl chains were in all-trans conformation. There was no change of the amide I and II bands compared with the results of the peptidolipid monolayer on pure water (Figure 5a). In contrast, the peak at 1580 cm-1 became stronger which indicated that the aromatic residue groups of Phe and His became more parallel to the air-water interface. The peaks of paraoxon at 1348, 1296, and 1273 cm-1 were observed; however, the intensity of the peak at 1348 cm-1 was much weaker than that in the s-polarized IRRAS spectra of paraoxon solution without the peptidolipid Langmuir monolayer (Figure 6a). This indicated that the orientation of the benzene ring of paraoxon was less parallel to

the interface after the peptidolipid Langmuir monolayer was spread at air-water interface. As for the p-polarized IRRAS spectra (Figure 7b), the peaks of asymmetric and symmetric stretching modes of CH2 (2920 and 2850 cm-1) were negative when the incident angle was below the Brewster angle and became positive when the incident angle was above the Brewster angle, which indicated that the bisector of CH2 group was parallel to the air-water interface. There was no change in the amide I and II bands compared to the results of the peptidolipid Langmuir monolayer on pure water. As for the peaks at 1273, 1232, and 1164 cm-1, they were similar to the peaks of paraoxon solution without the peptidolipid Langmuir monolayer. These results indicated that the orientation of PdO and P-O-Ar did not change after the peptidolipid was spread. However, the peak of paraoxon at 1348 cm-1 was not observed in the p-polarized IRRAS spectra, which indicated that the nitrobenzene group of paraoxon was not parallel to the air-water interface but tilted at a certain angle (Scheme 3a). This result was correlated to the result of s-polarized IRRAS. The peak at 1580 cm-1 assigned to the aromatic residue groups in Phe and His was not observed in the p-polarized IRRAS spectra of the peptidolipid Langmuir monolayer in the presence of paraoxon at 1.5 × 10-3 M. Therefore, even though the s-polarized IRRAS showed that Phe and His groups became more parallel to the air-water interface, their aromatic residues were still tilted at an angle to the normal of the air-water interface, in a similar orientation of the benzene ring of paraoxon. It is inferred that the nitrobenzene ring of paraoxon was possibly parallel to the aromatic groups of the peptidolipid. From previous results,15 it was concluded that the aromatic groups in the peptidolipid was important for the interaction between the peptidolipid and paraoxon and that the peptidolipid fluorescence was quenched by paraoxon owing to π-π interac-

7832 J. Phys. Chem. C, Vol. 111, No. 21, 2007 SCHEME 3: (a) Change of Orientation of Nitro Group of Paraoxon Before and After Spreading Peptidolipid at Air-water Interfacea

a (b) Schematic explanation of how paraoxon caused the monolayer surface potential to decrease.

SCHEME 4: Model of the Interaction between the Peptidolipid and Paraoxon

tions between the nitrobenzene group of paraoxon and aromatic groups in the peptidolipid. The IRRAS results presented here supported this explanation. The s-polarized IRRAS spectra with incident angle at 35° and p-polarized IRRAS with incident angle at 45° of the peptidolipid Langmuir monolayer at different surface pressures are shown in Figure 7c,d. For both s-polarized and p-polarized IRRAS spectra, the intensity of all the peaks increased with increasing surface pressures to the same extent. This means that neither the hydrophilic nor hydrophobic moieties of the peptidolipid changed orientation upon compression, with the change in intensity being merely caused by an increase of peptidolipid density in the Langmuir monolayer as the pressure was increased. 7. Model for Interaction between Peptidolipid and Paraoxon. The molecular-level information obtained from IRRAS makes it possible to explain qualitatively the surface potential data, in addition to establish a model for the interaction between peptidolipid and paraoxon. Two groups in paraoxon have a large dipole moment, namely PdO and nitrobenzene. On the basis of both s-polarized and p-polarized IRRAS spectra, we may assume that the orientation of PdO and P-O-Ar groups was not changed by the peptidolipid Langmuir monolayer, whereas the orientation of the nitrobenzene group changed. Therefore, the change of the orientation of the latter group should be responsible for the unusual ∆V-A isotherm of the peptidolipid Langmuir monolayer. Because the value of surface potential of the peptidolipid Langmuir monolayer decreased in the presence

Wang et al. of paraoxon, the nitro group should be positioned toward the air as shown in Scheme 3a. The peaks of nitro group of paraoxon disappeared in the p-polarized IRRAS even when the surface pressure was 5 mN‚m-1 (results not shown). This indicated that the nitro group of paraoxon changed its orientation as soon as the peptidolipid Langmuir monolayer covered the surface of the subphase. The unusual result of ∆V-A isotherm of the peptidolipid Langmuir monolayer in the presence of paraoxon may be explained as follows: For large areas per molecule in the peptidolipid Langmuir monolayer the surface potential was zero as it occurs for most monolayers. As the barriers began to compress the monolayer, the peptidolipid molecules should contribute positively to the surface potential, as they became increasingly ordered. On the other hand, with the increasing coverage of the subphase with peptidolipid molecules, an increasing number of nitro groups from paraoxon would be oriented toward the air, thus contributing negatively to the surface potential (Scheme 3b). For the paraoxon concentration of 1.5 × 10-3 M, the two competing contributions cancelled out during monolayer compression, leading to the unusual shape for the surface potential isotherm. At lower paraoxon concentrations, the positive contribution from the peptidolipid prevailed and the isotherms resembled that of the pure peptidolipid monolayer. The results above allowed us to propose a model for the interaction between the peptidolipid and paraoxon, which is depicted in Scheme 4. For the peptidolipid Langmuir monolayer on pure water, the alkyl chains had some gauche conformation and the aromatic residues were tilted to the air-water interface at a larger angle. After paraoxon was added to the subphase, there was a π-π interaction between the aromatic residues and paraoxon, with the aromatic groups sharing some common area which decreased the limiting molecular area. A more compact packing between the peptidolipid molecules made the alkyl chains change to the all-trans conformation. Conclusion The nature of the molecular recognition between paraoxon and the peptidolipid Langmuir monolayer was analyzed using surface potential and IRRAS measurements. From IRRAS, we inferred that the benzene ring of paraoxon was parallel to the air-water interface in the absence of the peptidolipid Langmuir monolayer, but was tilted as the peptidolipid monolayer was compressed. This change in orientation of the nitrobenzene group, which has a large dipole moment, was responsible for the unusual surface potential-area isotherm. Furthermore, the maximum surface potential for a peptidolipid Langmuir monolayer depended on the concentration of paraoxon in the subphase. Acknowledgment. This research was supported by the National Science Foundation (USA, CHE-0416095) and the U.S. Army Research Office (DAAD 19-03-1-0131). ONOJ is grateful to CNPq (Brazil) for the financial support. A special thank to Mr. Jhony Orbulescu for a sound discussion about the manuscript and thanks to Mr. Ting Wang from Bruker Optics for help in the calibration of the IRRAS equipment. References and Notes (1) Munnecke, D. J. Agric. Food Chem. 1980, 28, 105-111. (2) Mendoza, C. E. Thin-layer chromatography. In Pesticide Analysis; Dumas, K. G., Ed.; Marcel Dekker: New York, 1981; pp 1-44. (3) Das, K. G.; Kulkarni, P. S. Gas-liquid chromatography. In Pesticide Analysis; Dumas, K. G., Ed.; Marcel Dekker: New York, 1981.

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