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water interface by Brewster angle microscopy
Materials Letters 58 (2004) 1466 – 1470 www.elsevier.com/locate/matlet
In-situ observation of the aggregated morphology and interaction of dialkyldimethylammonium bromide with DNA at air/water interface by Brewster angle microscopy Lu Sun, Miao Xu, Xueliang Hou, Lixin Wu * Key Laboratory for Supramolecular Structure and Materials of Ministry of Education, Jilin University, Changchun 130023, PR China Received 14 July 2003; accepted 3 October 2003
Abstract The adsorption of DNA on the Langmuir film of a cationic surfactant, dioctadecyldimethylammonium bromide (DODABr), and the change of the aggregation morphology of the composite monolayer with respect to surface pressure have been investigated by Brewster angle microscopy (BAM). In contrast with the case of DODABr on pure water subphase, when DNA was dispersed into subphase, its adsorption to the interface monolayer through electrostatic interaction decreases the charge density and therefore promotes the formation of domain at low surface pressure. In addition, the electrostatic interaction changed the phase morphology of DODABr Langmuir monolayer under different surface pressure, that is, from flower-shaped crystalline domain on the pure water subphase to circular domain on the subphase dispersed with DNA. The result also shows that the monolayer of the composite at air/water interface under the high pressure is not homogeneous, but consists of incompletely fused domains. For the Langmuir film of the surfactant with shorter alkylchains, similar morphology can be observed both under the high and low surface pressure. But the tight-stacked circular domain is no longer observed. D 2003 Elsevier B.V. All rights reserved. Keywords: Air/water interface; Brewster angle microscopy; Surfactant; DNA; Complex
1. Introduction Due to its double helical structure and negative charged phosphate backbone, DNA is easy to form electrostatic complex with cationic surfactants [1,2]. This kind of cationic surfactant-DNA complex supporting with polymer micelle matrix has been used for gene transfer in vivo, and it also has been found very promising in the application of bio-function materials and biosensors [3 –6]. The cationic surfactant-DNA complex displays various aggregation morphologies under different conditions. The fluorescent microscopy has shown that the DNA molecule will gradually condense from linear to globular when it forms a complex with cationic surfactant in aqueous solution [7]. In organic solvent, the surfactant-DNA complex is proposed mostly existing in globular aggregate state due to hydrophilic and * Corresponding author. Tel.: +86-431-8499649; fax: +86-4318980729. E-mail address: [email protected] (L. Wu). 0167-577X/$ - see front matter D 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.matlet.2003.10.010
hydrophobic equilibrium. Okahata et al. have proved that DNA molecule adsorbs to the cationic surfactant in a rodlike form at the air/water interface. When the subphase was dilute dye-intercalated DNA aqueous solution, the Langmuir monolayer containing dye molecules at air/water interface could be transferred to solid substrate with the orientation [8]. However, as we know, few investigations related to the direct observation about aggregation morphology of the complex without introducing probe molecules under various surface pressures. Brewster angle microscopy (BAM) is a suitable technique for in-situ observation of the microdomain structure of phase separation at gas/liquid or liquid/solid interface without probe [9]. So it provides an effective method to check the interaction at interface. In this report, we present the measurement of in-situ phase transformation of double-chain cationic surfactant monolayer on DNA subphase. In this paper, aggregation and morphologic changes of the surfactant-DNA Langmuir monolayer accompanied with the increasing of the surface pressure were investigated.
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2. Experimental 2.1. Materials The sodium salts of fish testes DNA were purchased from AMRESCO Chemical and used as received. Didodecyldimethylammonium bromide (DDDABr), dioctadecyldimethylammonium bromide (DODABr) and acridine orange (AO) are the chemicals from ACROS Chemical. Other organic solvents are the analytical grade and the water for subphase is redistilled deionized water. 2.2. Instrument and measurement Langmuir trough (622D) used in the experiment is a production of Nima England. BAM (I-Elli2000) is from NFT Germany, which is equipped with a 50-mW semiconductor laser, an analyzer, a zooming microscope with a high sensitive CCD camera and a video recording system. The wavelength of p-polarized light is 532 nm. The lateral resolution reaches 2 Am. The scale of the images is 870650 Am. BAM observation to Langmuir monolayers was carried out by spreading 30 Al of DODABr chloroform solution (1.16 mg/ml) onto air/water or air/solution interface with pure water, DNA (3.5410 6 mol/l in base pair) and the mixture of DNA with AO in a molar ratio (base pair) 3:1 aqueous solution as subphases, respectively. The k – A isotherms and corresponding BAM images were recorded after standing 20, 60 and 60 min with above subphases in order to make the organic solvent evaporate off completely and to reach their equilibrium states, respectively. Repeated experiments for DDDABr (1.28 mg/ml) were under the same conditions as those for DODABr. The pH value of the subphase was maintained at 6 –7, and the temperature was controlled at 20F0.5 jC.
3. Results and discussion 3.1. p – A isotherms of DNA-surfactant complex system Fig. 1A shows k –A isotherms of DODABr on different subphases. The collapse pressure of DODABr on the pure water is about 43 mN/m. The phase transition can be clearly observed in the range of 25 –35 mN/m, which is in agreement with the reported results [10]. Vranken et al. have discussed the differences of k –A isotherms of DODABr performed by different research groups [11,12]. More in general, any impurities existing in the sample or the subphase could influence the shape of the isotherm and phase transition pressure when the spreading amount is large enough. However, neither have DODABr in our samples nor any others from the related literatures been presented details on the definite amount of impurities, which affect the results [10]. So we measure the k – A isotherms of
Fig. 1. (A) k – A isotherms of DODABr on subphase of (a) pure water, (b) DNA aqueous solution in which the concentration is 3.5410 6 mol/l and (c) DNA and AO aqueous solution which concentration proportion is 3:1. (B) k – A isotherms of DDDABr on subphase of (a) DNA aqueous in which the concentration is 3.5410 6 mol/l and (b) DNA and AO aqueous solution in which the concentration proportion is 3:1.
DODABr several times under preset experiment conditions. The results can be repeatedly pretty well and the k –A curves are quite similar to the reported shapes [13]. In contrast to the pure water subphase, the k –A isotherm of DODABr shows that the preliminary area increases and collapse area decreases on the subphase of DNA aqueous solution. The collapse pressure is up to 64 mN/m and the phase transition still existed. When the subphase was changed to the molar ratio MDNA/MAO=3:1 of DNA and AO aqueous solution, the k – A isotherm of DODABr shows little change comparing with that of it on DNA subphase. As a negative charged biopolyelectrolytes, DNA can absorb onto the surfactant with positive charges at air/ water interface forming electrostatic complex. That makes larger the molecular area due to the binding of DNA occupying larger surface area and smaller the charge repulsive effect, which lead to the change of stacking property of surfactant monolayer with surface pressure. Such a property is similar to the interaction of surfactant on general polyelectrolyte subphase as described in literature [14,15]. In order to understand the influence of electrostatic interaction between DNA and Langmuir monolayer on water, DDDABr, the surfactant with short chains that is
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more sensitive to interaction at interface, has been used for this purpose. It has poor spreading property on pure water subphase because it will dissolve in water during the pressing due to its strong hydrophilic feature. But this feature is much sensitive to the subphase chemical environment and become stable at the interface especially when there is anionic polyelectrolyte in the subphase. DDDABr forms stable monolayer on DNA aqueous solution due to the electrostatic interaction between them, which reduces its hydrophilic feature, as shown in Fig. 1B. However, the stability of DDDABr again decreases greatly after adding AO to DNA aqueous subphase, which is quite different from the case of DODABr and shows the sensitivity of DDDABr to the change of DNA aqueous subphase. One of the possible reasons is that the addition of AO neutralizes the negative charges of DNA and that leads to the decreasing of the interaction between DNA and surfactant.
30– 50 Am (Fig. 2B). It is noted that the patterned domain is quite uniform both in scale and in shape, which is different from larger and anisotropic leaf-like domains as reported in literature on high concentration, indicative of the growth of crystalline domains being a controllable dynamic process. From this point the variety of the patterned domain should be derived from the change of the temperature, barrier speed and so on. And further more the flower-like domains change back to dot structure with surface pressure increasing (Fig. 2C). Around the dot structure on the air/water interface is liquid condensed phase of DODABr based on increasing gray level, indicating that the Langmuir monolayer is composed of two phases when the surface pressure closes to collapse point. For DDDABr, no clear morphologic structure on pure water subphase has been observed because of its strong hydrophilic property (the surface pressure only getting to 9.3 mN/m).
3.2. In-situ BAM observation to the surfactant monolayer on pure water subphase
3.3. In-situ BAM observation to the surfactant monolayer on DNA subphase
The BAM image of DODABr on the pure water subphase shows that there is no visible morphologic structure at the air/water interface when the pressure just starts to increase, while the gray level getting high. Dot domains with diameter in the region of 8 – 10 Am can be observed when pressure reaches 20 mN/m (Fig. 2A). Both the amount and the size of domains increase as pressure getting high. When the pressure is up to 30 mN/m, flower-like domains have been observed and their diameter falls into the range of
Corresponding to the k – A isotherms in Fig. 1A(b), under lower surface pressure below the phase transition (Fig. 3A) such as 11 mN/m, dot domains have been observed and the amount of the dots increases with the pressure increasing. It is clear that DNA promotes the formation of dot domains at low pressure because the dot domain normally emerges at much higher surface pressure on pure water. Around the phase transition the sizes of domains can be observed evenly and the amount of the domains increases quite much. As the
Fig. 2. BAM Images of DODA aggregate on the pure water at surface pressures of (A) 21 mN/m, (B) 30 mN/m, (C) 39 mN/m, respectively, and (D) DDDA on the DNA and AO aqueous subphase at surface pressure of 9.3 mN/m.
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Fig. 3. BAM Images of DODABr aggregates on the DNA aqueous subphase at surface pressures of (A) 11 mN/m, (B) 22 mN/m, (C) 34 mN/m, (D) 39 mN/m, and on the DNA and AO aqueous subphase at surface pressure of (E) 35 mN/m and (F) 40 mN/m, respectively.
pressure higher than that of the phase transition, no flowerlike domains but big circular domains have been observed (Fig. 3C), and the size distribution quite uniform. We believe that the condensation of DNA and the formation of the DODA-DNA complex through electrostatic reaction restrain flower-like crystalline domain of DODABr but enhance the circular domain and then promotes the phase transition. Aggregation of DNA presents the biological significance because it is important to gene transfer and cellular karyokinesis. So it helps us to understand the process of DNA going through cellular membrane though the aggregation of DNA at air/water interface is different from that occurred in aqueous solution. With the surface pressure increasing from phase transition region to collapse point, the surface morphology of DODABr monolayer changes from circular domain to homogeneous domain (Fig. 3D). In this process, we confirm that the aggregation of DNA should be influenced by surface pressure, that is, the density of positive
charge, which is similar to that of DNA going through the cellular membrane. On the subphase of DNA with intercalated AO, the sizes of DODABr aggregate domains become smaller and denser (Fig. 3E,F) in contrast to the aggregates of DODABr on the DNA subphase and uniform Langmuir monolayer near the collapse pressure. As has been well known, AO adsorbs electrostatically and intercalates into the double helix of DNA leading to the change of surface charge and existing state of DNA in water [16 –18]. So we believe that the addition of AO influences on the formation of the DNA-DODA complex and the aggregated morphology. The effect of charge and agglomeration to the surface morphology is not clear now. DDDABr also adsorbs DNA through electrostatic interaction at the air/water interface forming interface complex, which makes more stable its Langmuir monolayer and much smaller the aggregate domain. At very low surface pressure dot domains of DDDABr monolayer have been observed on
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the subphase of DNA mixed with AO. In contrast to the situation of DODABr, the dot domain of DDDABr, appeared at low surface pressure such as 9.3 mN/m, as shown in Fig. 2D, does not grow larger with the surface pressure increasing until collapsed. In addition, DDDABr shows shorter responding time and lower responding pressure indicating the difference of the lengths of the alkyl-chains between DODABr and DDDABr because dot domain is immediately observed for DDDABr but not for DODABr as it needs a while and can be observed on a certain pressure.
4. Conclusion From above results, we confirm that DODABr and DDDABr form complexes with DNA at the air/water interface, respectively. Through in-situ observation by BAM, we find that the agglomeration of DNA strongly influence on the interface property of surfactant. Through its electrostatic interaction to DNA, the aggregated morphology of DODABr at air/water interface changes from small dot domains to a little bit larger uniform circular domains while DODABr itself changes from dot structure to flowerlike domain and then to dot structure again on pure water. The similar action is observed to more hydrophilic DDDABr with DNA at air/water interface. The intercalation of dye to DNA changes the morphology of aggregated domains of Langmuir monolayer.
Acknowledgements We gratefully acknowledge the financial support from the National Natural Science Foundation of China
(29992590-5), the Major State Basic Research Development Program (Grant No.G2000078102) and the Ministry of Education of China.
References [1] K. Eskilsson, C. Leal, B. Lindman, M. Miguel, T. Nylander, Langmuir 17 (2001) 1666. [2] K. Wagner, D. Harries, S. May, V. Kahl, J.O. Raedler, A. Ben-Shaul, Langmuir 16 (2000) 303. [3] R.I. Zhdanov, O.V. Podobed, V.V. Vlassov, Bioelectrochemistry 58 (2002) 53. [4] J. Zabner, Adv. Drug Deliv. Rev. 27 (1997) 17. [5] H.P. Haagsman, R.V. Diemel, Comp. Biochem. Physiol., Part A 129 (2001) 91. [6] M.C. Garnett, Ther. Drug Carr. Syst. 16 (1999) 147. [7] M. Melnikov, V.G. Sergeyev, K. Yoshikawa, J. Am. Chem. Soc. 117 (1995) 2401. [8] a. K. Tanaka, Y. Okahata, J. Am. Chem. Soc. 118 (44) (1996) 10679; b. Y. Okahata, T. Kobayashi, K. Tanaka, Langmuir 12 (1996) 1326. [9] M.A. Cohen Stuart, R.A.J. Wegh, J.M. Kroon, Langmuir 12 (1996) 2863. [10] N. Vranken, M.V. der Auweraer, F.C. De Schryver, H. Lavoie, C. Salesse, Langmuir 18 (2002) 1641. [11] S. Kirstein, H. Mohwald, M. Schimomura, Chem. Phys. Lett. 154 (4) (1989) 303. [12] L.G.T. Eriksson, P.M. Claesson, S. Ohnishi, M. Hato, Thin Solid Films 300 (1997) 240. [13] R.C. Ahuja, P.L. Caruso, D. Mobius, Thin Solid Films 242 (1994) 195. [14] P. Lehmann, D.G. Kurth, G. Brezesinski, C. Symietz, Chem. Eur. J. 8 (2001) 1646. [15] J. Schnitter, A. Engelking, R. Heise, D. Miller, H. Menzel, Macromol. Chem. Phys. 201 (2000) 1504. [16] J.C. Powers Jr., W.L. Peticolas, J. Phys. Chem. 71 (1967) 3191. [17] F. Zimmermann, B. Hossenfelder, J.C. Panitz, A. Wokaun, J. Phys. Chem. 98 (1994) 12796. [18] J.M. Kovaleski, M.J. Wirth, J. Phys. Chem. 99 (1995) 4091.
In-situ observation of the aggregated morphology and interaction of dialkyldimethylammonium bromide with DNA at air/water interface by Brewster angle microscopy Lu Sun, Miao Xu, Xueliang Hou, Lixin Wu * Key Laboratory for Supramolecular Structure and Materials of Ministry of Education, Jilin University, Changchun 130023, PR China Received 14 July 2003; accepted 3 October 2003
Abstract The adsorption of DNA on the Langmuir film of a cationic surfactant, dioctadecyldimethylammonium bromide (DODABr), and the change of the aggregation morphology of the composite monolayer with respect to surface pressure have been investigated by Brewster angle microscopy (BAM). In contrast with the case of DODABr on pure water subphase, when DNA was dispersed into subphase, its adsorption to the interface monolayer through electrostatic interaction decreases the charge density and therefore promotes the formation of domain at low surface pressure. In addition, the electrostatic interaction changed the phase morphology of DODABr Langmuir monolayer under different surface pressure, that is, from flower-shaped crystalline domain on the pure water subphase to circular domain on the subphase dispersed with DNA. The result also shows that the monolayer of the composite at air/water interface under the high pressure is not homogeneous, but consists of incompletely fused domains. For the Langmuir film of the surfactant with shorter alkylchains, similar morphology can be observed both under the high and low surface pressure. But the tight-stacked circular domain is no longer observed. D 2003 Elsevier B.V. All rights reserved. Keywords: Air/water interface; Brewster angle microscopy; Surfactant; DNA; Complex
1. Introduction Due to its double helical structure and negative charged phosphate backbone, DNA is easy to form electrostatic complex with cationic surfactants [1,2]. This kind of cationic surfactant-DNA complex supporting with polymer micelle matrix has been used for gene transfer in vivo, and it also has been found very promising in the application of bio-function materials and biosensors [3 –6]. The cationic surfactant-DNA complex displays various aggregation morphologies under different conditions. The fluorescent microscopy has shown that the DNA molecule will gradually condense from linear to globular when it forms a complex with cationic surfactant in aqueous solution [7]. In organic solvent, the surfactant-DNA complex is proposed mostly existing in globular aggregate state due to hydrophilic and * Corresponding author. Tel.: +86-431-8499649; fax: +86-4318980729. E-mail address: [email protected] (L. Wu). 0167-577X/$ - see front matter D 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.matlet.2003.10.010
hydrophobic equilibrium. Okahata et al. have proved that DNA molecule adsorbs to the cationic surfactant in a rodlike form at the air/water interface. When the subphase was dilute dye-intercalated DNA aqueous solution, the Langmuir monolayer containing dye molecules at air/water interface could be transferred to solid substrate with the orientation [8]. However, as we know, few investigations related to the direct observation about aggregation morphology of the complex without introducing probe molecules under various surface pressures. Brewster angle microscopy (BAM) is a suitable technique for in-situ observation of the microdomain structure of phase separation at gas/liquid or liquid/solid interface without probe [9]. So it provides an effective method to check the interaction at interface. In this report, we present the measurement of in-situ phase transformation of double-chain cationic surfactant monolayer on DNA subphase. In this paper, aggregation and morphologic changes of the surfactant-DNA Langmuir monolayer accompanied with the increasing of the surface pressure were investigated.
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2. Experimental 2.1. Materials The sodium salts of fish testes DNA were purchased from AMRESCO Chemical and used as received. Didodecyldimethylammonium bromide (DDDABr), dioctadecyldimethylammonium bromide (DODABr) and acridine orange (AO) are the chemicals from ACROS Chemical. Other organic solvents are the analytical grade and the water for subphase is redistilled deionized water. 2.2. Instrument and measurement Langmuir trough (622D) used in the experiment is a production of Nima England. BAM (I-Elli2000) is from NFT Germany, which is equipped with a 50-mW semiconductor laser, an analyzer, a zooming microscope with a high sensitive CCD camera and a video recording system. The wavelength of p-polarized light is 532 nm. The lateral resolution reaches 2 Am. The scale of the images is 870650 Am. BAM observation to Langmuir monolayers was carried out by spreading 30 Al of DODABr chloroform solution (1.16 mg/ml) onto air/water or air/solution interface with pure water, DNA (3.5410 6 mol/l in base pair) and the mixture of DNA with AO in a molar ratio (base pair) 3:1 aqueous solution as subphases, respectively. The k – A isotherms and corresponding BAM images were recorded after standing 20, 60 and 60 min with above subphases in order to make the organic solvent evaporate off completely and to reach their equilibrium states, respectively. Repeated experiments for DDDABr (1.28 mg/ml) were under the same conditions as those for DODABr. The pH value of the subphase was maintained at 6 –7, and the temperature was controlled at 20F0.5 jC.
3. Results and discussion 3.1. p – A isotherms of DNA-surfactant complex system Fig. 1A shows k –A isotherms of DODABr on different subphases. The collapse pressure of DODABr on the pure water is about 43 mN/m. The phase transition can be clearly observed in the range of 25 –35 mN/m, which is in agreement with the reported results [10]. Vranken et al. have discussed the differences of k –A isotherms of DODABr performed by different research groups [11,12]. More in general, any impurities existing in the sample or the subphase could influence the shape of the isotherm and phase transition pressure when the spreading amount is large enough. However, neither have DODABr in our samples nor any others from the related literatures been presented details on the definite amount of impurities, which affect the results [10]. So we measure the k – A isotherms of
Fig. 1. (A) k – A isotherms of DODABr on subphase of (a) pure water, (b) DNA aqueous solution in which the concentration is 3.5410 6 mol/l and (c) DNA and AO aqueous solution which concentration proportion is 3:1. (B) k – A isotherms of DDDABr on subphase of (a) DNA aqueous in which the concentration is 3.5410 6 mol/l and (b) DNA and AO aqueous solution in which the concentration proportion is 3:1.
DODABr several times under preset experiment conditions. The results can be repeatedly pretty well and the k –A curves are quite similar to the reported shapes [13]. In contrast to the pure water subphase, the k –A isotherm of DODABr shows that the preliminary area increases and collapse area decreases on the subphase of DNA aqueous solution. The collapse pressure is up to 64 mN/m and the phase transition still existed. When the subphase was changed to the molar ratio MDNA/MAO=3:1 of DNA and AO aqueous solution, the k – A isotherm of DODABr shows little change comparing with that of it on DNA subphase. As a negative charged biopolyelectrolytes, DNA can absorb onto the surfactant with positive charges at air/ water interface forming electrostatic complex. That makes larger the molecular area due to the binding of DNA occupying larger surface area and smaller the charge repulsive effect, which lead to the change of stacking property of surfactant monolayer with surface pressure. Such a property is similar to the interaction of surfactant on general polyelectrolyte subphase as described in literature [14,15]. In order to understand the influence of electrostatic interaction between DNA and Langmuir monolayer on water, DDDABr, the surfactant with short chains that is
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more sensitive to interaction at interface, has been used for this purpose. It has poor spreading property on pure water subphase because it will dissolve in water during the pressing due to its strong hydrophilic feature. But this feature is much sensitive to the subphase chemical environment and become stable at the interface especially when there is anionic polyelectrolyte in the subphase. DDDABr forms stable monolayer on DNA aqueous solution due to the electrostatic interaction between them, which reduces its hydrophilic feature, as shown in Fig. 1B. However, the stability of DDDABr again decreases greatly after adding AO to DNA aqueous subphase, which is quite different from the case of DODABr and shows the sensitivity of DDDABr to the change of DNA aqueous subphase. One of the possible reasons is that the addition of AO neutralizes the negative charges of DNA and that leads to the decreasing of the interaction between DNA and surfactant.
30– 50 Am (Fig. 2B). It is noted that the patterned domain is quite uniform both in scale and in shape, which is different from larger and anisotropic leaf-like domains as reported in literature on high concentration, indicative of the growth of crystalline domains being a controllable dynamic process. From this point the variety of the patterned domain should be derived from the change of the temperature, barrier speed and so on. And further more the flower-like domains change back to dot structure with surface pressure increasing (Fig. 2C). Around the dot structure on the air/water interface is liquid condensed phase of DODABr based on increasing gray level, indicating that the Langmuir monolayer is composed of two phases when the surface pressure closes to collapse point. For DDDABr, no clear morphologic structure on pure water subphase has been observed because of its strong hydrophilic property (the surface pressure only getting to 9.3 mN/m).
3.2. In-situ BAM observation to the surfactant monolayer on pure water subphase
3.3. In-situ BAM observation to the surfactant monolayer on DNA subphase
The BAM image of DODABr on the pure water subphase shows that there is no visible morphologic structure at the air/water interface when the pressure just starts to increase, while the gray level getting high. Dot domains with diameter in the region of 8 – 10 Am can be observed when pressure reaches 20 mN/m (Fig. 2A). Both the amount and the size of domains increase as pressure getting high. When the pressure is up to 30 mN/m, flower-like domains have been observed and their diameter falls into the range of
Corresponding to the k – A isotherms in Fig. 1A(b), under lower surface pressure below the phase transition (Fig. 3A) such as 11 mN/m, dot domains have been observed and the amount of the dots increases with the pressure increasing. It is clear that DNA promotes the formation of dot domains at low pressure because the dot domain normally emerges at much higher surface pressure on pure water. Around the phase transition the sizes of domains can be observed evenly and the amount of the domains increases quite much. As the
Fig. 2. BAM Images of DODA aggregate on the pure water at surface pressures of (A) 21 mN/m, (B) 30 mN/m, (C) 39 mN/m, respectively, and (D) DDDA on the DNA and AO aqueous subphase at surface pressure of 9.3 mN/m.
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Fig. 3. BAM Images of DODABr aggregates on the DNA aqueous subphase at surface pressures of (A) 11 mN/m, (B) 22 mN/m, (C) 34 mN/m, (D) 39 mN/m, and on the DNA and AO aqueous subphase at surface pressure of (E) 35 mN/m and (F) 40 mN/m, respectively.
pressure higher than that of the phase transition, no flowerlike domains but big circular domains have been observed (Fig. 3C), and the size distribution quite uniform. We believe that the condensation of DNA and the formation of the DODA-DNA complex through electrostatic reaction restrain flower-like crystalline domain of DODABr but enhance the circular domain and then promotes the phase transition. Aggregation of DNA presents the biological significance because it is important to gene transfer and cellular karyokinesis. So it helps us to understand the process of DNA going through cellular membrane though the aggregation of DNA at air/water interface is different from that occurred in aqueous solution. With the surface pressure increasing from phase transition region to collapse point, the surface morphology of DODABr monolayer changes from circular domain to homogeneous domain (Fig. 3D). In this process, we confirm that the aggregation of DNA should be influenced by surface pressure, that is, the density of positive
charge, which is similar to that of DNA going through the cellular membrane. On the subphase of DNA with intercalated AO, the sizes of DODABr aggregate domains become smaller and denser (Fig. 3E,F) in contrast to the aggregates of DODABr on the DNA subphase and uniform Langmuir monolayer near the collapse pressure. As has been well known, AO adsorbs electrostatically and intercalates into the double helix of DNA leading to the change of surface charge and existing state of DNA in water [16 –18]. So we believe that the addition of AO influences on the formation of the DNA-DODA complex and the aggregated morphology. The effect of charge and agglomeration to the surface morphology is not clear now. DDDABr also adsorbs DNA through electrostatic interaction at the air/water interface forming interface complex, which makes more stable its Langmuir monolayer and much smaller the aggregate domain. At very low surface pressure dot domains of DDDABr monolayer have been observed on
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the subphase of DNA mixed with AO. In contrast to the situation of DODABr, the dot domain of DDDABr, appeared at low surface pressure such as 9.3 mN/m, as shown in Fig. 2D, does not grow larger with the surface pressure increasing until collapsed. In addition, DDDABr shows shorter responding time and lower responding pressure indicating the difference of the lengths of the alkyl-chains between DODABr and DDDABr because dot domain is immediately observed for DDDABr but not for DODABr as it needs a while and can be observed on a certain pressure.
4. Conclusion From above results, we confirm that DODABr and DDDABr form complexes with DNA at the air/water interface, respectively. Through in-situ observation by BAM, we find that the agglomeration of DNA strongly influence on the interface property of surfactant. Through its electrostatic interaction to DNA, the aggregated morphology of DODABr at air/water interface changes from small dot domains to a little bit larger uniform circular domains while DODABr itself changes from dot structure to flowerlike domain and then to dot structure again on pure water. The similar action is observed to more hydrophilic DDDABr with DNA at air/water interface. The intercalation of dye to DNA changes the morphology of aggregated domains of Langmuir monolayer.
Acknowledgements We gratefully acknowledge the financial support from the National Natural Science Foundation of China
(29992590-5), the Major State Basic Research Development Program (Grant No.G2000078102) and the Ministry of Education of China.
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