Temperature dependent micellization behavior of as synthesized anionic SAILs in aqueous nonionic polymer solutions: conductivity, UV-visible probe and antimicrobial studies

Abstract As synthesized surface active ionic liquids (SAILs), Tetrapropylammonium Dodecylsulfate (TPADS), and Tetrabutylammonium Dodecylsulfate (TBADS) have been characterized by FTIR, 1H-NMR and 13C-NMR spectroscopic techniques. Conductometric and UV-Visible probe investigations have been attempted to study the aggregation behavior of these synthesized SAILs in aqueous solution of polymers Polyvinylpyrrolidone (PVP) and Polyethylene glycol (PEG) at different temperatures and concentrations. Two breaking points were identified from κ vs [SAILs] plots. First, the critical aggregation concentration (CAC) is recognized with the polymer-SAILs binding and the second being regarded as critical micellar concentration (CMC), suggested to represent saturation point for Polymer-SAIL aggregation. Thermodynamic parameters of micellization (ΔGmo,ΔHmo andΔSmo) are rationalized in terms of interactions prevailing between hydrophobic and hydrophilic regions of SAIL-Polymer system. Micellization process for the blend of SAIL-PVP is found to be more favored than SAIL-PEG at all temperatures. Further, the effect of PVP on the antimicrobial activities of TPADS/TBADS has been tested by measuring zone of inhibition which accounts for higher effectiveness of PVP-SAILs system against gram positive bacteria (Bacillus cereus and Staphylococcus aureus) as compared to gram negative bacteria (Pseudomonas aeruginosa). GRAPHICAL ABSTRACT


Introduction
Ionic liquids (ILs) are compounds of non-molecular organic/inorganic molten salts with melting point <373 K or even at room temperature due to the irregular packing of asymmetric bulky cations in crystal form.These liquids are known to possess interesting features, like low toxicity, high thermal stability and ionic conductivity, low vapor pressure, good solvent abilities, recyclability, non-flammability, and antimicrobial activity etc. [1][2][3] Because of such unique properties, ILs have the potential applications in various forms, like as solvent for nanomaterial synthesis, polymerization reactions, [4,5] as catalyst in organometallic synthesis and enzymatic processes, [4] as versatile novel lubricants for ceramic pairs of steel with aluminium/copper/silicon, as stationary phases in liquid and gas chromatography, [6,7] in separation and extraction science for liquid-liquid and metal ion extraction, [4,8] as diverse electrolyte in double layer and hybrid supercapacitors, [9,10] fuel and photochemical cells, [11,12] and as electrochemical sensors [13] etc. Furthermore, ILs have been modified as surface active ionic liquids (SAILs) having long hydrophobic alkyl chains which lead to their aggregation behavior at concentration > CMC as well as in encapsulating the environmental sensitive species, such as DNA, proteins, enzymes, and drugs etc. [14] Most of these SAILs have imidazolium and tetraalkylammonium groups as cationic parts.[16][17][18][19] However, imidazolium cation and halogen anions based SAILs have many environmental issues in comparison to SAILs bearing nitrate, sulfate, alkyl sulfonate as anionic parts and tetraalkyl ammonium group as cationic part. [20,21]Also, Low cost, biodegradable nature, antimicrobial and antibacterial properties of alkyl sulfate anion based tetraalkylammonium SAILs make them suitable candidate for variety of applications such as wood preservatives, antimicrobial properties, for solublization of hydrocarbons, catalytic conversion of green house gas CO 2 to useful solvent propylene carbonate etc. [22][23][24][25][26] Additives, like electrolytes, amino acids, proteins, polymers etc. are known to bring improvements in the properties of SAILs.[32] More specifically, polymer-surfactant aggregation occurs at a concentration identified as critical aggregation concentration (CAC).These polymer-surfactant aggregates (PSAs) have interesting characteristics of phase forming behavior in the bulk solution, comprising of precipitation zone and turbidity zone, depending on the ratio of polymers and surfactants with overall charge neutrality of PSAs.Additionally, the repulsions induced between electrostatic double layers present in these charged moieties also lead to the formation of stable dispersed phase.Hence, these clusters have potential to substitute the use of ultrafilteration membrane in their effect to remove heavy metal ions from dilute solutions. [33,34]These important characteristic features of PSA systems, has interested us to undertake investigations on self -aggregation behavior of tetraalkylammonium based synthesized SAILs TPADS and TBADS with the help of conductometric and UV-Visible probe studies along with antimicrobial action of TPADS/TBADS-PVP blend in aqueous medium as well as the variations of polymer concentration and temperature.

Synthesis
Equimolar SDS and TPABr/TBABr were dissolved in a particular amount of Dichloromethane (DCM) and the reaction system was magnetically stirred for 4 hrs at room temperature according to Equation (1).
Residue as left was filtered with the help of whatmann filter paper.The filtrate (i.e., organic layer) was washed with doubled distilled water till no pale yellow precipitates of AgBr were obtained in aqueous layer as tested with 0.1 M AgNO 3 .The Organic layer was dried in a vacuum oven for 48 hours in order to remove excess solvent content; the purity of the as obtained ionic liquids was confirmed from spectroscopic analysis. [20,22,35]

Conductivity measurements
Oakton700 Conductivity meter (Eutech Instruments, Thermo Fischer) was used for the determination of conductivity (j) measurements of the SAIL-Polymer mixtures.Conductivity cell was calibrated with KCl aqueous solutions having conductivities �84 and 1413 � 10 À 6 S�cm À 1 .Solution temperature was maintained within ± 0.1 � C by circulating thermostated water through it.The specific conductance of SAILs in 0.0%-0.3%w/w aqueous solution of polymers was measured over a range of temperature (293.15-313.15)K at an interval of 5 K. Shimadzu balance with a precision of ± 0.0001 g has been used for preparing all solutions. [36]

UV-visible probe studies
UV-VIS absorption spectra have been recorded with Genesys 10S Spectrophotometer (Thermoscientific, USA) using 2 � 10 À 3 M pyrene solution as spectroscopic probe.Quartz cuvette of path length 10 mm path length was employed for SAIL-Polymer sample analysis.Absorbance spectra was obtained in the wavelength range (200-400) nm at room temperature. [37]

Antimicrobial study
The antimicrobial activities of ionic liquids TPADS/TBADS in 0.2% w/w PVP was studied using 'well diffusion method'.Cultured Petri dishes containing nutrient broth was used for growing selected bacterial strains on it.Sterile borer was utilized for creating wells on the plates and different concentrations of SAILs in PVP poured with the help of micropipette into selected wells.For bacterial growth incubation of plates was done at 37 ± 0.5 � C for 24 hours.The Zone of Inhibition of plates (in mm) has been recorded. [38] Results and discussion

FTIR
Characterization of as synthesized samples was made via FTIR technique using L1600312 TWOLITA/ZnSe FTIR spectrometer supplied from Agilent Technologies.The spectra of TPADS and TBADS have been given in Figure 1 and Figure S1.The detailed information about the various peaks at different wave number has been assigned as, 2957-2922 cm À 1 for the asymmetric and symmetric stretching of the -CH 2 and -CH 3 groups of tetraalkylammonium cation and dodecylsulphate anion, C-H bending vibration at 1465 and 1376 cm À 1 showing the presence of long alkyl chain, 1252-1215 cm À 1 due to the asymmetric stretching of OSO 3 -, 1033-873 cm À 1 due to the symmetric stretching of the OSO 3 -, and 1107-1101 cm À 1 due to the stretching of C-N, showing the presence of ammonium cation. [39,40]

1 H and 13 C NMR
1 H-NMR and 13 C-NMR analysis were carried out with JNM ECX-500 NMR spectrometer, Jeol India with 500 MHz resonating frequency using D 2 O as a solvent.Chemical shifts values for 1 H-NMR and 13 C-NMR of TPADS have been given in Tables 2 and 3.
In 13 C-NMR of TPADS, the value of chemical shift at 10.05 ppm has been due to terminal (-CH 3 ) methyl groups carbon, 14.98 ppm due to the (-CH 2 -) methylene carbon attached to terminal methyl group and 59.89 ppm due to the methylene carbon attached to the positively charged nitrogen atom (-CH 2 -N þ ) and hence provide deshielded value of chemical shift in TPADS.
In DS -anion, chemical shift at 13.859 ppm due to terminal methyl group (-CH 3 ), 22.7 ppm due to the methylene carbon (-CH 2 -) attached to terminal methyl group, 67.97 ppm due to the methylene carbon attached to oxygen atom (-CH 2 -O) have been shown in the Table 3.Also the information about the 1 H-NMR and 13 C-NMR of TBADS has been provided in supporting information Tables S1  and S2.

Conductivity studies
The conductometric method is an incredibly efficient tool for explaining the micellization process of amphiphiles in terms of CMC values and thermodynamic parameters of SAILs TPADS/TBADS in aqueous polymeric solutions.Representative conductivity plots i.e., specific conductance (j) vs concentration of SAILs (TPADS and TBADS) in the Table 3. 13 C NMR spectrum of TPADS with chemical shift values of different carbon atoms.aqueous solution of PVP/PEG at different temperatures are given in Figure 2 and Figure S2.As the concentration of SAIL increases, its monomeric form begins to form self organized assembly called micelles.However, there is only one break point observed, which has been used to obtain the value of critical micellar concentration (CMC) for TPADS/TBADS in aqueous medium (Figure 1(a) and Figure S2 (a)).Whereas, in case of TPADS/TBADS in PVP/PEG .78 [22]4.96 0.173 [22] À  [22] 0.043 [22] À 13.250 313.15 0.584 À 37.32 À 28.570.028 À 8.768 0.656 À 37.78 À 38.87 À 0.003 [22] 0 aqueous solutions, the j with [SAIL] plots show two break points; the first one is regarded as beginning of interactions and aggregates formation among surface active ionic liquids and polymers due to different type of interactions and is known as critical aggregation concentration (CAC) thereof, and the second break point may be referred to as critical micellar concentration (CMC) i.e., the point where polymer molecules saturated by SAIL and at concentration > CMC, dynamic equilibrium has occurred between regular micelles and polymer-surfactant aggregates. [20,30,31,41,42] and Table S3.The CMC values for these SAILs tend to fall with increase in the concentration of polymers PVP/PEG.In case of PVP however, this behavior may be because of the hydophobicity of polymer and reduction in the electrostatic repulsions between the polar head groups of SAILs as a result of electrostatic interactions between the partial positive charge of PVP (i.e., N dþ center on pyrrolidone unit due to polar nature) and negatively charged polar head groups of tetraalkylammonium dodecylsulphate (TAADS), hence favors micellization process.However, with increase in the concentration of PVP, hydrophobicity and electrostatic interactions build up more aggressively resulting in lowering of CMC values to appreciable extent. [32,43]In case of PEG, it seems that the water molecules around the hydrophobic groups get reinforced through hydrogen bonding between PEG and water, moreover the non-polar portion of polymer directed toward hydrophobic alkyl chain of SAILs causing increase in hydrophobicity, hence favors micellization process.The CMC values of these SAILs in an aqueous solution of PVP are less as compared to PEG, due to its more hydrophobicity along with lowering of the electrostatic repulsion among the head groups of micelle. [32,44]In case of imidazolium based anionic SAILs in aqueous solution of nonionic polymers, opposite behavior has been observed for the micellization process. [20]he temperature effect on CMC of SAILs has been evaluated in terms of two major factors: (1) decrease in the hydrophilic hydration and (2) decrease in hydrophobic hydration due to the disruption of ordered water structure around the hydrophobic groups.In the present case, the former factor seems to dominate in the temperatures 293.15 and 298.15K resulting decrease in CMC values, while the latter dominates in the temperature range from 298.15 to 313.15 K as a result of this CMC values increase in magnitude, explaining the dip observed at 298.15 K, Figure 3. [22,45] Another important information as revealed from these studies is that TPADS has higher CMC values than TBADS in all studied systems, because of the increase in the counter-ion size from TPA þ to TBA þ .These counter-ions act as spacers between the anionic head groups of DS -, so greater the counter-ion size, larger would be spacing between the anionic head groups and lesser would be the electrostatic repulsions among them, hence facilitating the micellization process. [22,46]

Thermodynamics of micellization process
Standard thermodynamic parameters, namely standard Gibbs free energy of micellization (DG o m ), standard enthalpy of micellization (DH o m ) and standard entropy of micellisation (DS o m ) have been evaluated by using following Equations (2-4) [22,[36][37][38] and tabulated in Table 5.
where a is the degree of counter-ion dissociation calculated by using relation ¼ S 2 � S 1 ; S 1 and S 2 are slopes of pre-micellar and post-micellar regions in the j vs [concentration] plot.dðlnX CMC Þ=dT has been calculated by using differential form of second order polynomial.becomes more negative favoring micellization process to greater extent for TBADS than TPADS. [22,47]The higher negative DG o m values for PVP suggests that micellization process is more favored in case of PVP than PEG for both TPADS and TBADS.The positive DH o m values at temperatures 293.15K and 298.15K for all studied systems suggest that higher energy is required to break the ordered hydrogen bonded water structure around hydrophobic groups i.e., DS - and TAA þ .These results lead to conclude that hydrophobic   interactions favor micellisation as usual, [22] and the process is entropy driven.However, at temperature >298.15K, decrease in the DS o m values with the increment in PVP/PEG concentration, perhaps is due to the energy requirement to disrupt the network of water structures.Hence, DH o m values become more negative in the temperature range 303.15K À 313.15K for SAILs in polymeric solutions as compared to aqueous solutions.The observed process of micellization appears to be entropy driven at lower temperatures and enthalpy driven at higher temperatures for all studied systems.Also, the values of DS o m decrease with rise in temperature from 293.15 K to 313.15 K, which may be due to the clustering of surfactant molecules to form micelles.The values of DS o m show an increase with increment in PVP/PEG concentration at temperatures up to 298.15 K, which may be due to the rupture of the ordered structure of polymer-water around alkyl chains during the transfer of alkyl chains from solvent to the interior part of micelles. [44,45]Figures 4 and 5 and Figure S3 represent the variation of thermodynamic parameters with temperature and polymer concentration.From Figure 6, it is clear that entropy has more contribution toward DG o m as compared to enthalpic contribution at low temperature, whereas reverse trend follows at higher temperature. [38]Enthalpy-Entropy compensation phenomenon has a linear relationship between DH o m and DS o m as described by the following equation [22,37,44] :  S4). [22,48]Enthalpy-Entropy compensation curves for TPADS in 0.2% w/w PVP and PEG have also been given in Figure S4.

UV-visible probe studies
Pyrene has been used as probe molecule, as it provides wonderful information regarding the interactions among pyrene and surfactant molecules.Pyrene has been shown to have 12 multiple peaks in UV-Vis spectrum with strong peaks observed at 242, 272, 320, and 335 nm due to the multiple ring structure of this hydrophobic molecule. [37,38]For calculating the CMC of surfactants, the plot between total absorbance A T (i.e., the sum of absorbance values at four strong peaks of pyrene) and concentration of surfactants has been used.At lower surfactant concentration, because of hydrophilic environment, hence very small increment in the value of absorbance is obtained.But as the surfactant concentration reaches to CMC, there has been sudden increment in the value of absorbance due to the more incorporation of pyrene molecules in the non-polar hydrophobic interior of micelles.Boltzmann sigmoidal fitting has been used to evaluate CMC of surfactants wherein it has been found that total absorbance increases in sigmoidal manner with surfactant concentration (Figure 7 and Figure S5) as represented below by the relation: where x, a i , a f , x o , and Dx are surfactant concentration, sigmoidal initial and final limits, center of sigmoidal and independent variable i.e., x interval, respectively.Two CMC values have been obtained from the sigmoidal plot, that is at x o and ðx o þ 2DxÞ: The ratio, x o = Dx has depicted the correct value of CMC.If x o = Dx < 10, the CMC is given by x o ; otherwise the value is ðx o þ 2DxÞ: [49,50] In every case, the A T values found to be higher on addition of PVP/PEG as compared to SAILs in aqueous solution; hence more hydrophobic interactions are inferred. [38]The values obtained for CMC have been given in Table 4 and Table S3.

Antimicrobial studies
The antimicrobial study of SAILs TPADS and TBADS has been performed in 0.2% w/w aqueous solution of PVP against gram positive bacterial stains (B.cereus and S.aureus) and gram negative bacteria P. aeruginosa; Gram negative bacteria have membrane on outer side of peptidoglycan layer which is composed of lipopolysaccharides and phospholipids, whereas gram positive bacteria have no such membrane.This outer membrane in bacterial cell structure has the ability to protect them from intruding compounds, like TPADS/TBADS. [51,52]As shown in Table S5, on addition of 8.8 � 10 À 3 M and 6.6 � 10 À 3 M stock solution of TPADS/TBADS, respectively, from 20 to 50 lL; there occurs appearance of zone of inhibition which shows upsurge in the zone of inhibition values with rise in SAILs concentration meaning thereby greater action against microbes.This might be due to charge borne by the surfactant and concentration confinement, hence more effective interaction with bacterial cell membrane of strains resulting into their destruction. [53]Also, out of TPADS and TBADS, latter one has been found to be more effective against all the three bacterial strains in aqueous as well as aqueous polymeric solutions.Both TPADS and TBADS have efficient antimicrobial potential against gram positive bacteria as compared to gram negative bacteria may be due to additional layer of peptidoglycan.Zone of inhibition values and variation for TPADS and TBADS in 0.2% w/w aqueous solution of PVP have been given in Figure 8 and Table S5 along with pictures in Figure 9.It may be mentioned that SAILs and their composition with PVP showed no antimicrobial activities against bacterial strains S.typhi and E.coli.

Conclusion
As synthesized tetraalkylammonium Dodecyl SAILs have been characterized by FTIR and NMR spectroscopic techniques. [22]The focus of present study is to investigate the micellization of these benign SAILs in the presence of nontoxic and biodegradable polymers such as PVP and PEG.
From conductivity and UV-VIS probe investigations reveal the impact of polymers PVP and PEG on micellization behavior of anionic SAILs TPADS/TBADS.The CMC values for TBADS have been found to be lower than TPADS for all studied systems.Moreover, studies indicate that PVP facilitated the micellization for SAILs as compared to PEG, for electrostatic reasons.Thermodynamics of micellization has provided the information about spontaneity of the process along with entropic and enthalpic dependence of micellization processes with respect to temperature.Microbial studies reveal that SAILs with PVP have been found more effective against gram positive bacterial strains as compared to pure SAILs in aqueous solution.

Figure 6 .
Figure 6.Representative plots of TPADS for the contribution of DH o m and À TDS o m toward DG o m in aqueous solution of (a) 0.2% w/w PVP and (b) 0.2% w/w PEG.
) here T C represents compensation temperature and refers to solute-solvent interactions and de-solvation during micellization process, DH � m represents the chemical index of micellization process and characterize the solute-solute interactions.The DH o m vs DS o m plots provide DH � m and compensation temperature T C : DH � m is the change in enthalpy when DS o m ¼ 0, and quantifies the micelle stability.A reasonably good compensation between DH o m and DS o m values with correlation coefficient of 0.999 is obtained in the present work.The values of T C have been found to lie in the range 305-315 K (Table

Figure 8 .
Figure 8. Zone of inhibition values (in mm) for TPADS and TBADS in water and aqueous solution of 0.2% w/w PVP against (a) B.cereus (b) S.aureus and (c) P.aeruginosa.

Table 1 .
Specifications of used chemicals.