Hydrophobe Interface versus Partition into the

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Adsorption at Water/Hydrophobe Interface versus Partition into the Interior of the Hydrophobe: Quantitative Evaluation of the Solute Retention Selectivity at Water/Hydrocarbon Interface Keisuke Nakamura, Shingo Saito, and Masami Shibukawa J. Phys. Chem. C, Just Accepted Manuscript • DOI: 10.1021/acs.jpcc.7b12413 • Publication Date (Web): 02 Feb 2018 Downloaded from http://pubs.acs.org on February 5, 2018

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The Journal of Physical Chemistry C is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

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The Journal of Physical Chemistry

Adsorption at Water/Hydrophobe Interface versus Partition into the Interior of the Hydrophobe: Quantitative Evaluation of the Solute Retention Selectivity at Water/Hydrocarbon Interface

Keisuke Nakamura, Shingo Saito, Masami Shibukawa*

Graduate School of Science and Technology, Saitama University, 225 Shimo-Okubo, Sakura-ku, Saitama, 338-8570, Japan

*Corresponding

Author.

Tel:

+81-48-858-3520.

Fax:

[email protected]

-1-

ACS Paragon Plus Environment

+81-48-858-3520.

E-mail:

The Journal of Physical Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ABSTRACT The molecular level understanding of the chemistry at water/hydrophobe interface is crucial to chemical separation processes in aqueous media such as reversed-phase liquid chromatography and solid-phase extraction. However, direct observation of the behavior of molecules and ions at the interface in the reversed-phase separation systems still remains a major challenge and the probing techniques which provide the spatial information of the distribution of molecules and ions are required. In this paper, the molecular distribution between the aqueous solution and alkyl bonded silica particles is studied by surface-bubble-modulated liquid chromatography (SBMLC). We determine the distribution coefficients of various organic compounds referring to accumulations onto the water/alkyl chain interface and into the alkyl chain layer from the bulk water by SBMLC. The bulk water-to-alkyl chain layer distribution coefficient is corrected for the contribution of the end-capped silica surface to the solute retention using end-capped C8 and C18 bonded silica columns. The experimental data provide a picture of the spatial distribution of organic molecules in alkyl bonded silica particles exposed to water. It has been revealed that the water/alkyl chain interface exhibits quite different accumulation selectivity for organic compounds from the interior of the alkyl chain layer and the overall separation selectivity of the reversed-phase systems is determined by the relative sizes of the aqueous/hydrophobe interface and the hydrophobe itself.

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INTRODUCTION Various hydrophobic materials are now available for analytical and preparative scale separations of chemicals in aqueous solutions.1,

2

They are usually utilized as column packing

materials for reversed-phase liquid chromatography (RPLC) and reversed-phase solid phase extraction (RP-SPE). However, despite their importance and widespread application in chemical separation, the characterization of selective accumulation of solute compounds in aqueous media with hydrophobic materials remains fraught with uncertainties. A central question lingers about whether solute molecules adsorb on the water/hydrophobe interface or partition into the interior of the hydrophobe.3-10 Alkyl-bonded silica is the most widely used material in RPLC and RP-SPE so that extensive works have been devoted to revealing whether solute molecules are retained at water/alkyl chain interface or in alkyl chain layer bonded to silica. Spectroscopic investigations using nuclear magnetic resonance (NMR),11-13 Infrared (IR),14, 15 and Raman spectroscopy16, 17 have revealed the structure of the alkyl chains bonded to a silica surface in contact with water-organic mixtures such as water-acetonitrile and water-methanol as well as in a dry state. The distribution of the organic modifier into the alkyl chain layer has also been inferred from the change in structure of the alkyl chain layer. Montgomery et al.18 carried out a fluorescence spectroscopy study on orientational dynamics of a hydrophobic guest in a C18 surface using 1,4-bis(o-methylstyryl)benzene as a probe and showed that the large hydrophobic probe molecule resides at the surface of the chains. Henry et -3-



al.19 obtained the experimental evidence for adsorption of acetonitrile at a model interface, water/octadecylsiloxane and water/quartz, using sum-frequency generation spectroscopy. The mechanism of the retention process in RPLC has been extensively studied using molecular dynamics (MD) simulation in recent years since it can give a microscopic picture of local structure and dynamics of the RPLC interface on an atomic scale.20-25 Although most of the papers have been written about the RPLC retention process by adsorption of the solute molecule to the interface or by partition into an alkyl bonded layer, the simulation work has shown that it is neither of these two extremes but depends on the nature of the molecule; hydrophobic molecules are retained by both partition and adsorption, while the preferred location of a molecule with a hydrophilic group is the solution/alkyl chain layer interface for the solvent of pure water and the aqueous solutions including low concentrations of an organic modifier.22 It is now generally assumed that both adsorption onto the solution/alkyl chain layer interface and partition into the alkyl chain layer simultaneously contribute to the solute retention in reversed-phase systems.21-23, 26, 27 However, the respective contributions of the interface and the alkyl bonded layer itself as the components of the hybrid stationary phase to the accumulation of solute molecules in aqueous media have never been evaluated quantitatively by direct experimental measurements. Since the water/alkyl chain interface and the alkyl bonded layer may exhibit different solute accumulation selectivites from each other, the quantitative evaluation of the respective contributions of the interface and the alkyl chain layer itself to the retention on the -4-


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column packed with alkyl bonded silica is required to understand and/or predict the separation behavior of chemical compounds in the reversed-phase systems. The measurements of the retention volume of a solute compound by liquid chromatography cannot directly give a microscopic picture of the location of the solute molecule in the separation system because the value of the retention volume represents the overall magnitude of retention of a target compound in the column. However, we should be able to find out where and how much the solute compound is accumulated or retained by chromatographic measurements if the following requirements are fulfilled: (1) it can be assumed that each component of the hybrid stationary phase independently accumulates a solute compound, (2) the volumes or areas of the components of the hybrid stationary phase, i.e., the water/alkyl chain interface and the alkyl chain layer in the case of RPLC, can be determined, and (3) the volume or area of each component of the hybrid stationary phase can be changed independently of each other. Recently we presented a new type of liquid chromatography, surface-bubble-modulated liquid chromatography (SBMLC), which has a hybrid stationary phase consisting of gas phase as well as the alkyl bonded layer and the water/alkyl chain interface.28 We demonstrated that the stationary gas phase can be fixed in the pores of the hydrophobic porous material in water by delivering water into a dry column packed with the hydrophobic material. The formation of bubbles in the hydrophobic pores can be explained according to the Washburn equation:29

p=−

2γ cosθ r

(1)

where p is the induced capillary pressure, which extrudes a non-wetting liquid from a cylindrical -5-



pore of radius, r, or the pressure required to force the liquid into the pore, γ is the surface tension of the nonwetting liquid, and θ is the contact wetting angle in the liquid/surface/air system. The air trapped in the pores will be replaced by water vapor by equilibration of the column with degassed liquid water and the volume of the gas phase does not change if the pressure and temperature are kept constant. The incorporation of a gas phase onto the surface of the alkyl bonded silica in contact with water reduces the area of the water/alkyl chain interface. Therefore, one can control the area of the water/alkyl chain interface by pressure applied to the column because the volume of the gas phase can be changed by pressure according to eq 1. On the other hand, the amount of the alkyl chain layer remains constant independent of the volume of the gas phase and a solute molecule which can vaporize from the aqueous phase partitions into the alkyl chain layer through the gas phase. This means that SBMLC can meet the requirements described above and enable us to evaluate separately the contributions of the alkyl chain layer itself and the water/alkyl chain interface to the solute accumulation to the alkyl bonded silica material. We determined the bulk water-to-interface and the bulk water-to-C18 bonded layer distribution coefficients for various organic compounds by SBMLC assuming that the volume of the alkyl chain layer is constant regardless of the volume of liquid water in the pores and revealed that the hydrophobic compounds such as alkyl halides can penetrate into the C18 layer, whereas the compounds which have hydrophilic functional group are mostly accumulated at the interface between the bulk water and the C18 layer.28 -6-


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However, it is very likely from molecular perspective of the packing material that the molecules penetrating into the alkyl bonded layer may interact with the surface of silica substrate or end-capped silica. Therefore, the bulk water-to-C18 bonded layer distribution coefficients determined in the previous study may include the contributions not only of the alkyl chain layer but also of the end-capped silica surface to the solute retention. Although many researchers have pointed out that the surface of silica substrate significantly affects the distribution of a solute compound in RPLC,30-35 no one has quantitatively evaluated the contribution of the silica surface to the solute retention. In the present study, we determine the bulk water-to-interface and the bulk water-to-alkyl chain layer distribution coefficients for various compounds by SBMLC. The distribution coefficients are corrected for the contribution of the end-capped silica surface to the solute retention using end-capped C8 and C18 bonded silica columns. The change in the neat molecular surface area of the alkyl chains caused by its contact with liquid water has also been computed for accurate determination of the distribution coefficients. The unique ability of SBMLC to discriminate the contributions of the interface and the hydrophobic moiety to the solute retention will clearly give the picture of molecular distribution in the microheterogeneous structure of alkyl bonded silica in contact with water.

EXPERIMENTAL SECTION -7-



All chemicals used in this study were obtained from commercial sources and were of analytical reagent-grade unless otherwise stated. HPLC grade methanol and dichloromethane were obtained from Kanto Chemicals (Tokyo, Japan). Deuterium oxide (D2O) for NMR use was purchased from Wako Pure Chemicals (Tokyo, Japan). Water was purified subsequently with an Elix-Advantage 3-UV (Nihon Millipore, Tokyo, Japan) and an Arium 611 DI (Sartorius, Tokyo, Japan). The columns used were L-column2 ODS and L-column2 C8 (5 µm, 150 × 4.6 mm, Chemicals Evaluation and Research Institute, Tokyo, Japan). The schematic illustration of the surface structures of these packing materials is shown in Figure S1.36 A gas phase was incorporated into a column in a similar manner to the procedure described before.28 A packed column was filled with dichloromethane and then kept for 120 min at 80 °C in a liquid chromatograph oven (CTO-6A, Shimadzu, Japan) after the plugs of the both ends of the column had been removed. After drying the column completely, the column was set to the HPLC system and water was delivered into the column at a flow rate of 0.6 mL min-1 until the retention volume of D2O, VD2O , reached a constant value. The incorporation of the stationary gas phase into the column was ascertained by comparing VD2O with the total column void volume, V 0 ( VD2O < V0 ). The weight of each column packing material in the column was determined after the packing material was quantitatively transferred into a weighing bottle and then dried at 90 °C until a constant weight had been reached. Carbon and hydrogen contents of the packing materials were -8-


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determined by elemental analysis. Specific surface areas of the packing materials were measured by Seishin Enterprise (Tokyo, Japan) with a NOVA-4200e (Quantachrome Instruments, FL, USA) using the BET nitrogen sorption method. The molecular surface areas of the bonded alkyl chains of the packing materials used in this study were calculated using Chem3D (CambridgeSoft, MA, USA). The details are shown in the Supporting Information. SBMLC as well as RPLC was performed with a Shimadzu HPLC system equipped with a Model LC-30AD pump, a Model SIL-30AC auto sampler, a Model SPD-M20A photodiode array detector, and a Model RID-10A refractometric detector (Shimadzu, Tokyo, Japan). The columns were thermostated at 25 °C using a Shimadzu Model CTO-30A column oven. The injection volume was set at 10 µL. For the chromatographic measurements with a column filled with water (RPLC), the back pressure of the column was generated in order to suppress the formation of bubbles in the pores of the packing materials by attaching capillary PEEK tube of 65 µm I. D. and 150 cm long between the outlet of the column and the detector. All the eluents were filtered through a 0.45 µm membrane filter JHWPO 4700 obtained from Merck Millipore (Tokyo, Japan) before use. Elutions were carried out at a constant flow rate of ca. 0.6 mL min-1. The exact values of the volumetric flow rate were measured using a volumetric flask. The extra column volume was determined by measuring the elution volume of a sample solute through the system from which the column had been removed. Test solutions were prepared by dissolving analyte compounds in water. -9-



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Chromatographic data were collected and processed using a LC solution system (Shimadzu, Tokyo, Japan).

RESULTS AND DISCUSSION Estimation of the interfacial water volume, the gas phase volume, and the total molecular surface area of unwetted alkyl chains in an SBMLC column.

Shibukawa et al. have

shown that small inorganic ions and hydrophilic organic molecules differentiate the interfacial water formed on the hydrophobic surfaces from the bulk water and the retention of these solutes on a C18 silica column with pure aqueous systems can be interpreted with a partition between the bulk water and the interfacial water formed on the hydrophobic surface.37-39 The incorporation of the gas phase into the pores of the packing material brings about gas/water interface in addition to the water/alkyl chain interface. The interfacial water on the surface of the gas phase may act as another component of the hybrid separation medium. However, SBMLC with the column containing the stationary gas phase exhibited the same retention order for nonvolatile hydrophilic compounds such as ions and sugars as that observed in RPLC with the column filled with water. Since it can be assumed that these highly hydrophilic compounds do not partition into the alkyl chain layer, this result suggests that the gas/water interface has a similar solute accumulation selectivity to that of the water/alkyl chain interface or the total area of the gas/water interface is negligibly smaller than that of the water/alkyl chain interface. We thus assumed that the hybrid separation medium in SBMLC is -10-


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composed of the alkyl chain layer, the interfacial water at the surface of the alkyl layer, and the gas phase. We have shown that some compounds such as small inorganic ions distribute into the interfacial water, but not adsorb on the surface of alkyl chain layer. However, the partition into the interfacial water and the adsorption on the surface of alkyl chain layer cannot clearly be discriminated from each other for the compounds showing high affinity to the interface. In this study, the partition into the interfacial water and the adsorption onto the water/alkyl chain interface are thus evaluated inclusively in terms of the distribution into the interfacial water. This treatment is valid since the amount of a solute injected to the column is much smaller than the adsorption capacity of the alky chain layer surface. The structure of the hybrid separation medium in an SBMLC column packed with porous alkyl bonded silica incorporating the gas phase is schematically illustrated in Figure 1. It is required to estimate the amounts of the bulk water and each component of the hybrid separation medium in the column in order to determine the distribution coefficients of the solute compound between the bulk water and the individual components of the hybrid separation medium. The bulk water volume can be determined by measuring the retention volumes of two equally charged probe ions in two mobile phase electrolyte systems according to the following equation:37-40

VBW =

VAYXVBWZ − VAWZVBYX VAYX + VBWZ − VAWZ − VBYX

(2)

where V i jk is the retention volume of the probe ion i eluted with an aqueous solution of the -11-



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electrolyte jk. The probe ions and the eluent ions must fulfill the following requirements: (1) the association of a probe ion with counterion in solution can be neglected, (2) the ionic size is so small that the penetration of the ion into the pores is not restricted by size-exclusion or steric exclusion.40 In this study, the VBW values were determined by substituting the retention volumes of bromide, nitrate, iodide, and thiocyanate ions (A and B) eluted with 0.1 mol L-1 NaCl and NaClO4 (YX and WZ) aqueous solutions into eq 2. The VBW values determined for the L-column2 ODS and L-column2 C8 columns filled with water and the columns containing the fixed gas phase are tabulated in Table S1. The volume of the interfacial water, VIW , can be calculated by subtracting the VBW value from the total liquid water volume in the column, V L . When the column is filled with water, V L can be determined by measuring V D O and is identical to the column void volume, V0 . 2

(3)

VIW = VL − VBW = V0 − VBW

On the other hand, V L is not equal to V D O for an SBMLC column since D2O distributes not only 2

into the liquid water phase but also into the gas phase. We have thus estimated the VIW values for the SBMLC columns in the following manner. Since a nonvolatile and highly hydrophilic compound can be assumed to distribute only into the interfacial water from the bulk water, its retention volumes on the SBMLC column containing the gas phase, VR,S NV , and on the RPLC column filled with water, VR,RNV , are given by eq 4 and 5, respectively: S S V R,S NV = VBW + DIW,NVVIW

(4) -12-


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R R V R,RNV = VBW + DIW,NVVIW

(5)

where D IW, NV denotes the distribution coefficient of a nonvolatile hydrophilic compound defined as the ratio of its concentrations in the interfacial water and the bulk water ( = cIW,NV / cBW,NV ) and the superscripts S and R represent values for SBMLC and RPLC columns, respectively. From eq 5, the D IW, NV value of the compound is given by

DIW,NV =

cIW,NV c BW, NV

=

R VR,RNV − VBW R VIW

=

R VR,RNV − VBW R V0 − VBW

(6)

Hence, the volume of the interfacial water for an SBMLC column can be determined by the following equation:

S V IW =

S V R,S NV − V BW

(7)

D IW, NV

S The VIW values obtained from the retention volumes of thiourea, urea, uracil, glycerol, xylitol,

pentaerythritol, Br − , NO 3− , I − , and SCN - are listed in Table S2. The total liquid water volume S S for an SBMLC column, VLS , is given by the sum of V BW and VIW . The volume of the gas phase,

VG , can then be obtained by subtracting VLS from V0 . The values of V L , VBW , VIW , and VG for

the L-column2 ODS and L-column2 C8 columns are tabulated in Table 1. As shown in Table 1, the VIW value for the SBMLC column is less than 1 % of the value for the RPLC column due to the remarkable decrease in the area of the water/alkyl chain interface. The values of VBW as well as V L for the C18 and C8 silica columns in SBMLC mode are almost comparable, while the values for the columns filled with water in RPLC mode are quite different -13-



Page 14 of 45

from each other. These results indicate that liquid water is excluded from the pores of the packing materials and the internal space of the pores is nearly completely occupied by the gas phase in the SBMLC column. The fact that the VG value is larger for the C8 silica column than that for the C18 column demonstrates that the internal pore volume for the C8 silica particles is larger than that for the C18 silica, which can be understood on the basis of the thickness of the alkyl chain layer. As described above, the alkyl chain layer is so thin that the change in solute accumulation capacity of the alkyl chain layer due to the change in the area of the water/alkyl chain interface may not be neglected. Therefore we evaluated the capacity with the total molecular surface area of the unwetted alkyl chains instead of the volume of the alkyl chain layer. It has been reported that the alkyl bonded chains in contact with liquid water tend to bind or clump together.41-43 As a consequence, only outermost of alkyl chains comes into contact with liquid water, that is, the internal alkyl chains of bonded layer are unwetted. Therefore, the total molecular surface area of the unwetted alkyl chains, S C , can be given by the following equation. (8)

SC = ST − S W

where S T is the total van der Waals surface area of the bonded alkyl chains and S W is the surface area of the wetted alkyl chains. The values of S T for the C8 and C18 columns were calculated as follows:

ST =

Cwp N A S alkyl

(9)

mC nC 10 −20

where C and wp are the carbon content and weight of the packing material, NA is the Avogadro -14-


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constant, Salkyl and nC are the van der Waals surface area and carbon number of the bonded alkyl group (octyl or octadecyl), and mC is the atomic weight of carbon, respectively. The Salkyl values for octyl and octadecyl groups were calculated to be 429 Å2 and 197 Å2, respectively, using Chem 3D. We estimated the surface area of the wetted alkyl chains in the RPLC column filled with water, S WR , assuming that S WR is identical to the total surface area of the alkyl bonded silica particles determined by the BET method. Since the surface area of the wetted alkyl chains can be assumed to be proportional to the interfacial water volume, the values for the SBMLC columns, S WS , was calculated from the V IW values obtained for the RPLC and SBMLC columns as follows:

S WS =

S V IW S WR R V IW

(10)

The values of S T , S W , and S C determined for the L-column2 ODS and L-column2 C8 columns are tabulated in Table 2. It is interesting to note that the ratios of S TR / S WR for the C18 and C8 silica columns are 12.8 and 5.4 and agree well with the ratios of the surface areas of octadecyl and octyl groups to that of methyl group (34 Å2), 12.6 and 5.8, respectively. This may suggest that the alkyl chains bonded to the silica surface in the columns used in this study are densely packed and only the head methyl groups of the chains stick out from the bonded layer. The thickness of the interfacial water on the alkyl chain layer can be given by V IWR / S WR . The calculated values for the C18 and C8 silica materials are 1.3 and 1.2 nm, respectively. MD simulation studies have suggested the thickness of about 1 nm or less for the interface of hexane/water44 or the interfacial water layer at the surface of carbon tetrachloride.45Although the thickness of the -15-



Page 16 of 45

interfacial water estimated in this study is the distance from the surface of the alkyl chain layer at which small hydrophilic molecules and ions can sense the surface,38 the values obtained from these different studies are in fair agreement with each other. Evaluation of the contributions of the interfacial water, the alkyl chain layer, and the end-capped layer to the retention of organic compounds.

Now that the volume or area of every

component of the separation medium has been obtained, we can estimate the distribution coefficients of a solute for the individual parts of the hybrid stationary phase from its retention volumes on the RPLC column filled with water and the SBMLC column containing the stationary R gas phase. The retention volume of a solute compound in RPLC, VR(C18) , is expressed as

R R R R VR(C18) = VBW(C18) + DIW(C18)VIW(C18) + DAC(C18) S C(C18)

(11)

S On the other hand, the retention volume of the compound in SBMLC, VR(C18) , is given by

S S S S VR(C18) = VBW(C18) + DIW(C18)VIW(C18) + DAC(C18) S C(C18) + DGVG(C18)

(12)

In these two equations above, DIW(C18) , DAC(C18) , and DG are the solute distribution coefficients defined as follows:

DIW(C18) =

nIWVBW nBWVIW

(13)

DAC(C18) =

nACVBW nBW S C

(14)

DG =

nGVBW n BWVG

(15)

where nIW , nBW , n AC and nG are moles of the solute compound in the interfacial water at the surface of the C18 chain layer, the bulk water, the C18 chain layer, and the gas phase, respectively. -16-


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From eqs 11 and 12, we obtain

DIW(C18) =

R S R S ( aVR(C18) − VR(C18) ) − ( aVBW(C18) − VBW(C18) ) + DGVG(C18) R S aVIW(C18) − VIW(C18)

(16)

S R where a is the ratio of S C(C18) to S C(C18) . The D IW(C18) values of organic compounds of different

volatilities were thus estimated according to eq 16 assuming that DG can be given by

DG =

kH RT

(17)

where k H , R, and T are the Henry’s law constant with the dimension of pressure divided by molarity,46-48 the gas constant, and the column temperature, respectively. The D AC(C18) values were then determined by the following equation:

D AC(C18) =

S S S V R(C18) − V BW(C18) − D IW(C18) V IW(C18) − D G V G(C18) S S C(C18)

(18)

The DG , D IW(C18) , and D AC(C18) values obtained for various organic compounds are tabulated in Table 3. If the solute molecules interact only with the alkyl chains in the C18 bonded silica, D AC(C18) is expected to be related to liquid n-alkane/water partition coefficient, PLA , as follows:

DAC(C18) =

PLAVC ST

(19)

where V C is the volume of the alkyl chain layer. It has been shown that n-alkane/water partition coefficients are almost independent of the carbon number of n-alkanes (C6-C16) for various organic compounds.49 The D AC(C18) values were thus compared with the hexadecane/water partition coefficients, PC16 .49 Figure 2 shows the plots of log D AC(C18) against log PC16 . The inserted -17-



Page 18 of 45

straight line was drawn according to eq 19. The VC value for the C18 bonded layer was calculated by eq 20 assuming that the density of the C18 bonded layer, ρ C , is identical to that of liquid n-octadecane (0.7768 g mL-1 at 28 °C):50

VC =

wp − wsilica

(20)

ρC

where wsilica is the weight of the base silica in the column. We estimated the wsilica value from wp and the carbon content assuming that the origin of carbon of the packing material is only alkyl

group. According to eq 19, it is expected that all the data plots should fall on a straight line with the slope of unity and the intercept log VC / S T (the inserted line in Figure 2). However, the D AC(C18) values for the compounds having polar groups deviate from the line, indicating that polar compounds prefer the alkyl bonded silica to the bulk liquid n-alkane. As described above, the D AC(C18) may include the contributions not only of the alkyl chains but also of the end-capped silica

surface to the solute distribution into the alkyl bonded silica particles. If the end-capped silica surface and the bonded alkyl chain layer accumulate a solute compound independently of each other, R VR(C18) should be given by the following equation instead of eq 11:

R R R cor R V R(C18) = V BW(C18) + D IW(C18) V IW(C18) + D AC(C18) S C(C18) + D AE(C18) S E(C18)

(21)

where D AE is the adsorption equilibrium constant of the compound onto the end-capped silica cor surface, S E is the area of the end-capped silica surface, and DAC(C18) is the corrected D AC(C18) for

the adsorption to the end-capped surface. Similarly, the retention volume of the compound on a C8 -18-


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R , is expressed as silica column filled with water, VR(C8)

R R R cor R V R(C8) = V BW(C8) + D IW(C8) V IW(C8) + D AC(C8) S C(C8) + D AE(C8) S E(C8)

(22)

cor Since D AC indicates the magnitude of adsorption of a solute compound per unit area of

cor cor hydrocarbon, DAC(C8) can be assumed to be identical to DAC(C18) . In addition, the C18 and C8

bonded silicas used in this study, L-column2 ODS and L-column2 C8, are manufactured as fully end-capped materials by the same end-capping method. It is thus reasonable to assume that D AE(C8) cor is equal to D AE(C18) . From these assumptions, the D AC values can be calculated by the following

equation

D

cor AC

=

R R R R R R (VR(C18) − bVR(C8) ) − (VBW(C18) − bVBW(C8) ) − ( DIW(C18)VIW(C18) − bDIW(C8)VIW(C8) ) R R S C(C18) − bS C(C8)

(23)

where b is the ratio of S E(C18) to S E(C8) . The b value was calculated to be 1.02 assuming that S E is proportional to the total surface area of the bare silica particles in the column. The D IW(C8) value as well as the values for the volumes of the gas and liquid phases in the C8 silica column were determined in a similar manner to the calculation of the values for the C18 silica column. cor The validity of the D AC values was confirmed by comparison with the PC16 values. Figure 3

cor shows the plot of log D AC against log PC16 . Not only the plots for hydrophobic compounds but

also those for hydrophilic compounds having polar groups yield a straight line with the slope of approximately unity and the intercept of log VC / S T , which reveals that the alkyl chain layer behaves very similarly to liquid n-alkane as a separation medium. FT-IR studies on alkyl bonded silica gels as well as pure n-alkanes demonstrated that the conformational state of the alkyl chains -19-



depends on the chain length and specific position in the alkyl chain.14,

Page 20 of 45

15

On the contrary,

n-alkane/water partition coefficients depend little on the chain length of n-alkane as described above.49 This suggests that the chain conformation does not strongly affect the molecular interaction of the alkyl chains with a solute molecule, although the distribution of large molecules with rigid and well defined structures such as polycyclic aromatic hydrocarbons may be affected by the morphology of the alkyl chain layer. 13, 51 The results shown above indicate that one can evaluate quantitatively the respective contributions of the water/alkyl chain interface, the alkyl chain layer, and the end-capped silica surface to the retention of a solute compound in an aqueous solution by SBMLC. Figure 4 shows the relative contributions of the bulk water, the water/alkyl chain interface, the grafted alkyl chain layer, and the end-capped silica surface to the retention volumes of the organic compounds on an R R cor R L-column2 ODS column, which are represented by VBW(C18) , DIW(C18)VIW(C18) , DAC(C18) S C(C18) , and

D AE(C18) S E(C18) , respectively. The relative solute retentivities of the respective phases and interfaces

in an L-column2 C8 column are shown in Figure S2 for comparison. As seen in Figure 4 and Figure S2, the water/alkyl chain interface predominantly retains all the organic compounds studied, whereas the alkyl chain layer does not act as an effective retention medium for hydrophilic compounds although it has large retention capacity for highly hydrophobic compounds. It is worth noting that the end-capped silica surface as well as the water/alkyl chain interface exhibits quite different retention selectivity from the alkyl chain layer. Furthermore, alkyl halides and diethyl -20-


Page 21 of 45 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


ether substantially reside on the end-capped silica surface. The retention on the end-capped surface may in substantial part originate from the interaction between a solute molecule and the siloxane surface.

Effect of the bonded alkyl chain length on the separation selectivity in SBMLC and RPLC with alkyl bonded silica column.

A chromatographic separation of some organic compounds on

the L-column2 ODS was compared with that on L-column2 C8. In an SBMLC column containing the stationary gas phase, the alkyl chain layer and the end-capped surface predominantly act as the solute retention media because the water/alkyl chain interface is negligibly small and the capacity of the gas phase for retaining solute compounds is rather small compared to the other parts of the hybrid medium (see Tables 1 and 3). Therefore, when the C8 and C18 columns fully contain the stationary gas phase, the retention volumes of a solute compound on the C8 and C18 columns are represented by the following approximate equations: S S cor R VR(C8) ≈ VBW(C8) + DAC S C(C8) + DAE S E(C8)

(24)

S S cor R VR(C18) ≈ VBW(C18) + DAC S C(C18) + DAE S E(C18)

(25)

As shown above, the S E values for the tested C8 and C18 columns are approximately equal to each other ( S E(C18) / S E(C8) = 1.02). Consequently, it is expected that both of the columns exhibit similar retention selectivity in SBMLC mode. Figure 5 shows a chromatographic separation of some organic compounds on the L-column2 ODS and L-column2 C8 in SBMLC mode. As expected, the retention orders observed for the C8 and C18 silica columns are identical to each other. Every -21-



compound shows larger retention volume on the C18 column than on the C8 column, which can be understood on the basis of the fact that the C18 layer has larger S T value than the C8 layer. On an RPLC column filled with water, on the contrary, the water/alkyl chain interface plays a decisive role in the solute retention. As shown in Figure 6, the solute retention orders of the compounds on the L-column2 ODS and L-column2 C8 columns in RPLC mode are quite different from each other. The plots of log D IW(C8) against log D IW(C18) are shown in Figure 7. All the plots give a straight line with the slope of unity going through the origin, which indicates that D IW(C8) is equal to D IW(C18) or the water/C8 layer interface has the same retention characteristics as that for the water/C18 chain layer interface. The retention volumes of a solute compound on the C18 and C8 cor columns are given by eqs 21 and 22, respectively. Since DIW , DAC(C18) , D AE(C18) , and S E(C18)

values can be assumed to be independent of the length of the alkyl chain, the difference in separation selectivity between the two columns in RPLC mode is attributed to the differences in R and S CR . As shown in Table 3, Figure 4 and Figure S2, most of the organic compounds VIW

examined in this study are predominantly accumulated at the water/alkyl chain interface. On the other hand, the n-alkyl chains do not act as an effective retention medium for hydrophilic compounds but has large retention capacity for hydrophobic compounds. S CR increases as the bonded alkyl chain length increases, while VIW decreases as shown in Tables 1 and 2. Therefore, it is expected that the relative retention of hydrophobic compounds is larger on the C18 column than on the C8 column, while the hydrophilic compounds exhibit larger retention volumes on the C8 -22-


Page 22 of 45

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column than on the C18 column. The experimental results shown in Figure 6 are in accordance with the prediction. These results indicate that the separation selectivity in reversed-phase systems is determined by the relative sizes of the water/hydrophobe interface and the hydrophobe itself.

CONCLUSIONS We have elucidated the solute separation mechanism in the reversed-phase system with alkyl bonded silica by measuring the distribution coefficients of various organic compounds with a wide range of polarity or hydrophobicity between the bulk water and each component of the hybrid stationary phase, i.e., the water/alkyl chain interface, the alkyl chain layer, and the end-capped silica surface by SBMLC. The results obtained in this study reveal that a solute compound in an aqueous solution distributes into the alkyl chain layer bonded to silica surface similarly to the partition into a liquid n-alkane and the separation selectivity exerted by the adsorption onto the water/alkyl chain layer interface is quite different from that of the partition into the alkyl chain layer. It has also been clarified that the end-capped silica surface exhibits different separation selectivity from the alkyl chain layer presumably due to the interaction between a solute compound and siloxane bonding surface. Over the past decades, substantial progress has been made in the molecular description of aqueous/hydrophobe interfaces by surface sensitive spectroscopy, e.g., vibrational sum frequency generation spectroscopy or X-ray photoelectron spectroscopy.52 However, the determination of the spatial distribution of the compounds in the vicinity of the interface between aqueous solutions and -23-



hydrophobe is very challenging and new experimental methods are still required for understanding of the interfaces. Hydrophobic surfaces play a critical role in mass transfer processes in aqueous media such as the distribution of oils, drugs and toxins in the environmental and living systems. Adsorption of molecules and ions in an aqueous solution to a hydrophobic surface is thus a topic relevant to many fields. SBMLC can be one of the useful methods for elucidation of distribution behavior of a solute molecule at the aqueous/hydrophobe interfaces.

Glossary of Symbols The symbols used in this paper are listed below. Superscripts S and R in the volumes and surface areas denote SBMLC and RPLC, respectively.

V R : retention volume of a solute compound V0 : column void volume

V L : total liquid water volume VBW : bulk water volume VIW : interfacial water volume VG : volume of gas phase V C : volume of the alkyl chain layer

S T : total van der Waals surface area of the bonded alkyl chains S W : total surface area of the wetted alkyl chains -24-


Page 24 of 45

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S C : total van der Waals surface area of the unwetted alkyl chains

S E : total area of the end-capped silica surface DIW : bulk water-to-interface distribution coefficient DG : bulk water-to-gas phase distribution coefficient cor : adsorption equilibrium constant of a solute compound onto the alkyl chains D AC

DAE : adsorption equilibrium constant of a solute compound onto the end-capped silica surface

Supporting Information Calculation of the molecular surface areas; Bulk water volumes calculated for L-column2 ODS and L-column2 C8 columns filled with liquid water and the column containing the fixed gas phase (Table S1); Interfacial liquid water volumes determined for L-column2 ODS and L-column2 C8 columns containing the fixed gas phase (Table S2); Schematic illustration of the surface structures of L-column2 ODS and L-column2 C8 (Figure S1); Contributions of the bulk water, the interfacial water, the grafted alkyl chains, and the end-capped silica surface to the solute retention on a C8 silica column (Figure S2), and. This material is available free of charge via the Internet at http://pubs.acs.org.

ACKNOWLEDGMENTS This research was supported by a Grant-in-Aid for Scientific Research No. 16H04161 and No. -25-



25288062 from Ministry of Education, Culture, Sports, Science and Technology, Japan.

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Page 34 of 45

Table 1 The volumes (mL) of total liquid water (VL), bulk water (VBW), interfacial water (VIW), and fixed gas phase (VG) determined for an L-column2 ODS column (C18) and an L-column2 C8 (C8) column filled with liquid water (RPLC) and the columns containing the fixed gas phase (SBMLC) at 25 °C.

VL

VBW

VIW

VG

RPLC

1.648 ± 0.002

1.35 ± 0.03

0.30 ± 0.03

0

SBMLC

0.977 ± 0.001

0.975 ± 0.001

0.0022 ± 0.0007

0.671 ± 0.002

RPLC

1.828 ± 0.001

1.48 ± 0.03

0.35 ± 0.03

0

SBMLC

0.993 ± 0.001

0.990 ± 0.001

0.0030 ± 0.0006

0.835 ± 0.001

column

C18

C8

Table 2 The values of S T , S W , and S C (m2) determined for an L-column2 ODS column and an L-column2 C8 column filled with liquid water (RPLC) and the columns containing the fixed gas phase (SBMLC).

ST

SW

SC

RPLC

2916

234

2682

SBMLC

2916

2

2914

RPLC

1510

292

1218

SBMLC

1510

3

1507

column

C18

C8

-34-


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Table 3 The distribution coefficients of organic compounds for the bulk water-to-the gas phase (DG), the bulk water-to-the interfacial water (DIW(C18)), and the bulk water-to-the alkyl chain layer (DAC(C18)), obtained for an L-column2 ODS column at 25 °C compound

DG

DIW(C18)

D AC(C18) / nm

methanol

0.000186

2.8 ± 0.3

0.0077 ± 0.0008

ethanol

0.000215

8.2 ± 0.7

0.012 ± 0.002

1-propanol

0.000315

31 ± 9

0.033 ± 0.008

1-butanol

0.000315

129 ± 12

0.10 ± 0.03

0.00000454

333 ± 30

0.10 ± 0.08

acetonitrile

0.000835

6.6 ± 0.6

0.106 ± 0.002

propionitrile

0.00151

25 ± 2

0.265 ± 0.006

acetone

0.00157

22 ± 2

0.105 ± 0.005

2-butanone

0.00227

85 ± 8

0.30 ± 0.02

dichloromethane

0.117

37 ± 3

3.47 ± 0.01

1,2-dichloroethane

0.0560

98 ± 9

5.17 ± 0.02

chloroform

0.170

132 ± 12

11.50 ± 0.03

bromoethane

0.315

92 ± 8

9.89 ± 0.03

nitromethane

0.000909

6.6 ± 0.6

0.147 ± 0.002

nitroethane

0.00195

25 ± 2

0.426 ± 0.006

1-nitropropane

0.00341

101 ± 9

1.15 ± 0.02

diethyl ether

0.0518

210 ± 19

1.54 ± 0.05

benzene

0.227

340 ± 30

29.1 ± 0.1

benzyl alcohol

methyl acetate

0.00524

45 ± 4

0.26 ± 0.01

isopropyl acetate

0.0141

656 ± 59

2.0 ± 0.2

-35-



Page 36 of 45

Figure Captions Figure 1. Schematic illustration of the hybrid separation medium in an SBMLC column packed with a porous alkyl bonded silica. See the Glossary for other details.

Figure 2. Values of log D AC(C18) plotted against log PC16 .

cor Figure 3. Values of log DAC(C18) plotted against log PC16 .

R

R Figure 4. Contributions of the bulk water ( VBW(C18) ), the interfacial water ( DIW(C18)VIW(C18) ), the

cor R grafted alkyl chains ( DAC(C18) S C(C18) ), and the end-capped silica surface ( D AE(C18) S E(C18) ) to the

solute retention on a C18 silica column filled with water. Column: L-column2 ODS. Temperature: 25 °C.

Figure 5. Chromatograms of organic compounds obtained by SBMLC. Columns: L-column2 ODS and L-column2 C8. Temperature: 25 °C. Mobile phase: pure water. Flow rate: 0.6 mL min-1. Detector: deferential refractive index detector. Compounds: A, propionitrile; B, benzyl alcohol; C, diethyl ether; D, isopropyl acetate; E, bromoethane; F, chloroform; G, benzene.

Figure 6. Chromatograms of organic compounds obtained by RPLC. See Figure 5 for other details. -36-


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Figure 7. Values of log D IW(C8) plotted against log D IW(C18) . Temperature: 25 °C.

-37-



Figure 1

-38-


Page 38 of 45

Page 39 of 45

-1

n-alcohols nitriles ketones nitro alkanes alkyl halides esters diethyl ether benzyl alcohol benzene

-2

-3

log DAC(C18)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


-4

-5

-6

-7 -3

-2

-1

0

1

2

3

log PC16

Figure 2

-39-



-1


-2

Cor

log D AC(C18)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

-3

-4

-5

-6

-7 -3

-2

-1

0

1

2

3

log PC16

Figure 3

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Page 40 of 45

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VBW

DIW(C18) VIW(C18)

cor

R

D AC(C18) S C(C18)

DAE(C18) S E(C18)

bromoethane chloroform 1,2-dichloroethane dichloromethane methyl acetate diethyl ether 1-nitropropane nitroethane nitromethane 2-butanone acetone propionitrile acetonitrile 1-butanol 1-propanol ethanol methanol 0

15

30

45

60

VR/ mL

Figure 4

-41-


75


A, B

CD

F E

G C8 bonded silica

CD

A, B E F

0.2 µRIU

G C18 bonded silica

0

40

80

120

160

retention time/min

Figure 5

-42-


Page 42 of 45

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A

B E F

G C8 bonded silica

C

D B A

E F C

C18 bonded silica G

0.2 µRIU

D

0

60

120

180

240

300

360

420

retention time/min

Figure 6

-43-



3


2

log DIW(C8)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

1

0 0

1

2

3

log DIW(C18)

Figure 7

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Page 44 of 45

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TOC Graphic

bulk water interfacial water

alkyl chain end-capped silica surface silica support

-45-


Hydrophobe Interface versus Partition into the

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