Anal. Chem. 1997, 69, 4079-4081
Correspondence
Chromatography on Water-Ice Purnendu K. Dasgupta* and Youwen Mo
Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, Texas 70409-1061
Ice contains a thin film of water on its surface, the thickness ranging from nanometer to micrometer levels, depending on the bulk temperature and the composition of the solution used in making the ice. It follows that water held on the surface of an ice particle should endow a column packed with microparticulate ice the properties of a chromatographic column containing a pellicular packing. We demonstrate here the normal phase separation of two dyes on a column packed with millimeter-size ice particles, held at -18 °C. Water, the most abundant molecular form of matter on the surface of the earth, is omnipresent on this planet. Terrestrial life is dependent on and controlled by the physics and chemistry of water. The planetary climatology is largely associated with the phase changes of water, as modulated by the energy input of the sun. Volumes have been written on water. Of particular interest in the present context are the works of Frank,1 Eisenberg and Kauzmann,2 Hobbs,3 and Van der Leeden et al.4 After many decades of intense efforts beginning with the original work of Ro¨ntgen5 and continuing through those of Pauling,6 a structural model of liquid water that is able to quantitatively explain all of the physical properties of this enigmatic fluid still remains elusive.7 Compared to water, the structure of ice is much better understood.2 This is particularly impressive in view of the fact that water-ice exists in at least nine distinctly different crystalline forms, depending on pressure and temperature.3 Indeed, recent studies persuasively argue that the structure of liquid water and many of its properties are best understood as an admixture of two different crystalline forms of ice.8 Ice formed at ambient pressure by condensation on a cold surface at temperatures above about -80 °C is of the routinely encountered variety called ice-1h. It has a highly open hexagonal structure, resulting in a very low density (0.9167 at 0 °C). However, when contained water (e.g., in a beaker, a tube, or a capillary) is frozen, the solid mass formed is polycrystalline with (1) Frank, F. Ed. Water. A Comprehensive Treatise; Plenum: New York, 1973; Vols. 1-4. (2) Eisenberg, D.; Kauzmann, W. The Structure and Properties of Water; Oxford University Press: New York, 1969. (3) Hobbs, P. V. Ice Physics; Clarendon Press: Oxford, UK, 1974. (4) Van der Leeden, F.; Troise, F. L.; Todd, D. K. The Water Encyclopedia, 2nd ed.; Lewis Publishers: Chelsea, MI, 1990. (5) Ro ¨ntgen, W. C. Ann. Phys. 1892, 45, 91. (6) Pauling, L. Phys. Rev. 1930, 36, 430. (7) Bassez, M.-P.; Lee, J.; Robinson, G. W. J. Phys. Chem. 1987, 91, 5818. (8) Robinson, G. W., Zhu, S.-B., Singh, S., Evans, M. W. Water in Biology, Chemistry and Physics; World Scientific Publishing: London, 1996. S0003-2700(97)00423-X CCC: $14.00
© 1997 American Chemical Society
grains of variable size.3 As a chromatographic separation medium, this type of relatively impermeable solid mass may be of limited utility. Although the current practice of high-performance liquid chromatography (HPLC) is dominated by the bonded, reversed phase mode, the birth of modern liquid chromatography can be traced to the seminal paper by Martin and Synge,9 in which the operative mode was liquid-liquid partition chromatography. Indeed, one of the most widely used initial modes in HPLC was normal phase LC in which eluents of modest polarity were used in conjunction with silica packings, and as in the Martin-Synge experiment, water adsorbed on silica acted as the stationary phase. The advantages and disadvantages of this type of chromatography have been discussed in reference texts;10a for preparative separations, this mode still has some attraction. One important reason for the fall of such methods from grace has been the difficulty of maintaining a constant stationary phase loading. In the context of ice, it was Michael Faraday who first proposed that the surface of ice is covered by a layer of liquid water (see, e.g., ref 11 for a discussion and history). It has since been established that the liquid layer thickness ranges from 3-30 nm at -60 °C to 500-3000 nm at -1 °C, the lower values corresponding to ice made from distilled water and the higher values to ice made from 1 mM NaCl.12 In this regard, an ice particle represents a potentially useful chromatographic stationary phase on which it should be possible to carry out normal phase liquid chromatography. For such a packing, the active sorbent layer is on the surface of the particle, much like the commonly known superficially porous or pellicular packing.13 In the present report, we show the feasibility of using ice particles as a stationary phase for normal phase chromatographic separations. EXPERIMENTAL SECTION Preparation of Column Packing and Column. Techniques for making small (e100 µm) ice particles have been described in the literature;14 essentially, nebulized droplets are frozen in a suitably cold fluid medium much like individual raindrops are collected in liquid nitrogen for subsequent analysis.15 Ice particles made this way have been packed into columns to simulate snow (9) Martin, A. J. P.; Synge, R. L. M. Biochem. J. 1941, 35, 1358. (10) Snyder, L. R.; Kirkland, J. J. Introduction to Modern Liquid Chromatography, 2nd ed.; Wiley: New York, 1979. (a) pp 16-172; (b) p 36. (11) Dash, J. G. Contemp. Phys. 1989, 30, 89. (12) Conklin, M. H.; Bales, R. C. J. Geophys. Res. 1993, 98, 16851. (13) Horvath, Cs., Preiss, B.; Lipsky, S. Anal. Chem. 1967, 39, 1422. (14) Sommerfeld, R. A.; Freeman, T. L. Making Artifical Snow for Laboratory Use. USDA For. Serv. Res. Note RM-486, 1988. (15) Baechmann, K.; Haag, I.; Roeder., A. Atmos. Environ. 1993, 27A, 1951.
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beds and used for studying reactivities of ice toward various gases.16,17 However, a simpler approach appeared sufficient for this preliminary study. Crushed ice in a beaker from the in-house vending machines was cooled to dry ice temperatures by putting it on a block of dry ice in a Styrofoam cooler. A mortar and pestle was precooled in the same manner. After a precooling period of 30 min or more, approximately equal portions of the crushed ice and dry ice were taken in the mortar and manually milled as quickly as possible. More than one batch was processed if necessary to have an amount enough to pack the column. A 25 mL capacity buret of 1.2 cm i.d. was used as the column, using a bed of glass wool at the bottom as the bed support. The entire length of the buret, above the stopcock, was surrounded by a Plexiglas jacket tube, with inlet/outlet connections at the top/bottom through which 60% aqueous ethanol was circulated at -18 °C by a recirculating cooler (Model 2095, Forma Scientific, Marietta, OH). Solvents such as ethyl acetate, hexane, and acetone were precooled to at least -20 °C by putting these on dry ice in the cooler. Cold ethanol was circulated in the empty column jacket for at least 20 min. The column was then washed twice with precooled aliquots of 50:50 ethyl acetate/hexane. The powdered ice was loaded into the column and repeatedly washed with the above precooled solvent to remove entrained air bubbles. For the results reported here, the column bed height was ∼20 cm. Individual ice particles were taken with tweezers, put on a tared filter paper, and weighed to determine size. The weight of individual particles ranged from 0.1 to 0.4 mg in weight, equivalent spherical diameter 0.74 ( 0.17 mm. Visual microscopic examination also indicated that the particles are irregular in structure (most resembling a cube or a deformed version thereof) with significant variation in size. Chromatography. For the example discussed here, methyl red [2-[4-(dimethylamino)phenyl]azo]benzoic acid, MW 269.3)and methyl violet 2B (MW ∼403) were used as the test solutes. Saturated solutions were prepared in ethyl acetate and stored in a freezer. Concentrated solutions were deliberately used to test column loadability. Three to four drops of the test solution was applied from a 1 mL syringe (∼200-300 µL) to the top of the column bed, distributing it as evenly as possible. Elution was begun with precooled, water-saturated 50:50 ethyl acetate/hexane, slowly increasing the fraction of ethyl acetate until pure ethyl acetate is used. Acetone, in 5-10% amount, was added to the ethyl acetate in the final eluent aliquot to elute some minor components present in the samples that were strongly retained by the column. Optical detection of the eluate was accomplished with a Kratos Model 757 absorbance detector (Applied Biosystems). RESULTS AND DISCUSSION More than two decades ago, Hartkopf and Karger18 suggested that the uptake of various gases on ice can essentially be modeled from data obtained with gas chromatography using adsorbed water as the stationary phase, based on the fact that there is a liquid layer on ice. To our knowledge, actual chromatographic separations on ice have never been reported. (16) Conklin, M. H.,; Sigg, A.; Neftel, A.; Bales, R. C. J. Geophys. Res. 1993, 98, 18376. (17) Conklin, M. H.; Sommerfeld, R. A.; Laird, K. and Villinski, J. E. Atmos. Environ. 1993, 27A, 159. (18) Hartkopf, A.; Karger, B. L. Acc. Chem. Res. 1973, 6, 209.
4080 Analytical Chemistry, Vol. 69, No. 19, October 1, 1997
Figure 1. Separation of methyl red and methyl violet on a column of ice.
Figure 1 shows the chromatograms obtained at different detector wavelengths that allow the monitoring of the two solutes individually or together. Methyl red and methyl violet are both highly water-soluble dyes, the latter having the stronger affinity for water. The order of elution on the ice stationary phase is the same as that if the experiment is conducted instead using a stationary phase of silica gel containing adsorbed water, except that a more polar eluent composition is necessary to elute the dyes in a reasonable period of time. Although the present experiments are crude by any standard, they are sufficient to show the fundamental feasibility of carrying out separations on ice as a stationary phase. An obvious advantage of separation experiments is that separated dye bands can in fact be readily visually observed and the level of sophistication is such that the experiment can be conducted even in high school
laboratories for pedagogical purposes, complete with inexpensive light emitting diode-based optical detectors, if one so desired. However, even the limited amount of data presented here is worthy of some quantitative examination. The plate numbers for the two analyte peaks in Figure 1 fall between 225 and 375 plates; in other, repeated experiments, 200-400 plates have been observed for the same experiment. This equates to a plate height h of 0.05-0.1 cm for the 20 cm column, containing essentially pellicular packing of ∼0.075 cm diameter (dp) or a reduced plate height of ∼1, for a linear velocity, u, of ∼0.015 cm/s. As a comparison, Snyder and Kirkland10a suggested that, at a linear velocity of 0.1 cm/s, the plate height for 40 µm pellicular particles (presumably at room temperature, using aqueous eluents) is of the order of 0.01 cm. For pellicular packing, the important determinants of column efficiency are eddy diffusion and resistance to mass transfer in the mobile phase. The contribution of the first term to the plate height is proportional to the particle diameter while that of the second term is proportional to dp2u/ Dm, where Dm is the diffusion coefficient of the analyte in the mobile phase. Based on the first term alone, the plate height in the present case is expected to be a factor of ∼20 larger, due to the difference in diameter. Regarding the second term, ethyl acetate is a solvent with a viscosity about half that of water at room temperature. But we are operating at nearly -20 °C with fairly large analyte molecules. As a first approximation, we assume therefore that there is no significant change in Dm relative to the comparison benchmark. Based on dp2u, the second term is therefore likely to result in a value of h ∼50 times larger. It is fair to conclude that the separation performance is at least as good as can be expected on the basis of purely theoretical considerations. Obviously, smaller particles will be necessary to produce more efficient columns. Freezing of nebulized microdroplets is hardly a daunting task; in the right season and locale this would involve little more than nebulizing water in suitably small drops outdoor and collecting the ice particles, in much the same way artificial snow is made in large quantities in ski areas when nature is less cooperative. In the near future, we intend to generate smaller particles with a cooled grinder and slurry pack such small particles. Obviously, for any given particle dimension, pressure drop will increase as particle size decreases. It is of course wellknown that the melting point of ice decreases with increasing pressure, and the maximum permissible operating temperature must therefore decrease with decreasing particle size. The pressure drop for a 25 cm long column with a linear velocity of 0.1 cm/s for a liquid viscosity of 1 cP is plotted in Figure 2, as a function of particle size, according to10b
Figure 2. Operating pressure as a function of particle size and melting point at that pressure.
where ∆P is the pressure drop, η is the solvent viscosity, L is the length of the packing, and φ is assumed to have the worst case value of 1000, typical of irregular porous packing. It is clear that operating with essentially any particle size of ice attainable in the near future will not require impossibly low operating temperatures. Using ice as a stationary phase can potentially have many advantages, especially in preparative separations where extremely high efficiencies are not necessary while high loadability and a low cost of the stationary phase are desirable. When ingredients are incorporated into the ice surface by freezing an appropriate solution, it is noteworthy that such material should be readily recoverable. We feel that a particularly promising application in this regard is the separation of chiral isomers (for example, largescale separation of pharmaceuticals/biochemicals) using ice impregnated with chiral agents such as cyclodextrins. ACKNOWLEDGMENT This research was supported by the U.S. Department of Energy, Ofiice of Basic Energy Sciences, Division of Chemical Sciences through Grant DE-FG03-95ER14496. However, the manuscript has not been subject to review by the DOE and no endorsements should be inferred. Received for review April 23, 1997. Accepted July 18, 1997.X AC970423S
∆P ) φηL/dp2
(1) X
Abstract published in Advance ACS Abstracts, September 1, 1997.
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