Patent Application
20060113231
The present invention relates to methods of analytical sample preparation for instrumental analysis. More specifically, the present invention relates to capillary microextraction techniques for pre-concentrating trace analytes and microextraction devices.
Sample preparation is an important step in chemical analysis, especially when dealing with traces of target analytes dispersed in complex matrices. Such matrices are commonplace in samples from environmental, petrochemical, and biological origins. Samples of this nature are not generally suitable for direct introduction into analytical instruments. Such incompatibility is related to two factors. First, the complex matrices may have a detrimental effect on the performance of the analytical system or they may interfere with the analysis of the target analytes. Second, the concentration of the target analytes in the sample may be so low that it goes beyond the detection limit of the analytical instrument. In both cases sample preparation is necessary to make the sample compatible with analytical instrumentation. This is achieved through sample clean-up and sample pre-concentration. Sample derivatization is also sometimes necessary to facilitate analysis and detection of target compounds.
Sample preparation in chemical analysis often involves various extraction techniques to isolate and pre-concentrate target compounds from complex matrices in which they exist in trace concentrations. Conventional extraction techniques (e.g., liquid-liquid extraction (Majors, R. E. LC*GC. Int. 1997, 10, 93-101), Soxhlet extraction (Lopez-Avila, V.; Bauer, K.; Milanes, J.; Beckert W. F. J. AOAC Int. 1993, 76, 864-880), etc.) frequently used for this purpose are often time-consuming and involve the use of large volumes of hazardous organic solvents.
To address environmental and health concerns associated with the use of large volumes of organic solvents and to reduce sample preparation time, newer extraction techniques have been developed that use either reduced amounts of organic solutes such as solid-phase extraction (SPE), (Coulibaly, K; Jeon L. J. Food Rev. Int. 1996, 12, 131-151), accelerated solvent extraction (ASE) (Richter, B. E.; Jones, B. A.; Ezzel, J. L.; Porter N. L.; Abdalovic N.; Pohi, C. Anal. Chem. 1996, 1033-1039), microwave-assisted solvent extraction (MASE) (Zlotorzynski, A. Crit. Rev. Anal. Chem. 1995, 25, 43-76), etc. Another approach to address these problems was to develop sample preparation techniques using alternative, less hazardous extraction media, such as supercritical fluid extraction (SFE) (Hawthorne, S. B.; Anal. Chem. 1990, 62, 633A-642A). However, the extraction technique which is most fascinating from the environmental and occupational health and safety points of view is solid-phase microextraction (SPME) developed by Pawliszyn and coworkers (Berladi, R. P.; Pawliszyn, J. Water Pollut. Res. J. Can. 1989, 24, 179-91; Arthur, C. L.; Pawliszyn, J. Anal. Chem. 1990, 62, 2145). SPME completely eliminates the use of organic solvents for the extraction of analytes from a wide range of matrices. Another important feature of SPME is that, unlike conventional extraction techniques, it does not require exhaustive extraction-establishment of equilibria between the sample matrix and the stationary phase coating is sufficient to obtain quantitative extraction data. For most samples, the equilibration time is less than 30 minutes, which places SPME among the fastest extraction techniques.
In SPME, the outer surface of a solid fused silica fiber (approximately 1 cm at one end) is coated with a selective stationary phase. Thermally stable polymeric materials that allow fast solute diffusion are commonly used as stationary phases. The extraction operation is carried out by simply dipping the coated fiber into the sample matrix and allowing time for the partition equilibrium to be established. The sensitivity of the method is mostly governed by the partition coefficient of an analyte between the coating and the matrix. Extraction selectivity can be achieved by using appropriate types of stationary phases that exhibit high affinity toward the target analytes.
In its traditional format, SPME has a number of drawbacks. First, since the stationary phase coating is applied to the outer surface of the fiber, it is more vulnerable to mechanical damage. Second, traditional methods for the preparation of coatings fail to provide adequate thermal and solvent stability to the thick stationary phase (several tens of micrometers in thickness) coatings that are needed in SPME. This is due to the lack of chemical bonding between the coatings and the substrate to which they are applied.
In recent years, the extraction of analytes by GC stationary phase coatings on the capillary inner surface has received considerable attention. The introduction of in-tube SPME had the primary purpose of coupling SPME to high-performance liquid chromatography (HPLC) for automated applications. The in-tube SPME method uses a flow-through process where a coated capillary is employed for the direct extraction of the analytes from the aqueous sample. The extraction process involves agitation by sample flow in and out of the extraction capillary. Successful coupling of in-tube SPME with HPLC, as well as HPLC-MS, has been achieved for the specification of organoarsenic compounds, (Wu, J.; Mester, Z.; Pawliszyn, J. Anal. Chem. 1999, 71, 4237-4244) and determination of rantidine, (Kataoka, H.; Lord, H. L.; Pawliszyn, J. J. Chromatogr. B 1999, 731, 353-359) β-blockers, (Kataoka, H.; Narimatsu, S.; Lord, H.; Pawliszyn, J. Anal. Chem. 1999, 71, 4237-4244) carbamate pesticides, (Gou, Y.; Pawliszyn, J. Anal. Chem. 2000, 72 2774-2779; Gou, Y.; Eisert, R.; Pawliszyn, J. J. Chromatogr. A 2000, 873, 137-147; Gou, Y.; Tragas, C.; Lord, H.; Pawliszyn, J. J. Micro September 2000, 12, 125-134) and aromatic compounds (Wu, J.; Pawliszyn, J. J. Chromatogr. A 2001, 909, 37-52).
In spite of rapid on-going developments, especially in the areas of in-tube SPME applications, a number of fundamental problems remain to be solved. First, GC capillaries that are used for in-tube SPME typically have thin coatings that significantly limit the sample capacity (and hence sensitivity) of the technique. Conventional static coating techniques (Bouche, J.; Verzele, M. J. Gas Chromatogr. 1968, 6, 501-505; Janak, K.; Kahle, V.; Tesarik, K.; Horka, M. J. High Resolut. Chromatogr./Chromatogr. Commun. 1985, 8, 843-847; Sumpter, S R.; Woolley, C. L.; Hunag, E. C.; Markides, K. E.; Lee, M. L. J. Chromatogr. 1990, 517, 503-519) used to prepare stationary phase coatings in GC columns are designed primarily for creating thin (sub-micrometer thickness) coatings. Thus, developing an alternative technique to provide higher coating thickness suitable for in-tube SPME applications is very important.
Second, usually the stationary phase coatings used in GC capillaries are not chemically bonded to the capillary surface. In conventional approaches, these relatively thin coatings are immobilized on the capillary inner surface through free-radical cross-linking reactions. (Wright, B. W.; Peaden, P. A.; Lee, M. L.; Stark, T. J. J. Chromatogr. 1982, 248, 17-34; Blomberg, L. G. J. Microcol. September 1990, 2, 62-68). Immobilization of thicker coatings (especially the polar ones) is difficult to achieve. (Janak, K.; Horka, M.; Krejci, J. J. Microcol. September 1991, 3, 115-120; Berezkin, V. G.; Shiryaeva, V. E.; Popova, T. P. Zh. Analit. Khim. 1992, 47, 825-831). Third, because of the absence of direct chemical bonding between the stationary phase coating and the GC capillary inner walls, the thermal and solvent stabilities of such coatings are typically poor or moderate. When such extraction devices are coupled to GC, reduced thermal stability of thick GC coatings leads to incomplete sample desorption and sample carryover problems. (Buchholz, K. D.; Pawlyszyn, J. Anal. Chem. 1994, 66, 160-167; Zhang, Z.; Yang, M. J.; Pawliszyn, J. Anal. Chem. 1994, 66, 844A-853A.; Potter, D. W.; Pawliszyn, J. Environ. Sd. Technol. 1994, 28, 298-305).
Low solvent stability of conventionally prepared thick stationary phase coatings present a significant obstacle to the hyphenation of in-tube SPME with liquid-phase separation techniques that employ organic or organoaqueous mobile phase systems of the desorption of analytes. Solvent stability of the in-tube SPME coatings is, therefore, fundamentally important for further development of the technique. (Wu, J.; Pawliszyn, J. Anal. Chem. 2001, 73, 55-63). Thus, these three problems need to be solved in order to exploit full analytical potential of in-tube SPME.
In accordance with the present invention, there is provided methods of pre-concentrating trace analytes by extracting polar and non-polar analytes through a sol-gel coating and/or sol-gel monolithic bed. The present invention further provides microextraction methods including the steps of micro-extracting polar and non-polar analytes in a sol-gel coating and/or sol-gel monolithic bed, desorbing the analytes from the sol-gel and analyzing the extracted, desorbed analytes. The present invention also concerns microextraction devices useful for preconcentrating trace analytes, wherein the devices contain sol-gel extraction mediums. Another aspect of the present invention pertains a microextraction device in accordance with the present invention in hypenation with a chromatographic column, for example, a gas chromatographic column or a high-pressure liquid chromatographic column.
Other advantages of the present invention are readily appreciated as the same becomes better understood by reference to the following detailed description when considered in connection with the accompanying drawings wherein:
Generally, the present invention provides methods and apparatus for pre-concentrating trace analytes. Most generally, the methods involve the step of extracting polar and non-polar analytes through a sol-gel coating or monolithic bed. In a specific embodiment, the sol-gel has the formula:
wherein,
X=Residual of a deactivation reagent (e.g., polymethylhydrosiloxane (PMHS), hexamethyldisilazane (HMDS), etc.);
Y=Sol-gel reaction residual of a sol-gel active organic molecule (e.g., hydroxy terminated molecules including polydimethylsiloxane (PDMS), polymethylphenylsiloxane (PMPS), polydimethyldiphenylsiloxane (PDMDPS), poly(methyl-cyanopropylsiloxane) octadecylsilane, octylsilane, dendrimers, polystyrene, polystyrenedivinylbenzene, polyacrylate, molecularly imprinted polymers, polyethylene glycol (PEG) and related polymers like Carbowax 20M, polyalkylene glycol such as Ucon, macrocyclic molecules like cyclodextrins, crown ethers, calixarenes, alkyl moieties like octadecyl, octyl, etc.)
Z=Sol-gel precursor-forming chemical element (e.g., Si, Al, Ti, Zr, etc.)
l=An integer ≧0;
m=An integer ≧0;
n=An integer ÷0
p=An integer ≧0;
q=An integer ≧0;
and l, m, n, p, and q are not simultaneously zero.
Dotted lines indicate the continuation of the chemical structure with X, Y, Z, or Hydrogen (H) in space.
In order to achieve the desired sol-gels of the instant invention, certain reagents in a reagent system were preferred for the fabrication of the gels for the monolithic columns of the present invention. The reagent system included two sol-gel precursors, a deactivation reagent, one or more solvents and a catalyst. For the purposes of this embodiment, one of the sol-gel precursors contains a chromatographically active moiety selected from the group consisting of octadecyl, octyl, cyanopropyl, diol, biphenyl, phenyl, cyclodextrins, crown ethers and other moieties. Representative precursors include, but are not limited to: tetramethoxysilane, 3-(N-styrylmethyl-2-aminoethylamino)-propyltrimethoxysilane hydrochloride, N-tetradecyldimethyl(3-trimethoxysilylpropyl)ammonium chloride, N(3-trimethoxysilylpropyl)-N-methyl-N,N-diallylammonium chloride, N-trimethoxysilylpropyltri-N-butylammonium bromide, N-trimethoxysilylpropyl-N,N,N-trimethylammonium chloride, trimethoxysilyipropylthiouronium chloride, 3-[2-N-benzyaminoethylaminopropyl]trimethoxysilane hydrochloride, 1,4-bis(hydroxydimethylsilyl)benzene, bis(2-hydroxyethyl)-3-aminopropyltriethoxysilane, 1,4-bis(trimethoxysilylethyl)benzene, 2-cyanoethyltrimethoxysilane, 2-cyanoethyltriethoxysilane, (cyanomethylphenethyl)trimethoxysilane, (cyanomethylphenethyl)triethoxysifane, 3-cyanopropyldimethylmethoxysilane, 3-cyanopropyltriethoxysilane, 3-cyanopropyltrimethoxysilane, n-octadecyltrimethoxysilane, n-octadecyldimethylmethoxysilane, methyl-n-octadecyldiethoxysilane, methyl-n-octadecyldimethoxysilane, n-octadecyltriethoxysilane, n-dodecyltriethoxysilane, n-dodecyltrimethoxysilane, n-octyltriethyoxysilane, n-octyltrimethoxysilane, n-ocyldiisobutylmethoxysilane, octylmethyldimethoxysilane, n-hexyltriethoxysilane, n-isobutyltriethoxysilane, n-propyltrimethoxysilane, phenethyltrimethoxysilane, n-phenylaminopropyltrimethoxysilane, styrylethyltrimethoxysilane, 3-(2,2,6,6-tetramethylpiperidine-4-oxy)-propyltriethoxysilane, n-(3-triethoxysilylpropyl)acetyl-glycinamide, (3,3,3-trifluoropropyl)trimethoxysilane, and (3,3,3-trifluoropropyl)methyldimethoxysilane.
In one specific embodiment, a second sol-gel precursor, N-octadecyldimethyl[3-(trimethoxysilyl)propyl]ammonium chloride, was found to be desirable since it possessed an octadecyl moiety that allowed for chromatographic interactions of analytes with the monolithic stationary phase. Additionally, this reagent served to yield a positively charged surface, thereby providing the relatively high reversed electroosmotic flow necessary in capillary electrochromatography. However, it is considered within the scope to use any other reagent as known to one of ordinary skill in the art that would contain the octadecyl moiety for the purposes already set forth.
The deactivation reagent comprises a material reactive to polar functional groups bonded to the sol-gel precursor forming element in the coating or tube structure. Preferably, the deactivation reagent is reactive to hydroxyl functional groups. In one specific embodiment, the deactivation reagent is phenyldimethylsilane, polymethylhydrosiloxane, 1,1,1,3,3,3-hexamethyldisilazane, or a combination of any of the foregoing.
The sol-gel catalyst is generally either an acid, a base, or a fluoride compound. Preferably, the catalyst is trifluoroacetic acid (TFA). Advantageously, TFA can also function as a source of water. Preferably, the TFA contains about 5% water (i.e., TFA/water 95:5 v/v).
More specifically, the present invention provides the method of pre-concentrating both polar and non-polar analytes by feeding a sample through a sol-gel coated inner-surface of a tube or through a sol-gel monolithic bed and extracting analytes from the sample utilizing the sol-gel coating.
The preparation of the sol-gel coating on the inner surface of a tube includes the steps of providing the tube structure, a sol-gel solution comprising a sol-gel precursor, an organic material with at least one sol-gel active functional group, a sol-gel catalyst, a deactivation agent, and a solvent system as defined above. In one embodiment, the tube is hydrothermally pretreated. The sol-gel solution is then reacted with the inner surface of the tube under controlled conditions to produce a surface bonded sol-gel coating on that portion of the tube. The solution is then removed from the tube under pressure of an inert gas and is heated under controlled conditions to cause the deactivation reagent to react with the surface bonded sol-gel coating to deactivate and to condition the sol-gel coated portion of the tube structure. Preferably, the sol-gel precursor includes an alkoxy compound. The organic material includes a monomeric or polymeric material having at least one sol-gel active functional group. The sol-gel catalyst is taken from the group consisting of an acid, a base, and a fluoride compound. The deactivation reagent includes a material reactive to polar functional groups, such as hydroxyl groups, bonded to the sol-gel precursor-forming element in the coating or the tube structure.
The monolithic bed is made by first filling a tube with the sol-gel solution. By this single step filling, a sol-gel is produced that forms a porous matrix to be used for separation purposes. The matrix includes a positively charged surface within the matrix. That is, the microstructure of the sol-gel monolithic separation bed constitutes an infinite number of pathways through the porous matrix. The charges on the surface of the matrix generate an electroosmotic flow. Since the surface is positively charged, a reverse flow is created that is much easier to control than that of the prior art native silica surfaces. Such a monolithic bed provides a particle free sol-gel solution, which forms the preparation bed.
The sol-gel coating and monolithic bed can occur through numerous methodologies, alternative to the above. In a further coating method, the surface is coated with a sol-gel solution than includes a sol-gel precursor that includes, but is not limited to, alkoxysaline precursors such as methyltrimethoxysilane, trialkoxysilanes, and any other similar precursors known to those skilled in the art. Additionally, the sol solution contains a coating polymer that includes, but is not limited to hydroxy terminated polydimethylsiloxane. Finally, common to the preparation as discussed above, the solution contains a deactivation reagent and a sol-gel catalyst. The deactivation reagent can be, but is not limited to, PMHS and any other similar substance known to those skilled in the art. As for the sol-gel catalyst, an acid catalyst such as trifluoroacetic acid containing 5% water (TFA/H2O 95:5 v/v) is preferred. Once the sol-gel solution is placed onto the surface of the material, conditioning occurs under various parameters known to those skilled in the art. Furthermore, there are two major sets of reactions that take place during sol-gel processing the hydrolysis of the precursor and the polycondensation of the hydrolyzed products and other sol-gel active moieties in the system.
The present invention also encompasses capillary microextraction devices comprising transition metal oxide based hybrid organic-inorganic sorbent, sol-gel coatings. Advantageously, transition-based metal oxides exhibit stability across a wide range of pHs (Nawrocki, J. et al., J. Chromatogr. A 657 (1993) 229; Nawrocki, J. et al., J. Chromatogr. A 1028 (2004) 1; Nawrocki, J. et al., J. Chromatogr. A 1028 (2004) 31) and thus, can be utilized at both low pHs and high pHs where a silica based microextraction device would typically either become hydrolytically unstable (at extremely acidic conditions) or dissolve (at alkaline conditions beginning at a pH of about 8).
Suitable transition metal oxides include, without limitation, zirconia, alumina, and titania. Preferably, the transition metal oxide is zirconia, in part, because it exhibits superior alkali resistance over alumina and titania. Advantageously, zirconia is insoluble within a wide range of pH values (from about 1 to about 14), it exhibits resistance to dissolution at high temperatures and chemical inertness, and it possesses high mechanical strength (Kawahara, M. et al., J. Chromatogr. 515 (1990) 149; Trammell, B. C. et al., Anal. Chem. 73 (2001) 3323; Sun, L. et al. J. Colloid Interface Sci. 163 (1994) 464; Unger et al. High Performance Liquid Chromatography, Wiley, New York, 1989, p. 145; Bien-Vogelsang, U. et al., Chromatographia 19 (1984) 170; Yu, J. et al. J. Chromatogr. 631 (1993) 91).
An exemplary transition metal based sorbent, sol-gel coating is provided below.
wherein m=An integer ≧0;
n=An integer ≧0
x=An integer ≧0;
y=An integer ≧0;
and m, n, x, and y are not simultaneously zero. Dotted lines indicate the continuation of the chemical structure with Zr, Si, or Hydrogen (H) in space. The inner surface of the capillary is represented on the for left as thick bars.
Another aspect of the present invention concerns methods for preparing transition metal based sorbent, sol-gel microextraction devices. The methods comprise providing a hollow capillary and filling the capillary with a sol solution that can chemically bind to the inner surface of the capillary to form a sorbent extraction medium. In a specific embodiment, the capillary is a fused silica capillary. In yet another specific embodiment, the capillary is a hydrothermally treated capillary. In yet another specific embodiment, the capillary is a fused silica, hydrothermally pretreated capillary.
The sol solution includes a transition metal-based sol-gel precursor, a sol-gel active organic component, a chelating reagent, and a deactivating reagent in a solvent. The transition metal-based sol-gel precursor is selected from reactive transition-metal alkoxides. In one embodiment, the sol-gel precursor is selected from transition-metal alkoxides excluding silica alkoxides. Exemplary alkoxides are those with a zirconium component including, for example and without limitation, zirconium isopropoxide and zirconium tetrapropoxide. Preferably, the sol-gel precursor is zirconium(IV) butoxide.
In one embodiment, the sol-gel active organic component is selected from polydimethylsiloxane (PDMS), polymethylphenylsiloxane (PMPS), silanol-terminated polydimethyldiphenylsiloxane (PDMDPS), or poly(methylcyanopropylsiloxane). Preferably, the sol-gel active component is PDMDPS.
In yet another embodiment, the sol-gel active organic component is selected from octadecylsilane, octylsilane, dendrimers, polystyrene, polystyrenedivinylbenzene, polyacrylate, molecularly imprinted polymers, polyethylene glycol (PEG) and related polymers like Carbowax 20M, polyalkylene glycol such as Ucon, macrocyclic molecules like cyclodextrins, crown ethers, and calixarenes, and alkyl moieties, for example, octadecyl, and octyl.
The chelating reagent is utilized to slow the hydrolysis rate of the sol-gel active precursor and to prevent the transition metal from precipitating out of solution. Any ligand exchange reactant will work with the methods of the present invention, but exemplary chelating agents include, without limitation, glacial acetic acid, valeric acid, β-diketone, triethanolamine, and 1,5-diaminopentane.
The deactivation reagent is selected to derivatize any surface hydroxyl groups that are bound to metals. These sites are strongly adsorptive for polar solutes and their presence results in sample loss, sample carryover, and peak distortion and tailing. The deactivation reagent is a reactive silicon hydride. Preferably, the deactivating reagent is an alkyl hydrosilane or hexamethyldisilazane. In a specific embodiment, the sol solution comprises two deactivation reagents. Prefereably, the two deactivation reagents are poly(methylhydrosiloxane) and 1,1,1,3,3,3-hexamethyldisilazane.
The solvent utilized in the methods for preparing the transition metal-based sol-gel extraction mediums include proponol, 1-butanol, methylene chloride, and a combination of any of the foregoing.
Another aspect of the present invention pertains to an apparatus useful for preconcentrating and identifying target analytes in a sample. The apparatus comprises a microextraction device of the present invention in hyphenation with a chromatographic column.
As detailed in the experimental section below, specific formulations and methods are provided herein.
Any elements or limitations of any invention or embodiment thereof disclosed herein can be combined with any and/or all other elements or limitations (individually or in any combination) or any other invention or embodiment thereof disclosed herein, and all such combinations are contemplated with the scope of the invention without limitation thereto.
Three series of experiments were conducted demonstrating the applicability and utility of the present invention. Each section discussed below demonstrates both the open tube and monolithic bed columns ability to separate polar and non-polar analytes, even during the same extraction. Likewise, each set of experiments demonstrates the ability of the present invention to separate trace analytes at what was prior thought to be inconceivable trace amounts. Parts per quadrillion extractions were obtained utilizing the monolithic bed of the present invention.
Experimentation Series 1
Chemicals and materials. Fused silica capillary of 250 μm internal diameter (i.d.) was purchased from Polymicro Technologies (Phoenix, Ariz.). HPLC-grade methylene chloride and methanol were purchased from Fisher Scientific (Pittsburgh, Pa.). Trifluoroacetic acid (TFA) and polylrethylhydrosiloxane (PMHS) were procured from Aldrich Chemical Co. (Milwaukee, Wis.). Methyltrimethoxysilane (MTMS) was obtained from United Chemical Co. (Bristol, Pa.). Highly pure deionized water (18 Ω) was prepared in-house from a Barnstead model 04741 Nanopure deionized water system (Barnstead-Thermodyne, Debuque, Iowa). Eppendorf micro centrifuge tubes (1.5 mL) were purchased from Brinkman Instruments (Westbury, N.Y.).
Equipment. Gas chromatographic experiments were carried out on a Shimadzu 17 GC system (Shimadzu Scientific, Baltimore, Md.) equipped with a split/splitless injector 4 and a flame ionization detector (FID). A Barnstead model 04741 Nanopure deionized water system (Bamstead/Thermodyne, Debuque, Iowa) was used to prepare highly pure deionized water (18 Ω). A Microcentaur model APO 5760 centrifuge (Accurage Chemical and Scientific Corp., Westbury, N.Y.) was employed for necessary centrifugation of the sol-gel solution. A Fisher model G-560 Vortex Genie 2 system (Fisher Scientific, Pittsburgh, Pa.) was used for thorough mixing of the sol solution ingredients while preparing the sol solutions. A home-made gas-pressure operated filling/purging device (J. D. Hayes, A. Malik, Anal. Chem. 2000, 72, 4090-4099) was used for filing the fused silica capillary with the sol solution, as well as rinsing and purging with helium at various stages of column preparation.
Preparation of the sol-gel solution. For this, 0.1 g of the selected sol-gel-active polymer was dissolved in 100 μL of methylene chloride placed in a micro centrifuge tube (1.5 mL). PMHS (40 μL) and MTMS (100 μL) were added to this solution, and the contents of the centrifuge tube were thoroughly vortexed. Finally, 100 μL of TFA containing 5% water (sol-gel catalyst and source of water) was added and centrifuged for 4 mm at 13,000 rpm (15,682 G) to separate out any precipitate that might have formed during the mixing process. The clear sol solution from the top part of the centrifuge tube was then transferred to a clean vial for further use.
Preparation of Coatings for Capillary Microextraction. The sol-gel solution was used to prepare open tubular GC columns following a general procedure described in an earlier publication (D. X. Wang, S. L. Chong, and A. Malik, Anal. Chem., 1997, 69 (22), 4566-4576). Briefly, a hydrothermally treated fused silica capillary (10-m×250 μm i.d.) was filled with the sol-gel solution using a home-made filling/purging device (Hayes, J. D.; Malik, A. J. Chromatogr. B., 1997, 695, 3-13) under 100 psi helium pressure. The solution was allowed to stay inside the capillary for 15 min after which it was expelled from the capillary under the same helium pressure. Following this, the capillary was dried by purging it with helium for 30 min at room temperature. The capillary was then thermally conditioned under a continuous flow of helium: from 40° C. to 280° C. @ 1° C. min−1, holding the column at the final temperature for five hours. Finally, the column was rinsed with methylene chloride and dried under helium purge. At this point, the column was ready for analytical use.
Preparation of Monolithic beds for Microextraction. The monolithic extraction beds were prepared according to J. D. Hayes, A. Malik, Anal. Chem. 2000, 72, 4090-4099. Briefly, a fused silica capillary was filled with the sol solution and allowed to stay inside the capillary for an extended period (a few hours). During this time sol-gel reactions proceed inside the capillary and the sol solution transforms into a porous solid matrix. The capillary is then heated (from 30° C. to the final temperature, which may vary depending on the monolith material using a program rate of @ 0.2° C. min−1) with both ends sealed, holding it at the final temperature for two hours. After this, the monolithic capillary is rinsed with a series of appropriate solvents (e.g., methylene chloride, methanol and water). Finally, the monolith is thermally conditioned (e.g., at 300° C.) with continuous purge with an inert gas before use for extraction.
Result and Discussion
Experimentation Series 2
Equipment. SPME-GC experiments were carried out on a Varian Model 3800 capillary GC system equipped with an FID and a Varian Model 1079 temperature programmable split-splitless injector. Simple modifications to the split/splitless injector were made such that an extraction capillary could be inserted completely inside the injection port. An in-house designed liquid sample reservoir (
Chemicals and materials. Fused silica capillary (250-Ωm i.d.) with protective polyimide coating was purchased from Polymicro Technologies Inc. (Phoenix, Ariz.). Naphthalene and HPLC-grade solvents (tetrahydrofuran (THE), methylene chloride, and methanol) were purchased from Fisher Scientific (Pittsburgh, Pa.). The ketone, 4′-phenylacetophenone, was obtained from Eastman Organic Chemicals (Rochester, N.Y.). Hexamethyldisilazane (HMDS), poly(methylhydrosiloxane) (PMHS), trifluoroacetic acid (TFA), ketones (valerophenone, hexanophenone, heptonophenone, decanophenone, anthraquinone), aldehydes (benzaldehyde, nonylaldehyde, tolualdehyde, n-decylaldehyde, undecylic aldehyde), PAHs (acenaphthylene, flourene, phenanthrene, fluoranthene), and phenols (2,6-dimethylphenol, 2,5-dimethylphenol, 2,3-dimethylphenol, 3,4-dimethylphenol) were purchased from Aldrich (Milwaukee, Wis.). Hydroxy-terminated poly(dimethylsiloxane) (PDMS) and methylthrimethoxysilane (MTMS) were purchased from United Chemical Technologies, Inc. (Bristol, Pa.). Trimethoxysilane-derivatized polyethylene glycols (M-SIL-5000 and SIL-3400) were obtained from Shearwater Polymers (Huntsville, Ala.).
Preparation of Aqueous Standard Solutions for Capillary Microextraction. Stock solutions of polycyclic aromatic hydrocarbons (PAHs) (naphthalene, acenaphthylene containing 20% acenaphthene, fluorene, phenanthrene, fluoranthene) and ketones (4′-phenylacetophenone and anthraquinone) were prepared by dissolving 10 mg of each compound in 10 mL of THF in a 10 mL volumetric flask at room temperature. A 25-μL portion of this standard solution was diluted with deionized water to give a total volume of 25 mL that corresponded to a 1 ppm PAH aqueous solution. Preparation of 100 ppb and 1 ppb PAH solutions were accomplished by further dilution of this stock solution with deionized water. Stock solutions of ketones (valerophenone, hexanophenone, heptonophenone, decanophenone) or aldehydes (benzaldehyde, nonylaldehyde, tolualdehyde, n-decylaldehyde, undecyclic aldehyde) were also prepared using THF as the initial organic solvent. Stock solutions of dimethylphenol (DMP) isomers (2,6-dimethylphenol, 2,5-dimethylphenol, 2,3-dimethylphenol, 3,4-dimethylphenol) were prepared in an analogous way-using methanol as the initial organic solvent. Prior to extraction, all glassware was deactivated. The glassware was cleaned using Sparkleen detergent and rinsed with generous amounts of deionized water, and dried at 150° C. for two hours. The inner surface of the dried glassware was then treated with 5% v/v solution of HMDS in methylene chloride, followed by placing of the glassware in an oven at 250° C. overnight. The glassware were then rinsed sequentially with methylene chloride and methanol, and further dried in the oven at 100° C. for 1 hour. Before use, they were rinsed with generous amounts of deionized water and dried at room temperature in a flow of helium.
Preparation of Sol-gel Coated Capillaries. Sol-gel PDMS and PEG extraction capillaries, as well as the sol-gel open-tubular GC columns, were prepared according to procedures described elsewhere and as follows. (Wang, D. X., Chong, S. L., Malik, A. Anal. Chem. 1997, 69, 4566-4576). Briefly a previously cleaned and hydrothermally treated fused silica capillary was filled with a specially designed sol solution using a helium pressure operated filling/purging device (
The sol-gel coated capillary was further purged with helium for one hour, and conditioned in a GC using temperature programming (from 40° C. @ 1° C./min). The capillary was held at the final temperature (350° C. for sol-gel PDMS and 300° C. for sol-gel PEG) for five hours. During conditioning, the capillary was constantly purged with helium at a linear velocity of 20 cm/s. Before using for extraction, the capillary was sequentially rinsed with methylene chloride and methanol followed by drying the capillary in a stream of helium under temperature programming (from 40° C. to 250° C., @ 4° C./min, 60 min at 250° C.).
Gravity-Fed Sample Reservoir for Capillary Microextraction. The gravity-fed sample reservoir for capillary microextraction (
Thermal Desorption of Extracted Analytes in the GC Injection Port. To facilitate thermal desorption of the extracted analytes from sol-gel microextraction capillary for their subsequent introduction into the GC capillary column, the Varian Model 1079 split/splitless injector was slightly modified. For this, the quartz wool was removed from the glass insert to accommodate a two-way fused silica connector within the insert. With the glass insert (now without the quartz wool) in place, the injection port was cooled down to ambient temperature. The metallic nut at the top of the injector together with the rubber septum, the septum support, and the glass insert were temporarily removed from the injection port. The injector end of the GC capillary column was then pushed for the bottom of the injector to pass it through the injection port and the glass insert (now located outside the port) such that about 10-15 cm of the capillary column extends out from the top of the insert. This end of the column was press-fitted into the lower end of the deactivated two-way fused silica connector. The two-way connector with press-fitted column end was then secured inside the glass insert, which was subsequently placed back into injection port so that the two-way butt connector with the attached GC column head remained within the glass insert. The septum support was replaced on top of the glass insert. The septum was replaced, and the injector nut was tightened down. Finally, the capillary column was secured inside the oven by tightening the ferrule connection at the bottom of the injection port. After performing capillary microextraction, the extraction capillary was connected to the system in the following way. The capillary column nut at the bottom of the injector was loosened and the column was slid up. The extraction capillary was passed through the septum support and pressed-fitted into the fused silica two-way butt connector. The column was then pulled down until the extraction capillary disappeared below the septum support and remained inside the glass insert (
Sol-gel Capillary Microextraction-GC Analysis. Sol-gel PDMS and sol-gel PEG-coated capillaries were used for extraction. Prior to extraction, the sol-gel coated extraction capillary was first thermally conditioned with a simultaneous flow of helium through it. This involved using helium carrier gas at 10 psi and setting the initial GC temperature at 40° C. with the extraction capillary connected to the injection port. The GC temperature was increased at a rate of 5° C./min until 250° C. was reached. The rate of GC temperature increase was then changed to 1° C./min. For sol-gel PDMS- and sol-gel PEG-coated capillaries the final conditioning temperatures were 350° C. and 300° C., respectively. The extraction capillaries were held at the final temperatures for 60 minutes with continuous helium flowing through them.
After conditioning, the extraction capillary was cooled to room temperature, removed from the GC, and installed on the homemade sample reservoir (
To perform capillary microextraction the extraction capillary was vertically connected to the empty reservoir by removing the PEEK tubing nut from the bottom screw cap, inserting the capillary into the peek tubing (so that ˜1 cm of capillary remained extended from the bottom and ˜0.5 cm from the top of the tubing), and reassembling the apparatus. The aqueous sample (25 mL) was placed in the reservoir, and allowed to flow through the extraction capillary under gravity for 30 minutes for equilibrium to be established. After this, the microextraction capillary was removed from the gravity-fed apparatus and immediately connected to the homemade capillary filling/purging device (
The extracted analytes were then thermally desorbed from the capillary by rapid temperature programming of the injector (@ 100° C./min starting from 30° C.). The nature of the coating used in the capillary determined the final temperature of the ramp (330° C. for sol-gel PDMS and 280° C. for sol-gel PEG-coated capillaries). The desorption was performed over the five-minute period, whereby the released analytes were swept over by the carrier gas into the GC column. The thermal desorption step was accomplished in the splitless mode, keeping the column temperature at 30° C. to promote effective solute focusing at the column inlet. After thermal desorption, the split vent remained closed throughout the course of the chromatographic run. The GC separations were performed using in-house prepared sol-gel-coated open tubular PDMS columns (10 m×0.25 mm i.d.). After the sample was introduced into the column, the column oven temperature was increased at a rate of 15° C./min. GC analysis were carried out using helium as the carrier gas. Analyte detection was performed using a flame ionization detector (FID). The FID detector temperature was maintained at 350° C.
Results and Discussion.
Sol-gel technology provides an elegant synthetic pathway to advanced materials (Novak, B. M. Adv. Mater. 1993, 5, 422-433; Livage, J. In Applications of Organometallic Chemistry in the Preparation and Processing of Advanced Materials, Harrod, J. F., Lame, R. M. Eds.; Kiuwer: Dordrecht, The Netherlands, 1995; pp. 3-25; Walsh, D.; Whiton, N. T. Chem. Mater. 1997, 9, 2300-2310) with a wide range of applications. (Aylott, J. W. et al. Chem. Mater. 1997, 9, 2261-2263; Collinson, M. M. et al. Chem. 2000, 72, 702A-709A; Lobnik, A. et al. Sens. Actuators B 1998, 51, 203-207; Vorotilov, K. A. et al. J. Sol-gel Sci. Technol. 1997, 8, 581-584; Reisfeld, R.; Jorgenson, C. K. (eds.), Spectroscopy, Chemistry, and Applications of Sol-gel Glasses, Springer-Verlag, Berlin, 1992; Lev, O. et al. S. Chem. Mater. 1997, 9, 2354-2375; Atik, M. et al., J. Sd. Gel. Sci. Technol. 1997, 8, 517-522; Haruvy, Y. et al. N. Chem. Mater. 1997, 9, 2604-2615; Fabes, B. D. et al. J. Am. Ceram. Soc. 1990, 73, 978-988; Sakka, S. et al. In Chemistry, Spectroscopy, and Applications, Reisfeld, R., Jorgenson C. K. Eds.; Springer-Verlag: Berlin, 1992; pp. 89-118). In the context of analytical microseparations, it allows for the in situ creation of hybrid organic-inorganic stationary phases within separation columns in the form of coatings, (Rodriguez, S. A. et al. Chem. Mater. 1999, 11, 754-762; Rodriguez, S. A. et al. Anal. Chem. Acta 1999, 397, 207-215; Guo Y. et al. J. Microcol. September 1995, 7, 485-491; Guo, Y. et al. Anal. Chem. 1995, 67, 2511-2516; Guo, Y. et al. Chromatographia 1996, 43, 477-483; Guo, Y. et al. J. Chromatogr. A. 1996, 744, 17-29; Narang, P. et al. J. Chromatogr. A. 1997, 773, 65-72; Hayes, J. D. et al. J. Chromatogr. B 1997, 695, 3-13; Hayes, J. D. et al. Anal. Chem. 2001, 73, 987-996) monolithic beds, (Cortes, H. J. et al. J. High. Resolut. Chromatogr./Chromatogr. Commun. 1987, 10, 446-448; Fields, S. M. Anal. Chem. 1996, 68, 2709-2712; Hayes, J. D. et al. Anal. Chem. 2000, 72, 4090-4099; Nakanishi, K. et al. J. Sol. gel. Sci. Technol. 1997, 8, 547-552; Duly, M. T. et al. Anal. Chem. 1998, 70, 5103-5107; Fujimoto, C. J. High Resol. Chromatogr. 2000, 23, 89-92; Roed, L. et al. J. Micro September 2000, 12, 561-567) and stationary phase particles. (Reynolds, K. J. et al. J. Liq. Chromatogr. & Rel. Technol. 2000, 23, 161-173; Pursch, M. et al. Chem. Mater. 1996, 8, 1245-1249) Excellent chromatographic and electromigration separations have been demonstrated using separation columns with sol-gel stationary phases. (Wang, D. X. Sol-gel Chemistry-Mediated Novel Approach to Column Technology for High-Resolution Capillary Gas Chromatography, Ph.D. Dissertation, University of South Florida, Department of Chemistry: Tampa, Fla., 2000; Wang, D. X. et al. Anal. Chem. 1997, 69, 4566-4576; Guo, Y., et al. Anal. Chem. 1995, 67, 2511-2516; Hayes, J. D. et al. J. Chromatogr. B 1997, 695, 3-13; Hayes, J. D. et al. Anal. Chem. 2001, 73, 987-996; Hayes, J. D. “Sol-Gel Chemistry-Mediated Novel Approach to Column Technology for Electromigration Separations,” Ph.D. dissertation, Department of Chemistry, University of South Florida, Tampa, Fla., USA, 2000; Tang, Q. et al. J. Chromatogr. A 1999, 837, 35-50; Cabrera, K. et al. J High Resol. Chromatogr. 2000, 23, 93-99; Chen, Z., et al. Anal. Chem. 2001, 73, 3348-3357; Roed, L. et al. J. Chromatogr. A 2000, 890, 347-353.) Applicants introduced Sol-gel coatings for gas chromatography (Wang, D. X. et al. A. Anal. Chem. 1997, 69, 4566-4576) and solid-phase microextraction (Chong, S. L. et al. Anal. Chem. 1997, 69, 3889-3898) in 1997, and demonstrated significant thermal and solvent stability advantages inherent in sol-gel coated GC columns (Wang, D. X. Sol-gel Chemistry-Mediated Novel Approach to Column Technology for High-Resolution Capillary Gas Chromatography, Ph.D. Dissertation, University of South Florida, Department of Chemistry: Tampa, Fla., 2000) and SPME fibers. (Malik, A. et al. In Applications of Solid-phase Microextraction, Pawliszyn, J. Ed.; Royal Society of Chemistry (RSC): Cambridge (UK), 1999; pp. 73-91) Since then several other groups have got involved in sol-gel research for solid phase microextraction (Gbatu, T. P. et al. Anal. Chim. Acta 1999, 402, 67-79; Wang, Z. Y. et al. J. Chromatogr. A 2000, 893, 157-168; Zeng, Z. et al. Anal. Chem. 2001, 73, 2429-2436) and solid-phase extraction. (Senevirante, J. et al. Talanta 2000, 52, 801-806).
Because of the advanced material properties, sol-gel coatings and monolithic beds can also be expected to serve as excellent extraction media in capillaries as an effective means of solventless microextraction, and call such a microextraction technique as Sol-gel Open Tubular Microextraction (OTME). Sol-gel OTME is synonymous with in-tube solid-phase microextraction (in-tube SPME) on sol-gel coated capillaries. Both sol-gel OTME and sol-gel monolithic microextration (MME) can be combined under a general term—Capillary Microextraction (CME). The new terminology provides a better reflection of the techniques, since “In-Tube Solid-phase Microextraction” is not necessary limited to the use of only “solid phases” as the extraction media. In fact, liquid stationary phase coatings are commonly used both in in-tube SPME as well as conventional SPME.
Sol-gel technology allows of the creation of coatings on the inner surface of open tubular GC, CE, and CEC columns (Wang, D. X. et al. Anal. Chem. 1997, 69, 4566-4576; Guo, Y. et al. Anal. Chem. 1995, 67, 2511-2516; Hayes, J. D. et al. Anal. Chem. 2001, 73, 987-996) as well as on the outer surface of substrates of different shapes and geometry (e.g., SPME fibers. (Chong, S. L. et al. Anal. Chem. 1997, 69, 3889-3898; Gbatu, T. P., et al. Anal. Chim. Acta 1999, 402, 67-79; Wang, Z. Y. et al. J. Chromatogr. A 2000, 893, 157-168; Zeng, Z. et al. Anal. Chem. 2001, 73, 2429-2436.). It is applicable to the creation of silica-based, (Her, R. K. The Chemistry of Silica, Wiley, New York, 1979; Brinker, C. J.; Scherer, G. W. Sol-Gel Science, Academic Press, San Diego, Calif., 1990; Rabinovich, E. M. In Sol Gel Technology for Thin Films, fibers, Pre forms, Electronics, and Specialty Shapes, Klein, L. C. Ed.; Noyes Publications: Park Ridge, N.J., 1988; pp. 260-294) and transition metal-based (Livage, J. et al. Prog. Solid. St. Chem. 1988, 18, 259-341; In, M. et al. J. Sol. gel. Sd. Technol. 1995, 5, 101-114; Jiang, Z., et al. Anal. Chem. 2001, 73, 686-688; Silva, R. B. et al. J. Sep. Sd. 2001, 24, 49-54; Palkar, V. R. Nanostructured Mater. 1999, 11, 369-374; Chaput, F. et al. J. Non. Cryst. Solids. 1995, 188, 11-18) and silica/nonsilica mixed systems. (Dutoit, D. C., et al. J. Catal. 1995, 153, 165-176; Jones, S. A. et al. Chem. Mater. 1997, 9, 2567-2576; Kosuge, K. et al. J. Phys. Chem. B1 999, 103, 3562-3569).
In the context of capillary separation and sample pre-concentration techniques, the most important attribute of sol-gel coating technology is that it provides surface coatings that become automatically bonded to the substrate surfaces containing sol-gel-active functional groups (e.g., silonal groups). This direct chemical bonding results in enhanced thermal and solvent stability of sol-gel coatings. The attributes of thermal and solvent stability of the stationary phase coatings are enormously important in analytical separation and sample pre-concentration.
Advantageously, sol-gel technology also allows for the stationary phase coating, its immobilization, and deactivation to be achieved in one single step (Wang, D. X. et al. Anal. Chem. 1997, 69, 4566-4576) instead of multiple time-consuming steps involved in conventional coating technology.
The use of the stationary phase coating on the inner surface of a fused silica capillary eliminates coating scraping problem inherent in fiber-based SPME and significantly reduces the possibility of sample contamination. Furthermore, the protective polyimide coating on the outer surface of the fused silica extraction capillary adds flexibility to the extraction device as compared with traditional SPME fibers.
Sol-gel PDMS- and PEG-coated were used in conjunction with the gravity-fed sample reservoir (
Sol-gel capillary microextraction typically uses a short length of fused silica capillary coated internally with sol-gel stationary phase. The extraction is carried out by attaching the extraction capillary to an in-house designed gravity-fed extraction apparatus (
Sol-gel coating technology can easily produce thick coatings (Chong, S. L. et al. Anal. Chem. 1997, 69, 3889-3898; Wang, Z. Y. et al. J. Chromatogr. A 2000, 893, 157-168; Zeng, Z. et al. Anal. Chem. 2001, 73, 2429-2436) (df>μm). For example, Zeng et al. recently reported SPME on sol-gel coated fibers with a coating thickness of 76 μm. The use of microextraction capillaries with thick sol-gel coatings should lead to higher sensitivity of capillary microextraction. It can be expected that the use of capillaries with larger inner diameter and thicker sol-gel coatings should lead to further enhancement of this extraction sensitivity.
In this work, the extraction capillary length was relatively short—only 3.5 cm. The use of such a short length was dictated by the linear dimensions of the glass insert of the injection port and that of the press-fit connector. In principle longer extraction capillaries can be employed using a different configuration of the coupling, between the extraction capillary and the open tubular GC column.
In this work, aldehydes were extracted and analyzed without derivatization. This became possible due to outstanding material properties of sol-gel PDMS coating used both in the microextraction capillary as well as in the GC separation column. Organic-inorganic nature of the sol-gel PDMS coating provides sorption sites both of the polar and non-polar analytes. High quality of column deactivation achieved through sol-gel column technology (Wang, D. X. et al. Anal. Chem. 1997, 69, 4566-4576) allows the GC analysis of aldehydes without derivatization. From an analytical standpoint, the possibility of extraction and gas chromatographic analysis of underivatized aldehydes by OEME-GC is important and should provide simplicity, speed, sensitivity, and accuracy in aldehyde analysis.
The, sol-gel OTME-GC for aldehydes and ketones described herein provides a number of important advantages over sample preparation techniques coupled to HPLC. First, the fact that no derivatization is needed makes the procedure faster, simpler, and more accurate. Second, since the flame ionization detector used for GC analysis inherently possess several order of magnitude higher sensitivity compared with the UV detector commonly used with the HPLC analysis, the described procedure also provides sensitivity advantage. The OTME-GC analysis of the aldehydes and ketones is also characterized by low run-to-run RSD values (Table II). For five replicate measurements, RSD values of under 6% and 0.4% were obtained for solute peak area and retention time, respectively, the only exception was benzaldehyde that had a retention time RSD value of 1.9% which is significantly higher than the RSD values for the rest of the aldehydes and ketones studied.
Highly polar compounds, such as alcohols, amines, and phenols, have higher affinity for water. Conventional non-polar phases (e.g., PDMS) are usually not very efficient for their extraction from an aqueous phase. Polar coatings are normally, used for the extraction of these highly polar analytes. However, creation of thick coatings of polar stationary phases and their immobilization on a substrate are associated with technical difficulties. (Janak, K. et al. J. Microcol. September 1991, 3, 115-120). Previously, Applicants showed (Chong, S. L. et al Anal Chem. 1997, 69, 3889-3898; Janak, et al. 1991) that these polar compounds can be satisfactorily extracted and analyzed using sol-gel PDMS coatings. This becomes possible thanks to the organic-inorganic hybrid nature of the sol-gel PDMS coatings characterized by the presence of both polar and non-polar sorption sites. Sol-gel coating technology allows for the creation of both polar (Hayes, J. D. et al. J. Chromatogr. B 1997, 695, 3-13; Wang, Z. Y. et al. J. Chromatogr. A 2000, 893, 157-168) and non-polar (Chong et al. 1997; Gou et al., 2000; Wang, D. X. 2000; Wang, D. X. et al. 1997; Hayes, J. D., et al. 2001; Malik et al. 1999; Gbatu, T. P. et al. 1999) coatings with equal ease and versatility. In the present work, we demonstrate the possibility of efficient extraction of these polar analytes from an aqueous environment using open tubular capillary microextraction on sol-gel polyethylene glycol (sol-gel PEG) coatings.
The capillary-to-capillary reproducibility for open tubular microextraction was evaluated for the two types of sol-gel coatings used in this work—sol-gel PDMS and sol-gel PDMS and sot-gel PEG coatings. For this, three identical segments of each type of sol-gel coated capillary were used for extraction. Fluorene was used at the test solute for the sol-gel PDMS coated capillary while decanophenone served the same purpose for the sol-gel PEG coated microextraction capillary. A total of six extractions (30 min each) were carried out on each capillary using 1 ppm aqueous solutions containing the respective test solute. The relative standard deviations (RSD) of the mean GC peak area for the two test solute on sol-gel PDMS and sol-gel PEG capillaries were 3.9% and 3.0%, respectively. These low RSD values are indicative of excellent capillary-to-capillary reproducibility in sol-gel open tubular micro extraction.
In the present work, OTME was performed using 3.5 cm long sol-gel coated capillary segments. The length of the used extraction capillary was limited by the linear dimensions of the glass insert in the injection port of the used GC (Varian 38000) and the length of the two-way press-fit connector. In this format, the entire length of the extraction capillary was contained inside the GC injection port. However, sol-gel coated capillary segments of greater lengths can be used in GC systems that employ longer glass inserts, (e.g., Shimadzu 17) which should lead to enhanced sensitivity. Even in the present configuration, the coated segment was more than three times longer than coated segments used on conventional SMPE fibers. The desorption of the extracted analytes can be achieved by making a press-fit connection between the extraction capillary and the GC column outside the injection port. (Koivusalmi, E. et al Anal. Chem. 1999, 71, 86-91) Such a configuration will also allow for the use of sol-gel coated capillaries of greater lengths, significantly enhancing the sensitivity of the technique.
For the first time, sol-gel coated capillaries were used for solventless microextraction and sample pre-concentration, and the technique was termed sol-gel capillary microextraction (sol-gel CME). Two types of sol-gel coatings (sol-gel PDMS and sol-gel PEG) were effectively used for the extraction of analytes belonging to various chemical classes. Parts per trillion (ppt) and parts per quadrillion (ppq) level detection sensitivities were achieved for polar and non-polar analytes. Further sensitivity enhancements should be possible through the use of thicker sol-gel coatings in conjunction with longer extraction capillaries of larger inner diameter. The sol-gel coated capillaries are characterized by enhanced thermal and solvent stabilities (a prerequisite for efficient analyte desorption), making them very suitable for coupling with both GC and HPLC. Sol-gel capillary microextraction showed remarkable run-to-run and capillary-to-capillary repeatability, and produced peak area RSD values of less than 6% and 4% respectively.
Experimentation Series 3
Equipment. All CME-GC experiments were performed on a Shimadzu Model 14A capillary GC system equipped with a flame ionization detector (FID) and a split-splitless injector. On-line data collection and processing were done using ChromPerfect (version 3.5) computer software (Justice Laboratory Software, Denville, N.J.). A Fisher Model G-560Vortex Genie 2 system (Fisher Scientific, Pittsburgh, Pa.) was used for thorough mixing of various sol solution ingredients. A Microcentaur model APO 5760 microcentrifuge (Accurate Chemical and Scientific Corp., Westbury, N.Y.) was used to separate the sol solution from the precipitate (if any) at 13,000 rpm (15,682×g). A Nicolet model Avatar 320 FTIR instrument (Thermo Nicolet, Madison, Wis.) was used to acquire infrared spectra of the prepared sol-gel materials. A Bamstead Model 04741 Nanopure deionized water system (Barnstead/Thermodyne, Dubuque, Iowa) was used to obtain ˜16.0MΩ water. Stainless steel mini-unions (SGE Inc., Austin, Tex.) were used to connect the fused silica capillary GC column with the microextraction capillary, also made of fused silica. An in-house-designed liquid sample dispenser was used to facilitate gravity-fed flow of the aqueous sample through the sol-gel microextraction capillary. A homebuilt, gas pressure-operated capillary filling/purging device (Hayes, J.; Malik, J. Chromatogr. B. 1997, 695, 3-13) was used to perform a number of operations: (a) rinse the fused silica capillary with solvents; (b) fill the extraction capillary with the sol solution; (c) expel the sol solution from the capillary at the end of sol-gel coating process; and (d) purge the capillary with helium after treatments like rinsing, coating, and sample extraction.
Chemicals and materials. Fused-silica capillary (320 and 250 μm, i.d.) with a protective polyimide coating was purchased from Polymicro Technologies Inc. (Phoenix, Ariz.). Naphthalene and HPLC-grade solvents (methylene chloride, methanol) were purchased from Fisher Scientific (Pittsburgh, Pa.). Hexamethyldisilazane (HMDS), poly (methylhydrosiloxane) (PMHS), ketones (valerophenone, hexanophenone, heptanophenone, and decanophenone), aldehydes (nonylaldehyde, n-decylaldehyde, undecylic aldehyde, and dodecanal), polycyclic aromatic hydrocarbons (PAHs) (naphthalene, acenaphthene, fluorene, phenanthrene, pyrene, and naphthacene), were purchased from Aldrich (Milwaukee, Wis.). Two types of silanol-terminated poly(dimethyldiphenylsiloxane) (PDMDPS) copolymers (with 2-3% and 14-18% contents of the diphenyl-containing component) were purchased from United Chemical Technologies Inc. (Bristol, Pa.).
Preparation of sol-gel zirconia-PDMDPS coating. The sol solution was prepared in a clean polypropylene centrifuge tube by dissolving the following ingredients in a mixed solvent system consisting of methylene chloride and butanol (250 μL each): 10-15 μL of zirconium(IV) butoxide (80% solution in 1-butanol), 85 mg of silanol-terminated poly(dimethyldiphenylsiloxane) copolymer, 70 mg of poly(methylhydrosiloxane), 10 μL of 1,1,1,3,3,3-hexamethyldisilazane, and 2-4 μL of glacial acetic acid. The dissolution process was aided by thorough vortexing. The sol solution was then centrifuged at 13,000 rpm (15,682×g) to remove the precipitate (if any). The top clear sol solution was transferred to a clean vial and was further used in the coating process. A hydrothermally treated fused silica capillary (2 m) was filled with the clear sol solution, using pressurized helium (50 psi) in the filling/purging device (Hayes, J.; Malik, J. Chromatogr. B. 1997, 695, 3-13). The sol solution was allowed to stay inside the capillary for a controlled period of time (typically 15-30 min) to facilitate the formation of a sol-gel coating and its chemical bonding to the capillary inner walls. After that, the free portion of the solution was expelled from the capillary, leaving behind a surface-bonded sol-gel coating within the capillary. The sol-gel coating was then dried by purging with helium. The coated capillary was further conditioned by temperature programming from 40 to 150° C. at 1° C./min and held at 150° C. for 300 min. Following this, the conditioning temperature was raised from 150 to 320° C. at 1° C./min and held at 320° C. for 120 min. The extraction capillary was further cleaned by rinsing with 3 mL of methylene chloride and conditioned again from 40° C. to 320° C. at 4° C./min. While conditioning, the capillary was constantly purged with helium at 1 mL/min. The conditioned capillary was then cut into 10 cm long pieces that were further used to perform capillary microextraction.
Preparation of the samples. PAHs, ketones, and aldehydes were dissolved in methanol or tetrahydrofuran to prepare 0.1 mg/L stock solutions in silanized glass vials. For extraction, fresh samples with ppb level concentrations were prepared by diluting the stock solutions with deionized water.
Gravity-fed sample dispenser for capillary microextraction. The gravity-fed sample dispenser for capillary microextraction was constructed by in-house modification of a Chromaflex AQ column (Kontes Glass Co., Vineland, N.J.) consisting of a thick-walled glass cylinder coaxially placed inside an acrylic jacket. The inner surface of the thick-walled cylindrical glass column was deactivated by treating with a 5% (v/v) solution of HMDS in methylene chloride followed by overnight heating at 100° C. The column was then cooled to ambient temperature, thoroughly rinsed with methanol and liberal amounts of deionized water, and dried in a helium flow. The entire Chromaflex AQ column was subsequently reassembled.
Sol-gel capillary microextraction-GC analysis. To perform capillary microextraction, a previously conditioned sol-gel zirconia-PDMDPS coated microextraction capillary (10 cm×320 μm i.d. or 10 cm×250 μm i.d.) was vertically connected to the bottom end of the empty sample dispenser. The aqueous sample (50 mL) was then placed in the dispenser from the top and allowed to flow through the microextraction capillary under gravity. While passing through the extraction capillary, the analyte molecules were sorbed by the sol-gel zirconia-PDMDPS coating residing on the inner walls of the capillary. The sample flow through the capillary was allowed to continue for 30-40 min for an extraction equilibrium to be established. After this, the microextraction capillary was purged with helium at 25 kPa for 1 min and connected to the top end of a vertically placed two-way mini-union connecting the microextraction capillary with the inlet end of the GC column. Approximately, 6.5 mm of the extraction capillary remained tightly inserted into the connector, as did the same length of GC column from the opposite side of the mini-union facing each other within the connector. The installation of the capillary was completed by providing a leak-free connection at the bottom end of the GC injection port so that 9 cm of the extraction capillary remained inside the injection port. The extracted analytes were then thermally desorbed from the capillary by rapidly raising the temperature of the injector (up to 300° C. starting from 30° C.). The desorption was performed over a 8.2 min period in the splitless mode allowing the released analytes to be swept over by the carrier gas into the GC column held at 30° C. during the entire desorption process. Such a low column temperature facilitated effective solute focusing at the column inlet. Following this, the column temperature was programmed from 30 to 320° C. at a rate of 20° C./min. The split vent remained closed throughout the entire chromatographic run. Analyte detection was performed using a flame ionization detector (FID) maintained at 350° C.
Result and Discussion
Capillary microextraction (Bigham, S.; Kabir, A. et al. Anal Chem. 2002, 74, 752) uses a sorbent coating on the inner surface of a capillary and thereby, overcomes a number of deficiencies inherent in conventional fiber-based SPME such as susceptibility of the sorbent coating to mechanical damage due to scraping during operation, fiber breakage, and possible sample contamination. In CME, the sorbent coating is protected by the fused silica tubing against mechanical damage. The capillary format of SPME also provides operational flexibility and convenience during the microextraction process since the protective polyimide coating on the outer surface of fused silica capillary remains intact. Inner surface-coated capillaries provide a simple way to perform extraction in conjunction with a gravity-fed sample dispenser and thus, avoid typical drawbacks of fiber-based SPME, including the need for sample agitation during extraction as well as the sample loss and contamination problems associated with this.
The sol-gel process is a straightforward route to obtaining homogeneous gels of desired compositions. In recent years, it has received increased attention in analytical separations and sample preparations due to its outstanding versatility and excellent control over properties of the created sol-gel materials that proved to be promising for use as stationary phases and extraction media.
A general procedure for the creation of sol-gel stationary phase coating on the inner walls of fused silica capillary GC columns was first described by Malik and co-workers (Wang, D.; Chong, S. L.; Malik, A.; Anal Chem. 1997, 69, 4566). In the present work, a judiciously designed sol solution ingredients (Table III) was used to create the sol-gel zirconia-PDMDPS coating on the fused silica capillary inner surface. Zirconium(IV) butoxide (80% solution in 1-butanol) was used as a sol-gel precursor and served as a source for the inorganic component of the sol-gel organic-inorganic hybrid coating.
The sol-gel zirconia-PDMDPS coating presented here was generated via two major reactions: (1) hydrolysis of a sol-gel precursor, zirconium(IV) butoxide; and (2) polycondensation of both the precursor and hydrolysis products between themselves and other sol-gel-active ingredients in the coating solution, including silanol-terminated PDMDPS. The hydrolysis of the zirconium(IV) butoxide precursor is represented by Scheme 1 (Yoldas, B. E. J. Non-Cryst. Solid, 1984, 63, 145).
Condensation of the sol-gel polymer growing in close vicinity of the capillary walls with silanol group on the capillary surface led to the formation of an organic-inorganic coating chemically anchored to the capillary inner walls (Scheme 2).
A major obstacle to preparing zirconia-based sol-gel materials using zirconium alkoxide precursors (e.g., zirconium butoxide) is the very rapid sol-gel reaction rates for these precursors. Even if the solution of zirconium alkoxide is stirred vigorously, the rates of these reactions are so high that large agglomerated zirconia particles precipitate out immediately when water is added (Chang, C. H.; Gopalan, R.; Lin, Y. S. J. Membr. Sci. 1994, 91, 27). Such fast precipitation makes it difficult to reproducibly prepare zirconia sol-gel materials. Ganguli and Kundu (J. Mater. Sci. Lett. 1984, 3, 503) addressed the fast precipitation problem by dissolving zirconium propoxide in a non-polar dry solvent like cyclohexane. The hydrolysis was performed by exposing the coatings prepared from the solution to atmospheric moisture. Heating to 450° C. was necessary to obtain transparent films. The hydrolysis rates of zirconium alkoxides can also be controlled by chelating with ligand-exchange reagents. Acetic acid (Kandu, D; Biswas, P. K.; Ganguli, D. Thin Solid Films, 1988, 163, 273; Noonan, G. O.; Ledford, J. S. Chem. Mater 1995, 7, 1117), valeric acid (Severin, K. G.; Ledford, J. S.; Torgerson, B. A.; Berglund, K. A. Chem. Mater., 1994, 6, 890), β-diketones (Percy, M. J., Barlett, J. R., Spiccia, L., West, B. O., Woolfrey, J. L. J. Sol-Gel Sci. Technol. 2000, 19, 315; Peshev, P.; Slavova, V. Mater. Res. Bull. 1992, 27, 1269; Papet, P.; Le Bars, N.; Baumard, J. F.; Lecomte, A.; Dauger, A. J. Mater. Sci. 1989, 24, 3850), triethanolamine (Okubo, T.; Takahashi, T; Sadakata, M.; Nagamoto, H. J. Membr. Sci. 1996, 118, 151), and 1,5-diaminopentane (Percy, M. J., Barlett, J. R., Spiccia, L., West, B. O., Woolfrey, J. L. J. Sol-Gel Sci. Technol. 2000, 19, 315) have been used as chelating reagents for zirconia sol-gel reactions. In general, chelation occurs when the added reagent replaces one or more alkoxy groups forming a strong bond. The formation of this bond reduces the hydrolysis rate by decreasing the number of available alkoxy groups (Bradley, D. C.; Mehrotra, R. C.; Gaur, D. P. Metal Alkoxide, Academic Press, London, 1978, p. 162).
In experimentation series 3, the hydrolysis rate of zirconium butoxide was controlled by using glacial acetic acid (Wu, J. C. S.; Cheng, L. C. J. Membr. Sci. 2000, 167, 253) as a chelating agent as well as a source of water released slowly through the esterification with 1-butanol (Guizard, C.; Cygankiewicz, N.; Larbot, A.; Cot, L. J. Non-Cryst. Solids 1986, 82, 86; Larbot, A.; Alary, J. A.; Guizard, C.; Cot, L.; Gillot, J. J. Non-Cryst. Solids 1988, 104, 161). Two silanol-terminated poly (dimethyldiphenylsiloxane) copolymers (with 2-3% and 14-18% diphenyl-containing blocks) were used as sol-gel-active organic components to be chemically incorporated in the sol-gel network through polycondensation reactions with the zirconium butoxide precursor and its hydrolysis products. An IR spectrum of the pure co-polymers (the one with 2-3% phenyl-containing block) is presented in
This advantageous chemical incorporation of an organic component into the sol-gel network is responsible for the formation of an organic-inorganic hybrid material system that can be conveniently used for in situ creation of surface coating on a substrate like the inner walls of a fused silica capillary. Besides, the organic groups help to reduce the shrinkage and cracking of the sol-gel coating (Thouless, M.D.; Olsson, E.; Gupta, A. Acta Metall. Mater., 1992, 40, 1287; Paterson, M. J. McCulloch, D. G.; Paterson, P. J. K.; Ben-Nissan, B. Thin Solid Film, 1997, 311, 196). Furthermore, the sol-gel process can be used to control the porosity and thickness of the coating and to improve its mechanical properties (Sorek, Y.; Reisfeld, R.; Weiss, A.M. Chem. Phys. Lett. 1995, 244, 371). Poly(methylhydrosiloxane) and 1,1,1,3,3,3-hexamethyldisilazane served as deactivation reagents to perform chemical derivatization of the strongly adsorptive residual hydroxyl groups on the resulting sol-gel material. These reactions minimized the strong adsorptive interactions between polar solutes and the sol-gel sorbent that may lead to sample loss, peak tailing, sample carry-over and other deleterious effects. In the presented method for the preparation of the sol-gel zirconia coated microextraction capillary, the deactivation reactions were designed to take place mainly during thermal conditioning of the capillary following the sol-gel coating procedure.
Hydrolytic polycondensation reactions for sol-gel-active reagents are well established in sol-gel chemistry (Puccetti, G.; Leblanc, R. M. J. Phys. Chem. B 1998, 102, 9002; Golubko, N. V.; Yanovskaya, M. I.; Romm, I. P.; Ozerin, A. N. J. Sol-Gel Sci. Technol 2001, 20, 245; Brinker, C.; Scherer, G. Sol-gel Science, The Physics and Chemistry of Sol-Gel Processing, Academic Press, San Diego, USA, 1990; Dire, S.; Campostrini, R.; Ceccato, R. Chem. Mater., 1998, 10, 268) and constitute the fundamental mechanism in sol-gel synthesis. The condensation between sol-gel-active zirconia and silicon compounds is also well documented (Mori, T.; Yamamura, H.; Kobayashi, H.; Mitamura, T. J. Am. Ceram. Soc. 1992, 75, 2420; Toba, M.; Mizukami, F.; Niwa, S. I.; Sano, T.; Maeda, K.; Annila, A.; Komppa, V. J. Mol. Catal., 1994, 94, 85; Zhan, Z.; Zeng, H. C. J. Non-Cryst. Solids, 1999, 243, 26). According to published literature data (Dang, Z.; Anderson, B. G., Amenomiya, Y.; Morrow, B. A. J. Phys. Chem. 1995, 99, 14437; Guermeur, C.; Lambard, J.; Gerard, J.-F.; Sanchez, C. J. Mater. Chem. 1999, 9, 769), the characteristic IR band for Zr—O—Si bonds is located in the vicinity of 945-980 cm−1.
Metal-bound hydroxyl groups on the created sol-gel coating represent strong adsorptive sites for polar solutes. In the context of analytical microextraction or separation, the presence of such groups is undesirable and may lead to a number of deleterious effects including sample loss, reproducibility problems, sample carryover problems, and peak distortion and tailing. Therefore, appropriate measures need to be taken to deactivate these adsorptive sites. This may be accomplished by chemically reacting the hydroxyl groups with suitable derivatization reagents. Like silica-based sol-gel coatings, the surface hydroxyl groups of sol-gel zirconia coating can be derivatized using reactive silicon hydride compounds such as alkyl hydrosilanes (Fadeev, A. Y.; Helmy, R.; Marcinko, S. Langmuir 2002, 18, 7521; Marcinko, S.; Helmy, R.; Fadeev, A. Y. Langmuir 2003, 19, 2752) and hexamethyldisilazane (Wu, N. L.; Wang, S. Y.; Rusakova, I. A. Science 1999, 285, 1375). A mixture of polymethylhydrosiloxane and hexamethyldisilazane was utilized for this purpose: the underlying chemical reactions are schematically represented in Scheme 21 (A and B) illustrate the chemical structure of the sol-gel zirconia surface coating before and after deactivation, respectively.
One of the most important undertakings in CME is the creation of a stable, surface-bonded sorbent coating on the inner walls of a fused silica capillary.
Sol-gel zirconia-PDMDPS-coated capillaries allowed the extraction of analytes belonging to various chemical classes. Experimental data highlighting CME-GC analysis of polycyclic aromatic hydrocarbons using a sol-gel zirconia-PDMDPS coated capillary is shown in
CME-GC experiments were performed on an aqueous sample with low ppb level analyte concentrations. Experimental data presented in Table IV shows that CME-GC with a sol-gel zirconia-PDMDPS coating provides excellent run to-run repeatability in solute peak areas (3-7%) and the used sol-gel GC column provided excellent repeatability in retention times (less than 0.2%). It should be pointed out that the column used for GC analyses was also prepared in-house using a sol-gel method described in a previous publication (Wang, D.; Chong, S. L.; Malik, A. Anal. Chem. 1997, 69, 4566).
The reproducibility of the newly developed method for the preparation of sol-gel hybrid organic-inorganic zirconia coated capillaries was evaluated by preparing three sol-gel zirconia PDMDPS-coated capillaries in accordance with the methods of the present invention and following their performance in CME-GC analysis of different classes of analytes extracted from aqueous samples. The GC peak area obtained for an extracted analyte was used as the criterion for capillary-to-capillary reproducibility, which ultimately characterizes the capillary preparation method reproducibility. The results are presented in Table V. For each analyte, four replicate extractions were made on each capillary and the mean of the four measured peak areas was used in Table V for the purpose of capillary-to-capillary reproducibility. The presented data show that the capillary-to-capillary reproducibility is characterized by an RSD value of less than 5.5% for all three classes of compounds used for this evaluation. For a sample preparation method, a less than 5.5% R.S.D. is indicative of excellent reproducibility.
In capillary microextraction technique, the amount of analyte extracted into the sorbent coating depends not only on the polarity and thickness of the coated phase, but also on the extraction time.
Sol-gel zirconia-PDMDPS coatings showed high pH stability and retained excellent performance after rinsing with 0.1M NaOH (pH 13) for 24 h. Chromatograms in
As is evident from
For comparison, the same experiment was conducted using a 10 cm piece of a conventionally coated commercial PDMDPS-based GC column as the microextraction capillary. The results are shown in
These data suggest that the created hybrid sol-gel zirconia-based coatings have significant pH stability advantage over conventional silica-based coatings, and that such coatings have the potential to extend the applicability of capillary microextraction and related techniques to highly basic samples, or analytes that require highly basic condition for the extraction and/or analysis.
Sol-gel zirconia-based hybrid organic-inorganic sorbent coating was developed for use in microextraction. Principles of sol-gel chemistry was employed to chemically bind a hydroxy-terminated silicone polymer (polydimethyldiphenylsiloxane) to a sol-gel zirconia network in the course of its evolution from highly reactive alkoxide precursor (zirconium tetrabutoxide) undergoing controlled hydrolytic polycondensation reactions. For the first time, sol-gel zirconia-PDMDPS coating was employed in capillary microextraction. The newly developed sol-gel zirconia-PDMDPS coating demonstrated exceptional pH stability: its extraction characteristics remained practically unchanged after rinsing with a 0.1M solution of NaOH (pH 13) for 24 h. Solventless extraction of analytes was carried out simply by passing the aqueous sample through the sol-gel extraction capillary for approximately 30 min. The extracted analytes were efficiently transferred to a GC column via thermal desorption, and the desorbed analytes were separated by temperature programmed GC. Efficient CME-GC analyses of diverse range of solutes was achieved using sol-gel zirconia-PDMDPS capillaries. Parts per trillion (ppt) level detection limits were achieved for polar and non-polar analytes in CME-GC-FID experiments. Sol-gel zirconia-PDMDPS coated microextraction capillaries showed remarkable run-to-run repeatability (R.S.D.<0.27%) and produced peak area R.S.D. values in the range of 1.24-7.25%.
Throughout this application, various publications, including United States patents, are referenced by author and year and patents by number. Full citations for the publications are listed below. The disclosures of these publications and patents in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains.
The invention has been described in an illustrative manner, and it is to be understood that the terminology which has been used is intended to be in the nature of words of description rather than of limitation.
Obviously, many modifications and variations of the present invention are possible in light of the above teachings. It is, therefore, to be understood that within the scope of the appended claims, the invention can be practiced otherwise than as specifically described.
*Experimental conditions for capillary microextraction and GC analysis are same as in FIGS. 12 (PAHs), 13 (Aldehydes), and 14 (Ketones).
**Detection limits were calculated for a signal-to-noise ratio (S/N) of 3 using the data presented in FIGS. 12 (PAHs), 13 (Aldehydes), and 14 (Ketones).
* Experimental conditions for capillary microextraction and GC analysis are same as in FIGS. 16 (Phenols), 17 (Alcohols and Amines).
**Detection limits were calculated for a signal-to-noise ratio (S/N) of 3 using the data presented in FIGS. 16 (Phenols), 17 (Alcohols and Amines).
This application is a continuation-in-part of co-pending U.S. Ser. No. 10/710,212, filed Jun. 25, 2004, which is a divisional of U.S. Ser. No. 10/001,489, filed on Oct. 23, 2001, now U.S. Pat. No. 6,783,680, which claims priority from U.S. Ser. No. 60/242,534, filed Oct. 23, 2000, which are all incorporated herein by reference in their entirety including any figures, tables, or drawings.
| Number | Date | Country | |
|---|---|---|---|
| 60242534 | Oct 2000 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 10001489 | Oct 2001 | US |
| Child | 10710212 | Jun 2004 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 10710212 | Jun 2004 | US |
| Child | 11247445 | Oct 2005 | US |
