Anal. Chem. 2004, 76, 5974-5981
Implantable Flow-Through Capillary-Type Microdialyzers for Continuous in Situ Monitoring of Environmentally Relevant Parameters Manuel Miro´*,† and Wolfgang Frenzel‡
Department of Chemistry, Faculty of Sciences, University of the Balearic Islands, Carretera de Valldemossa, Km. 7.5, 07122-Palma de Mallorca, Illes Balears, Spain, and Institut fu¨r Technischen Umweltschutz, Technische Universita¨t Berlin, Strasse des 17. Juni 135, D-10623 Berlin, Germany
In this paper, a simple, flexible, and cost-effective flowthrough microdialyzer hyphenated with a miniaturized differential potentiometric detector is proposed for continuous diffusion-controlled sampling of analytes of environmental interest. The analytical performance of the dedicated configuration involving merely a single cellulose regenerated hollow fiber is critically compared with that of commercially available concentric probes commonly exploited for in vivo monitoring of the extracellular space in living tissues and that of large dialysis-based probes furnished with flat membranes. The outstanding feature of the capillary-type design is the ability of adapting the extraction fractions (EF) to the requirements of the assays and flow-through detectors by selection of appropriate membrane length/perfusion rate ratios. Passive sampling under steady-state conditions (EF ∼ 100%) has proven feasible for environmentally relevant ions, such as chloride, by perfusing a 3-cm-long capillary with water at a flow rate of 2.0 µL/min. Hence, there is no need for recalibration of the flow setup after implantation of the purpose-made probe. The effect of physical and chemical variables on the diffusive flux is discussed in detail for the various flow-through membrane separation devices assessed. Effective means to attain identical dialysate concentrations of target species under dynamic regime irrespective of the matrix ingredients are also presented. The dedicated microdialyzer features extreme tolerance to high molecular weight interfering matrix compounds (g5000 mg/L humic acid) at the 5% interference level, which makes it especially suited for the interference-free potentiometric determination of ionic species in environmental samples containing high levels of organic matter. The potentials of the membrane separation unit were assessed for continuous monitoring of chemical changes in the interstitial/pore water of organic soils via stimulusresponse strategies. Microdialysis is a sampling and cleanup technique conventionally used in the clinical field for measuring quasi-continuously the * Corresponding author. E-mail: hotmail.com. † University of the Balearic Islands. ‡ Technische Universita¨t Berlin.
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5974 Analytical Chemistry, Vol. 76, No. 19, October 1, 2004
concentration of administered therapeutic drugs and their metabolites (pharmacokinetic studies) as well as endogenous compounds, such as neurotransmitters, in the interstitial tissue fluid of organs (e.g., brain, liver, kidney) and in body fluids of living systems (e.g., plasma, bile).1-4 Since no fluid is removed from or introduced into the probed environment during microdialysis, minimum changes in the fluid balance and metabolic processes result. Moreover, only low molecular weight species present in the extracellular fluid can cross the dialysis membrane, thus stopping any further metabolism due to enzymatic actions in the dialysate. In addition, diffusates can be directly analyzed without any further sample pretreatment. As a consequence of its inherent advantages and potentials, microdialysis sampling has been progressively extended to other research areas, such as biotechnology, for the removal of cellular and particulate matter as well as proteins from fermentation broths.5,6 Scarce examples are, however, found in the literature regarding the exploitation of the virtues of microdialysis for environmental assays.7 The characterization of carbohydrates in storage septic tanks8 as well as the monitoring of metal uptake by plants9 should be highlighted as promising approaches, which denote the versatility and potentialities of microdialysis for in situ sampling in highly complex matrixes. Current research in areas of neuroscience, pharmacology, and biotechnology is being performed by resorting to short-size commercially available probes with loop-type or concentric configurations10-12 though dedicated flow-through coaxial-type (1) Dittrich, P.; Tomaselli, F.; Maier, A. Bioforum Int. 2003, 7, 42-43. (2) Davies, M. I.; Cooper, J. D.; Desmond, S. S.; Lunte, C. E.; Lunte, S. M. Adv. Drug Delivery Rev. 2000, 45, 169-188. (3) Robinson, T. E., Justice, J. B., Jr., Eds. Microdialysis in the Neurosciences; Elsevier: Amsterdam, 1991. (4) Weiss, D. J.; Lunte, C. E.; Lunte, S. M. Trends Anal. Chem. 2000, 19, 606616. (5) Torto, N.; Laurell, T.; Gorton, L.; Marko-Varga, G. Anal. Chim. Acta 1999, 379, 281-305. (6) van de Merbel, N. C.; Lingeman, H.; Brinkman, U. A. Th. J. Chromatogr., A 1996, 725, 13-27. (7) Torto, N.; Mwatseteza, J.; Laurell, T. LC-GC Eur. 2001, 14, 536-546. (8) Torto, N.; Lobelo, B.; Gorton, L. Analyst 2000, 125, 1379-1381. (9) Torto, N.; Mwatseteza, J.; Sawula, G. Anal. Chim. Acta 2002, 456, 253261. (10) Min, R. W.; Rajendran, V.; Larsson, N.; Gorton, L.; Planas, J.; Hahn-Hagerdal, B. Anal. Chim. Acta 1998, 366, 127-135. (11) Tseng, W.-C.; Sun, Y.-C.; Yang, M.-H.; Chen, T.-P.; Lin, T.-H.; Huang, Y.-L. J. Anal. At. Spectrom. 2003, 18, 38-43. 10.1021/ac049406a CCC: $27.50
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shunt probes for sampling flowing fluids have also been described.13,14 Small designs permit probing restricted areas, such as organs and plant tissues, and concomitantly prevent external disturbance of the sampling site. In addition to their high costs, the major shortcoming of the commercial built-in microdialyzers is the user’s inability to adapt the dimensions of the probe to the requirement of the assays and sensitivity of the analytical instrumentation. Despite the efforts conducted by several researchers to enhance the performance of concentric-type microdialysis probes by optimizing the internal geometry, i.e., the dimensions of the inner cannulas,15 typical dialysis measurements are performed under non-steady-state conditions.16 As a consequence, the system needs proper calibration to accurately calculate the extraction fractions and determine the concentration of the target analytes. Furthermore, the narrow range of ultraslow perfusing rates applicable results in large response times that may hinder the implementation of such probes in continuous monitoring schemes. In this paper, a novel, reliable, and cost-effective microdialysis unit involving a single hollow-fiber membrane of adjustable length is coupled to miniaturized tubular ion-selective electrodes (ISEs) placed in a Y-connector for on-line monitoring of ionic species in environmental and agricultural samples. This is the first application dealing with the implementation of a microdialysis probe into a soil body. It should be stressed that the direct applicability of conventional analytical methods for the determination of key parameters in soil extracts is often limited to samples containing low levels of organic matter.17 The presence of high molecular weight colored compounds causes improper performance of spectrophotometric procedures due to the increase of background absorbance and spectral overlapping. Sample pretreatments are equally demanded for multielemental determinations involving chromatographic separations to prevent adsorption of humic acid and humin species onto the analytical column.18 Despite the improved tolerance of potentiometric schemes to turbid and selfcolored samples, procedures involving silver-based ISEs are specially susceptible to interferences due to humic substances.19 On-line microdialysis sampling is, thus, exploited in this work as a powerful tool to isolate the analyte from high molecular weight matrix constituents without requiring time and laborious batch cleanup protocols.20 The potentials of the proposed configuration are assessed by the potentiometric determination of low concentrations of chloride in soils containing concomitantly high levels of organic matter. Chloride was selected as a model of analyte since it is a core parameter routinely monitored in soils for fertility assessment.17 The analytical performance of the dedicated hollow(12) Mayer, B. X.; Petsch, M.; Tschernko, E. M.; Muller, M. Electrophoresis 2003, 24, 1215-1220. (13) Gunaratna, C.; Lunte, S. M.; Zuo, H. Curr. Sep. 1994, 13, 80-83. (14) Fang, Q.; Shi, X.-T.; Sun, Y.-Q.; Fang, Z.-L. Anal. Chem. 1997, 69, 35703577. (15) Torto, N.; Mikeladze, E.; Gorton, L.; Cso ¨regi, E.; Laurell, T. Anal. Commun. 1999, 36, 171-174. (16) Martı´nez-Martı´nez, M. S.; Gutie´rrez-Hurtado, B.; Colino-Gandarillas, C. I.; Martı´nez-Lanao, J.; Sa´nchez-Navarro, A. Anal. Chim. Acta 2002, 459, 143150. (17) Hesse, P. R. A Textbook of Soil Chemical Analysis; Chemical Publishing: New York, 1972. (18) Smith, R. M. J. Chromatogr., A 2003, 1000, 3-27. (19) Sikora, F. J.; Stevenson, F. J. Soil Sci. Soc. Am. J. 1987, 51, 924-929. (20) Ferreira, A. M. R.; Rangel, A. O. S. S.; Lima, J. L. F. C. Commun. Soil Sci. Plant Anal. 1996, 27, 1437-1445.
Figure 1. Schematic representation of the microdialysis flow setup furnished with a miniaturized differential potentiometric detector for continuous monitoring of chloride.
fiber probe was compared with that of a commercial concentric microdialyzer and that of a large dialysis probe furnished with a flat membrane. The proposed design is especially suitable for implantation into soils without disturbance of the sampling site because of the low dimensions of the overall setup. A thorough discussion is also included in the bulk of the text with regard to the dependence of the phase-transfer rate of ionic species on the matrix composition, which has not been investigated in previous papers dealing with on-line microsampling of low molecular weight ions.9 EXPERIMENTAL SECTION Reagents and Solutions. All chemicals were of analyticalreagent grade and employed without further purification. Deionized water (specific resistivity >18 MΩ cm) was used for the preparation of the various solutions. The stock chloride solution of 10 g/L was prepared by dissolving 16.490 g of dried sodium chloride in 1 L of distilled water. Standard solutions within the 0.5-800 mg/L chloride range were prepared from the stock solution by serial dilution. Electrolyte solutions containing 0.25 or 0.50 mol/L potassium nitrate were supplied to the tubular ISEs to ensure stable and repeatable potentiometric measurements. A stock solution of 10 g/L of a natural humic acid (Ferak, Berlin), selected as a model of high molecular weight compounds, was used to assess the maximum tolerance of the microdialyzers to interfering organic species. Flow System and Miniaturized Differential Potentiometric Detector. The continuous-flow system arranged for the various microdialyzer designs is shown schematically in Figure 1. The flow manifold is constructed from PTFE tubing of 0.3-mm i.d. A conventional multichannel peristaltic pump (Type IPS-8, Ismatec) furnished with Tygon tubing was employed as a liquid driver at dispensing rates higher than 6.0 µL/min (especially suitable for the flat-type microdialyzer probe). For perfusing rates below 6.0 µL/min, a unidirectional microsyringe infusion pump (type 102, CMA/microdialysis, Axel Semrau GmbH&Co, Sprockho¨vel, Germany) equipped with two 1-mL syringes was used. One of the syringes contains the perfusion liquid for the dialyzer. The diffusate zone feeds the indicator electrode through a 4-cm-long PTFE tube. The second syringe filled with an electrolyte solution was connected directly to the reference electrode. The ancillary line feeding the indicator electrode with a potassium nitrate solution for ionic strength adjustment was connected to the flow network via a Y-piece. This auxiliary stream was, in all instances, supplied by the peristaltic pump. The miniaturized flow-through potentiometric detector was constructed from two identical silver tubes (10-mm length, 1.2Analytical Chemistry, Vol. 76, No. 19, October 1, 2004
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Figure 2. Magnified view of the various microdialysis probes assessed for environmental assays. (A) Commercial CMA/20 needle-type microdialyzer with height-adjustable holder. Prior to accommodation into real-life samples, the microdialysis probe is housed within the protective sieve shown in the photograph. (B) Dedicated capillary-type microdialyzer. The perforated cover also illustrated was designed to prevent mechanical stress on the membrane surface after insertion into the soil layer. For clarity reasons, only the lower half of the cover is shown together with the concentrically arranged hollow fiber (See text for further details).
mm i.d., 0.15-mm wall thickness, 99.99% Ag, Goodfellow) pressfitted into a KEL-F Y-connector (2 cm long, 2.5 cm wide, 1 cm thick) that was specially made to provide a minimum distance (
