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[ 22 ] L. Mathiasson, G. Nilve, B. Ulen, Int. J. Environ. Anal. Chem. 45 ( 1991 ) 117. [ 23 ] Y. Shen, L. Mathiasson, J.A. Jonsson, J. Microcol. Sep. 10 ( 1998 ) 107. [ 24 ] B. Bernhardsson, E. Martins, G. Johansson, Anal. Chim. Acta 167 ( 1985 ) 111. Jan Ake Jonsson is docent and senior lecturer at the Department of Analytical Chemistry, Lund University, Lund, Sweden. He is a titular member of the IUPAC Commission of Separation Methods in Analytical Chemistry, and his main research interest are the fundamentals of chromatography and modern techniques for sample preparation, especially membrane methods in biological and environmental analysis. Lennart Mathiasson is docent and senior lecturer at the Department of Analytical Chemistry, Lund University, Lund, Sweden. His main research interests are the development of modern extraction techniques, especially SFE and microwave-assisted extraction, as well as membrane methods and their applications to environmental, food and occupational hygiene problems.

[ 11 ] M.D. Luque de Castro, I. Papaefstathiou, Trends Anal. Chem. 17 ( 1998 ) 41. [ 12 ] J.A. Jonsson and L. Mathiasson, Trends Anal. Chem. 18 ( 1999 ) 325. [ 13 ] B. Lindegard, H. Bjork, J.A. Jonsson, L. Mathiasson, A.M. Olsson, Anal. Chem. 66 ( 1994 ) 4490. [ 14 ] E. Thordarson, S. Palmarsdottir, L. Mathiasson, J.A. Jonsson, Anal. Chem. 68 ( 1996 ) 2559. [ 15 ] J.A. Jonsson, P. Lovkvist, G. Audunsson, G. Nilve, Anal. Chim. Acta 227 ( 1993 ) 9. [ 16 ] L. Chimuka, N. Megersa, J. Norberg, L. Mathiasson, J.A. Jonsson, Anal. Chem. 70 ( 1998 ) 3906. [ 17 ] M. Knutsson, J. Lundh, L. Mathiasson, J.A. Jonsson, P. Sundin, Anal. Lett. 29 ( 1996 ) 1619. [ 18 ] T. Miliotis, M. Knutsson, J.A. Jonsson, L. Mathiasson, Int. J. Environ. Anal. Chem. 64 ( 1996 ) 35. [ 19 ] Y. Shen, L. Gronberg, J.A. Jonsson, Anal. Chim. Acta 292 ( 1994 ) 31. [ 20 ] P. Wieczorek, J.A. Jonsson, L. Mathiasson, Anal. Chim. Acta 337 ( 1997 ) 183. [ 21 ] M. Knutsson, G. Nilve, L. Mathiasson, J.A. Jonsson, J. Agric. Food Chem. 40 ( 1992 ) 2413.

Liquid membrane extraction in analytical sample preparation

II. Applications
Jan Ake Jo nsson*, Lennart Mathiasson
Department of Analytical Chemistry, Lund University, P.O. Box 124, S-221 00 Lund, Sweden Membrane-based extraction techniques offer efcient alternatives to classical sample preparation techniques. In this review a number of examples from the elds of environmental and biomedical analysis are discussed. High selectivity and enrichment factors, as well as the possibility of automated interfacing to chromatographic and other analytical instruments, are shown for quantitative analysis. z1999 Elsevier Science B.V. All rights reserved.
Keywords: Membrane-based extraction techniques; Liquid^liquid extraction; Supported liquid extraction

1. Introduction
In our companion paper [ 1 ], we described the basic principles and theory for two membrane-based extraction techniques that can be applied efciently to sample pre-treatment before analysis by chromatographic or other methods. The techniques described are supported liquid membrane (SLM ) extraction [ 2,3 ] and microporous membrane liquid^liquid extraction (MMLLE ) [ 4 ]. SLM is an aqueous^organic^aqueous extraction technique, which is mainly applicable to polar compounds such as organic acids or bases, charged compounds, and metal ions. MMLLE, which involves an aqueous^organic extraction, is more suited for non-polar organic compounds. Both of these complementary techniques are characterized by high selectivity, ease of automation, and high enrichment ( especially in the case of SLM ), which make them useful tools for pre-treatment of complex
1999 Elsevier Science B.V. All rights reserved.

*Corresponding author.
0165-9936/99/$ ^ see front matter PII: S 0 1 6 5 - 9 9 3 6 ( 9 9 ) 0 0 1 0 3 - X

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is very efcient. However, the peaks observed after the extraction from blood plasma are lower, reecting the protein binding of the drugs; section 2.7 in our companion paper [ 1 ] gives a discussion of this. Similar gures showing a similar high degree of selectivity have been published for other drug analysis applications using SLM^gas chromatography ( GC ) [ 4 ], MMLLE^GC [ 6 ] and capillary electrophoresis ( CE )^SLM [ 7 ], all referring to blood plasma samples; also Section 3.3, below. A particularly `dirty' matrix is manure. In an early application work [ 8 ], suspensions of manure from animals ( swine, poultry and cows ) were extracted with SLM ( after ltration and centrifugation ). The extract was derivatized and nally analyzed for carboxylic acids by capillary GC. In the resulting chromatograms, the only peaks appearing originated from the analytes and from the derivatization reagents, showing a very selective extraction and permitting a proper quantication.
2.2. SPE^SLM comparison
Fig. 1. ( a ) Chromatograms of Amperozide ( I ), its metabolite ( II ), and homologue ( III ) with the subsequent blank after enrichment from blood plasma. ( b ) The corresponding chromatograms after enrichment from an aqueous buffer solution. Concentrations 4 Wg / ml of I and II, 8 Wg / ml of III. Reprinted from Lindegard et al. [ 5 ] with permission. Copyright 1994 American Chemical Society.

samples ( e.g., biological samples such as blood plasma or urine ) or for organic or inorganic trace analysis in natural waters. The examples we give below are chosen to illustrate the potential of these techniques with regard to selectivity, enrichment and automation.

For some classes of analytes, a direct comparison can be made between samples extracted with solidphase extraction (SPE ) according to established practice, and SLM extraction. In Fig. 2, an example is shown, of the determination of certain triazine herbicides in natural water [ 9 ]. It is seen that SLM provides a considerably cleaner extract, leading to more reliable identication and lower detection limits than SPE. However, the SLM procedure is slower, as applied in the cited work. A similar example concerning the determination of sulfonylurea herbicides in natural waters has been presented by Nilve et al. [ 10 ].
2.3. Selectivity tuning

2. Selectivity
2.1. Biomedical applications

The extraction of trace analytes from biological samples constitutes a challenge of great importance for the pharmaceutical industry and medicine. The SLM technique can provide the necessary selectivity. Fig. 1 shows liquid chromatography ( LC ) chromatograms of some basic drugs extracted with SLM both from water solutions and from blood plasma [ 5 ]. These chromatograms cannot be distinguished in practice in terms of disturbing peaks or baseline appearance, so the removal of the blood plasma matrix

By the use of various reagents usually added to the membrane phase, the mass-transfer rates of the different compounds, and thus the selectivity, can be greatly inuenced. This can improve the extraction efciency and permit extraction of classes of compounds other than simple acids and bases.
2.3.1. Improvement of mass transfer for polar analytes The extraction of small carboxylic acids should be straightforward, using a system similar to that described for bases in [ 1 ], but with a reversed pH gradient, i.e., with an acidied sample and a basic acceptor. However, it was found that some of these

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acids were extracted less efciently. By the addition of the well-known hydrogen-bonding reagent tri-octylphosphine oxide (TOPO ) this situation was improved [ 11 ]. In Fig. 3 it is seen that the extraction efciency of the most polar acids ( formic and lactic acids ) is improved considerably by the addition of TOPO, while the extraction of the less polar butanoic acid, which was satisfactory also without TOPO, is less inuenced.
2.3.2. Extraction of amino acids Extraction of amino acids is complicated by their zwitterionic properties. Amino acids are charged at all pH values and thus are not directly extractable. It is therefore necessary either to derivatize one of the functional groups [ 12 ] or to use ion-pairing reagents. One possibility, utilizing the second approach, is to use the

Fig. 3. Inuence of TOPO content in di-n-hexyl ether on extraction efciency E. After Shen et al. [ 11 ], with permission from Elsevier Science B.V.

extractant di-2-ethylhexylphosphoric acid ( DEHPA ), dissolved in the membrane liquid ( tri-2-ethylhexyl phosphate ) [ 13 ]. Under moderately acidic conditions, the amino acids ion-pair with this reagent and can be transported to the stagnant, more acidic acceptor ( see Fig. 4A ). This is an example of counter-ion transport extraction, whose driving force is provided by the gradient of protons moving in the direction opposite to the analytes. Another possibility [ 14 ] is to use a quaternary ammonium reagent, in this case methyltrioctylammonium chloride ( Aliquat 336 ), to extract amino acids from a basic donor into a neutral acceptor containing a high concentration of chloride ions ( see Fig. 4B ), providing the counter-ion transport gradient. In both these cases, satisfactory extraction efciencies and enrichment factors were obtained. There are also other possibilities, utilizing complexation equilibria, for the SLM extraction of amino acids ^ for example, involving crown ethers [ 15,16 ] and platinum complexes [ 17 ].
2.3.3. Extraction and speciation of metals A rich literature exists on the extraction of metal ions with LLE and SLM. Many of the reagents described can be employed for sample enrichment and pre-treatment in trace analysis of metals in environmental and biological matrices. Using the wellknown reagent 8-hydroxyquinoline, added to the sample in the donor phase, and a stagnant acceptor solution of DTPA ( diethylenetriaminepentaacetic acid ), extraction efciencies up to 70% were obtained for Cu [ 18 ]. A more versatile extraction system involved extraction of metal ions, as thiocyanate complexes, with a membrane containing Aliquat 336, into an acceptor with DTPA. Atomic absorption spectropho-

Fig. 2. Chromatograms ( LC^UV ) of methoxy-s-triazine herbicides. ( A ) SPE extraction of spiked river water ( 1.0 Wg / l of each analyte in 1 l ). ( B ) SLM extraction of spiked river water ( 0.5 Wg / l of each analyte ). Peak designations: 1, Simetone; 2, Atratone; 3, Secbumetone; 4, Terbumetone. For more details see [ 9 ]. Reproduced with permission from Elsevier Science B.V.

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both in combination with AAS. The possibility of using ion-pair chromatography combined with SLM extraction for metal determination was presented recently [ 21 ]. SLM extraction of Pb in urine, with detection by potentiometric stripping analysis (PSA ), was successfully realized [ 22 ]. This approach has potential for on-site eld measurements, and an instrument based on this principle would be fairly simple. Parthasarathy and Bufe [ 23 ] used the SLM technique with a crown ether and laurie acid in the membrane phase for the extraction and speciation of Cu in lake water. Their conclusion is that the SLM selectively extracts free Cu2 ions in the water. In another recent application regarding the speciation of chromium [ 24 ], two SLM units were serially connected in an automated system, the rst one extracting Cr( III ) ( i.e., Cr3 ) using the DEHPA system, and the second one extracting Cr(VI ) ( e.g., Cr2 O23 ) by ion-pairing with Aliquat 336. Using this 7 process, it was possible to determine chromium in its different oxidation states separately in natural water from an abandoned tannery site.
2.3.4. Articial receptors as extractants By incorporating specially designed extractants in the membrane phase, a very high degree of selectivity can be obtained. Preliminary ( unpublished ) experi-

Fig. 4. ( A ) Mechanism of transport of amino acid ( A ) across a liquid membrane containing DEHPA ( HR ) as extractant. From Wieczorek et al. [ 13 ], with permission from Elsevier Science B.V. ( B ) Mechanism of transport of amino acid ( A ) across a liquid membrane containing Aliquat 336 ( C ) as extractant. From Dzygiel et al. [ 14 ], with permission from Marcel Dekker, Inc.

tometry ( AAS ) was used for the analysis of the extracts. This ion-pairing system worked satisfactorily for Cu, Cd, Co, Ni and Zn [ 18 ]. Concentration enrichment factors of up to 500 times were obtained, but the limit of detection was set by the purity of the reagents, rather than the enrichment levels. The DEHPA reagent ( see Section 2.3.2 and Fig. 4A ) can also be used for trace enrichment of heavy metals. This was applied to the determination of Cu, Cd and Pb in river water [ 19 ] and of Pb in urine [ 20 ],

Fig. 5. Extraction efciency of barbiturates versus receptor concentration in dioctyl phthalate. F, Phenobarbital; 8, diallylbarbital; R, hexobarbital.

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ments have been performed with SLM extraction of barbiturates using the so-called Hamilton receptor [ 25 ]. This is a molecule that `ts' snugly around the barbiturate structure and has been shown to increase selectivity in LLE [ 26 ]. In Fig. 5 it is seen that the extraction efciency of phenobarbital and diallylbarbital is greatly improved by addition of the receptor. Hexobarbital, which has a methyl group hindering the binding to the receptor, is practically uninuenced. In fact, the receptor used was not the original molecule as described by Hamilton, but an analogue with alkane chains in order to reduce the water solubility. Other types of synthetic extraction reagents have also been described. Of special interest is the borate reagent described by Smith [ 27 ] for extraction of sugars and catecholamines. Preparative SLM extractions which are selective for fructose over glucose have been demonstrated [ 28 ].
2.3.5. Immuno-SLM A promising idea for increasing the selectivity of SLM extraction is to employ soluble antibodies in the acceptor liquid. Thus, compounds that are extracted through the organic membrane phase are trapped very selectively in the acceptor as antigen^ antibody complexes, that could be transported to a connected on-line ow-immunoassay system for detection. Preliminary experiments are being conducted with this technique's application to the extraction of triazine herbicides and nitrophenols.

enrichment factors up to 3000 were obtained for lead by Parthasarathy and co-workers [ 30 ]. Aniline was enriched up to 2000 times from natural water [ 29,31 ], and various alkyl-s-triazine herbicides about 80 times from natural water [ 32 ].
3.2. Time-integrating sampling

A true time-integrating sampling was achieved by SLM extraction. In co-operation with the Swedish Agricultural University, phenoxy acid herbicides were sampled from a brook in an agricultural area during the spring [ 33,34 ]. During this time, herbicides are applied to the growing crops at different time intervals. This together with occasional rains led to a large variability of the herbicide concentration in the brook, making conventional grab-sampling unreliable for determination of the total run-off of herbicides from the elds. It was shown that using SLM extraction, a true time-integrating sampling was obtained, leading to reliable measurements of mean values over 24 h. It is possible to extend such sampling periods to longer times, e.g., a week, but the extraction system has to be carefully designed ( with low extraction efciencies and efcient analyte trapping in the acceptor ) in order not to exceed the theoretical maximum enrichment factor, which would lead to erroneous results.
3.3. Enrichment in biomedical analysis

3. Enrichment
3.1. Environmental applications

In most of the biomedical applications mentioned in Section 2.1, enrichment factors of 30^70 times have been obtained. The available volume of a sample ( e.g.,

In many cases, it is possible to design systems which principally permit very large concentration-enrichment factors. However, the price to be paid is a longer time for the extraction. As discussed in our companion paper [ 1 ], it is possible to estimate theoretically the maximum enrichment factor attainable, and this procedure has been veried experimentally [ 29 ]. As briey mentioned above, it has been routinely possible to achieve over 100 times enrichment of heavy metal ions, as well as various organic pollutants, for the purpose of determining these compounds at trace levels ( 6 Wg / l ), mainly in environmental samples. For example, Pb was enriched from urine 200 times [ 20 ], and several heavy metals were enriched 200 times from natural waters [ 19 ]. Using a hollowber geometry and crown ethers in the membrane,

Fig. 6. Schematic diagram of ow system for SLM extraction of chlorinated phenols. For a description, see the text. From Knutsson et al. [ 36 ], with permission from Vieweg Publishing.

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Fig. 7. Experimental system for SLM^LC determination of basic drugs in blood plasma. For a description, see the text. Reprinted from Lindegard et al. [ 5 ] with permission. Copyright 1994 American Chemical Society.

blood plasma ) is limited, and these systems are usually designed so that the entire extract from a ca. 1 ml sample results in one chromatographic run, maximizing the analyte signal ( see Section 4.1 ). Especially worth mentioning is an SLM^LC^CE system [ 35 ] where the SLM extract of a drug was rst transferred to microcolumn LC, and the appropriate eluting analyte plug then transferred to a CE capillary, nally leading to a total concentration enrichment of about 40 000 times and a detection limit of about 0.15 nM in the blood plasma ( with conventional UV detection through the capillary ). Even at these concentrations, clean blanks were obtained and no matrix peaks interfered with the detection of the analyte.

4. Interfacing
4.1. Liquid chromatography

Membrane extraction techniques are well suited to automated interfacing with various chromatographic instruments, most straightforwardly with liquid chromatography. In several papers we have shown ow systems built up around peristaltic pumps and pneumatic valves, and controlled by electronic timers, integrators or computer systems. A typical system is shown in Fig. 6. It was set up for SLM extraction of chlorinated phenols from natural waters [ 36 ] and comprises two peristaltic pumps ( 1 ), four pneumatic valves ( 2,5,6,8 ), and a membrane unit ( 4 ). The sample is acidied and pumped through the donor channel. By means of a valve ( 2 ) the sample can be substituted for reagent water for washing the donor channel between the samples. The alkaline

acceptor is kept stagnant during extraction and by switching a valve ( 5 ), it can be transported further on, neutralized and transferred to a precolumn ( 9 ) where the analytes are adsorbed and focused. By switching the injection valve ( 8 ) the analytes are transferred to the analytical column. There are provisions for rinsing the precolumn with acid between runs. This type of system is mainly used for the extraction of relatively large amounts of water with large membrane units ( channel volumes 1 ml ). The integral precolumn ensures that all the extracted analytes from one sample are transferred to the column to be analyzed in one chromatographic run. Essentially the same set-up has been used for other applications [ 10,37 ]. To circumvent the disadvantage that the membrane unit is subject to a rather high pressure during the transfer of the sample to the precolumn, a similar system with an intermediate storage loop has been constructed [ 31 ]. Another variation is to employ a membrane unit that is sufciently small, so that the entire extract can be contained in an injection loop and thus injected directly into the liquid chromatograph without a precolumn [ 38,39 ]. In some cases [ 32,40 ], the extract from large membrane units is collected manually and injected into the HPLC using an autosampler. This leads to lower overall sensitivities but permits duplicate chromatographic analyses. In order to handle biological samples of about 1 ml, the peristaltic pump approach is not suitable. A fully automatic system, built around an `intelligent sample processor' Gilson 231 ( Gilson SA, Villiers-le-Bel, France ) ^ the basis of the known ASPEC and ASTED instruments ^ was constructed for the determination of basic drugs in blood plasma [ 5 ]. Fig. 7 shows a schematic diagram of that system. The samples to be extracted are contained in vials ( 1 ) in an autosampler rack. Immediately before extraction, alkaline buffer is added and the alkaline sample is transferred by means of the syringe pump ( 3 ) and a robotic needle through the donor channel ( 4 ) in the membrane unit ( 6 ). The channel volume is 10 Wl in this case. After the extraction is completed, the contents of the acceptor channel ( 7 ) are pushed into an injection loop ( 8 ), and subsequently injected into the chromatographic column ( 9 ). By synchronizing the operations of the sample preparation system with the chromatographic computer system, one sample is extracted during the chromatographic run of the previous sample. Thus the sample throughput is determined by the length of the chromatogram, in this case ca. 15 min ( cf. Fig. 1 ).

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Fig. 8. Instrumental system for blood plasma analysis using MMLLE^GC. Reprinted from Shen et al. [ 4 ] with permission. Copyright 1998 American Chemical Society.

4.2. Gas chromatography

ane, which was passed to a loop-type large volume ( 300 Wl ) GC injection device [ 46 ] and injected into a capillary column system, comprising a retention gap, a retaining precolumn and an analysis column. The entire system was completely automated, and handled three samples simultaneously: while one sample was being chromatographed, the next one was in the phaseswitching unit, and the third one was extracted. The system permitted a high degree of selectivity and sensitivity. However, even if the described SLM^GC system was operable, it was complex and difcult to operate. Therefore, the principle of MMLLE was applied, leading directly to an extract in hexane that could be injected directly into the GC via the large-volume injection system. In Fig. 8, the resulting apparatus is shown. This permitted a smooth integration of extraction and GC, and the analysis could be performed completely automatically, with a cycle time of 25 min per sample, again determined by the chromatographic run time. Thus, one blood plasma sample is extracted while the previous one is separated in the GC. The results obtained were similar to those obtained with the SLM^GC system described above, but the MMLLE^GC system was operated more easily and more rugged.
4.3. Capillary electrophoresis

The combination of SLM extraction and packedcolumn gas chromatography has mainly been made in two ways. In early work [ 41,42 ], a so-called LC^ GC interface (Varian ) was employed. This now obsolete device, essentially an autosampler syringe, was set up to inject a selected portion of the acceptor phase into the standard GC injection port. Also, a conventional six-port valve for direct injection of liquid was used [ 43,44 ]. In both these approaches, aqueous samples are injected into the GC, which makes them less suited for capillary column applications. To permit direct interfacing of SLM extraction and capillary column GC, a solvent change is required, followed by a large-volume injection into the column. With this principle, a SLM^GC system for drugs in blood plasma was constructed [ 6 ]. The extraction was made essentially as shown in Fig. 7 and the acceptor solution was transferred to a solid-phase precolumn, where the analytes were adsorbed. Subsequently, nitrogen was passed through the precolumn, thereby removing the water, using a principle described earlier by Brinkman and Vreuls [ 45 ]. Then the analytes, still adsorbed on the precolumn, were desorbed with hex-

In CE, there is a large demand for selective sample pre-treatment. The injected amounts are usually very small, necessitating enrichment of the analytes in many cases and, as the system generally is more sensitive to various matrix inuences, necessitating an efcient clean-up. SLM extraction has been used in connection with CE in several cases, mainly in offline modes [ 7,35,47 ]. A completely direct and automatic connection was not accomplished. However, a special, low-volume membrane unit utilizing a hollow-ber geometry with 1.3 Wl acceptor volume was developed [ 48 ]. After enrichment, this volume was transferred to the separation capillary, which was temporarily connected ( via its detector end ) to the SLM unit. After the transfer, the capillary was placed manually in the electrode vessel and a double stacking procedure was started [ 49 ], by which the analytes were concentrated in the injector end of the capillary, followed by the CE separation. Satisfactory clean-up and enrichment were demonstrated, to permit determination of a basic drug in blood plasma in concentrations of 4 nM.

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volume. This is equivalent to saying that the extraction efciency is constant with the sample volume. There is an upper limit for this constancy [ 29 ], as discussed in Section 3.1. Within this limit, it is possible to increase the enrichment factors, and thus reduce the detection limits by increasing the extraction time, and in some cases it is possible to reach very low detection limits. In environmental applications involving extraction of organic or inorganic pollutants, limits of detection well below Wg / l are routinely achieved. In some cases, the detection limit is set by the purity of the available reagents [ 18 ].
Fig. 9. Experimental system for metal enrichment. From Djane et al. [ 20 ], with permission from the Royal Society of Chemistry.

5.2. Extraction efciency

4.4. Atomic absorption spectrometry

For trace determinations of heavy metals, an interfacing of SLM and AAS is desired. This has been accomplished by pumping the extract from the acceptor to an empty vial in a fraction collector, from which the vials were transferred to the GFAAS autosampler. Alternatively, the contents could be introduced directly into a ame AAS or ICP nebulizer. Fig. 9 shows a typical system, which additionally employs peristaltic pumps and pneumatic valves in a way similar to that shown in Fig. 6. A completely automated system [ 50 ] comprising four SLM extraction units working in parallel to improve throughput was connected directly to the vials in the autosampler dispensing unit of a GFAAS. This principle is advantageous from the point of view of contamination, as it is possible to create a nearly closed system with a nitrogen overpressure.

5. Quantitative determinations
5.1. Calibration curves

In SLM extraction, it is usually not practical to work with 100% extraction efciency ( E ). As detailed in our companion paper [ 1 ], it is more time-efcient to work at lower E values. In most cases where quantitative determinations are made, E is between 20 and 80%. This is not to be confused with recovery, as E enters the calibration as a constant factor and, provided that it is constant under the selected conditions, does not inuence the accuracy. This situation can be compared with other techniques such as AAS ( where the nebulization efciency is about 10%), dialysis ( where often only a few percent of analyte is transferred ), and MS ( where the ionization efciency may be 6 1%, depending on the ionization technique ). There are examples where the values of E in different sample matrices may be different. In blood plasma, changes in ( the apparent ) E can be caused by protein binding [ 5,6 ]. In environmental matrices such as natural water, other components might inuence the extraction [ 19,36 ], while in other cases, no signicant difference between natural waters and reagent water is found [ 34 ]. In the determination of carboxylic acids in soil solutions [ 52 ], the recovery of eight acids, spiked to different soil solutions, and pure water, was in no case signicantly different from 100%.
5.3. Comparisons

In a number of cases, straight line calibration curves have been demonstrated for natural matrices, for both environmental [ 18,31,34,37,39,51 ] and biological [ 4^6 ] applications. This refers to conventional calibration curves ( extraction of sample with equal volumes spiked with different concentrations of analytes ). Additionally, it has been demonstrated that in many cases the extraction process is linear over time ( or volume ), so the concentration found in the extract increases proportionally with the extracted sample

For validation, comparisons with reference materials and other methods is valuable. The concentration of Pb in urine, as determined by SLM^AAS, was compared successfully to measurements by ICP [ 20 ]: three metals were determined in a riverine water reference material [ 19 ] with satisfactory agreement ( e.g., 32 5 vs. 28 4 ng / l of Cd ), and various phenol derivatives compared well with an Aquacheck standard sample ( unpublished ) at the 2^4 Wg / ml level.

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5.4. Stability of the liquid membrane

The need for a reasonable long-time stability of the liquid membrane extraction system necessitates some consideration. Obviously, the organic solvent must be selected to be insoluble in water and still sufciently non-viscous. This is easily accomplished with nonpolar liquids. The most commonly used liquid is nundecane, which forms membranes that can be used continuously for several weeks without physical and chemical changes. When there is a need for more polar membranes, di-n-hexyl ether or mixtures of this solvent with n-undecane are often used, and such membranes are practically as stable as pure n-undecane membranes. To achieve special selectivity, as described above ( see Section 2.3 ) various additives to the liquid membrane are required. This may compromise stability, and the use of additives which are as hydrophobic as possible ( maybe modied with alkyl chains, as mentioned above in Section 2.3.4 ) is advantageous. For practical use, an SLM preparation should be stable for at least one working day, as is the case in all the applications described above. The material of the liquid membrane support seems to inuence the stability somewhat, the most commonly used PTFE membranes being slightly better than polypropylene membranes. Also, the pore size inuences the membrane stability. The regeneration of the SLM is made in a few minutes by simply soaking the membrane support in the desired liquid, wiping, and reinstalling the SLM in the membrane holder. For the hollow-ber membranes, in situ regeneration has been shown to work well [ 48 ] and this should also be the case for at membranes.
5.5. Memory effects

acceptor contents after successive replacing the extract with clean acceptor buffer or, alternatively, by extracting a blank sample directly after a real sample. In some cases, negligible amounts were then found [ 5,18,31,32 ], while in other cases the memory effects demand a scheme for washing the membrane system between each sample [ 36,37 ]. A detailed study for carboxylic acid extraction has been undertaken [ 11 ].

6. Conclusions
SLM and MMLLE are procedures that can provide selective enrichment and clean-up in various sample types of environmental or biological origin and for various classes of analytes. Depending on the needs, the focus can be either on the enrichment aspect with enrichment factors of several hundred times or on the clean-up aspect with efcient removal of even difcult matrices such as blood plasma. The procedures can be utilized in quantitative determinations. Analytical procedures involving SLM or MMLLE extraction have to be validated with real samples, just as with any other approaches.

Acknowledgements
This work was nancially supported by the Swedish Natural Science Research Council (NFR ), the Swedish Environmental Protection Agency (SNV ), the Swedish Council for Forestry and Agricultural Research (SJFR ), the Swedish Institute, the Swedish International Development Co-operation Agency (SIDA ), the Crafoord Foundation and the European Community ( DG XII ). The companies Pharmacia AB, Astra Draco AB and Astra Pain Control AB have also contributed. A number of co-workers, including graduate and undergraduate students, have made important contributions and their diligent work is highly appreciated.

In all quantitative applications of SLM and MMLLE, memory effects must be considered, i.e., incomplete transfer of analyte from the membrane to the acceptor phase. The memory effect not only causes a reduction in the recovery but also leads to carry-over effects in sequential extractions in automated systems. The memory effect in SLM is largely determined by the composition of the membrane liquid in such way that a liquid giving a high partition coefcient might also give high memory effects. In MMLLE, the main cause seems to be kinetic, i.e., the rate of mass transfer from the liquid in the membrane pores to the bulk liquid in the acceptor channel. Practically, the memory effects are studied by analyzing several portions of

References
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