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Developments in multiple headspace extraction
J. Biochem. Biophys. Methods 70 (2007) 229 – 233 www.elsevier.com/locate/jbbm
Review
Developments in multiple headspace extraction Minna Hakkarainen ⁎ Department of Fibre and Polymer Technology, School of Chemical Science and Engineering, Royal Institute of Technology, 100 44 Stockholm, Sweden Received 1 June 2006; accepted 23 August 2006
Abstract This paper reviews new developments in multiple headspace extraction (MHE), especially its combination with two miniaturized extraction techniques, solid-phase microextraction (SPME) and single-drop microextraction (SDME). The combination of the techniques broadens the applicability of SPME and SDME to quantitative determination of analytes in complex liquid and solid matrixes. These new methods offer several advantages over traditional liquid–solid, liquid–liquid and headspace extraction techniques. The potential applications include extraction of volatiles and semivolatiles from environmental and physiological samples and from different polymer products such as medical and biomedical materials, food packaging and building materials. The theoretical principals of the techniques are also briefly reviewed. © 2006 Elsevier B.V. All rights reserved. Keywords: Solid-phase microextraction; Single-drop microextraction; Multiple headspace extraction; Solid matrix
Contents 1. Introduction . . . . . . . . . . . . . . . . . . . 2. Multiple headspace extraction . . . . . . . . . 3. Multiple headspace solid-phase microextraction 4. Multiple headspace single-drop microextraction 5. Adsorption systems . . . . . . . . . . . . . . . 6. Conclusions. . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . .
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1. Introduction The extraction and quantitative analysis of volatiles and semivolatiles in solid or complex matrixes is an analytical challenge. Different liquid–solid extraction techniques, such as soxhlet extraction, microwave-assisted extraction (MAE), supercritical fluid extraction (SFE) and sonication are commonly used for the extraction of analytes from solid samples. However, these techniques are either expensive, time and labour intensive and/or use large amounts of toxic organic solvents. It is also difficult to perform accurate liquid–solid extraction in the case
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of volatile substances. If headspace techniques are used, partition of analytes from solid sample into the gaseous phase is often reduced, because of the interactions between the analytes and the matrix [1]. Internal and external calibration techniques rarely produce acceptable results due to the matrix effects, that cause considerable differences in the partition coefficients and release rates for different analytes. In 1977 Kolb and Pospisil presented a technique called discontinuous gas extraction [2]. The method was later renamed as multiple headspace extraction (MHE) [3]. The method removes the matrix effects and enables, thus, direct quantitative determination of analytes in solid matrixes by headspace techniques. This paper summarises new developments in multiple headspace extraction and especially its combination with solid-phase microextraction (SPME) and single-drop microextraction (SDME), which broadens the
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M. Hakkarainen / J. Biochem. Biophys. Methods 70 (2007) 229–233
applicability of SPME and SDME to quantitative determination of analytes in solid and complex aqueous matrixes. 2. Multiple headspace extraction The multiple headspace extraction is a stepwise headspace extraction for the quantitative analysis of volatiles in solid or complex liquid samples [2]. The method theoretically calculates the total amount of analyte in a solid sample after only a few successive extractions and makes it possible to quantitatively determine volatile analytes in solid matrixes. The theoretical principals of multiple headspace extraction (MHE) were presented in 1981 by Kolb and Pospisil [3]. When a portion of the headspace is removed in the first extraction, the equilibrium between the analyte in the condensed sample and the headspace is disturbed. As the sample is allowed to re-equilibrate, more analyte migrates from the condensed phase into the headspace. The concentrations in the two phases will now be smaller than during the first extraction, but the ratio between the analyte concentration in the two phases will be the same. The second extraction and analysis, thus, results in a smaller peak and by continuing this procedure it is possible to strip off all the volatiles from the sample. If carried out ad infinitum, the various peak areas are summed up to get the total peak area, which corresponds to the total amount of the analyte in the sample. The influence of sample matrix is thus eliminated by the exhaustive extraction. As the MHE procedure follows a logarithmic function it is not required that the extractions are carried out until all the analyte is removed from the sample matrix. Instead, the logarithms of the various area values from the consecutive analyses are plotted versus the number of analyses in a linear scale and the total area value is obtained by regression calculation from the areas obtained in only a few extraction steps [4]. The total amount of a volatile analyte in a sample can be calculated by summarizing all individual peak areas (Ai), where i is the number of the extraction. As this is a converging geometrical progression, the sum can be derived as: iX ¼l i¼1
Ai ¼
A1 1−e−q
ð1Þ
3. Multiple headspace solid-phase microextraction Under the last decade there have been great improvements in the area of organic trace analysis and several new miniaturized extraction techniques have arised. Solid phase microextraction (SPME) was for the first time presented in 1990 by Pawliszyn and Arthur [13]. It is based on a thin fused silica fibre coated with polymeric absorbent or adsorbent. For sampling the SPME fibre is immersed directly into aqueous samples or in the headspace over the liquid or solid sample matrix. Several fibre materials with different selectivities are commercially available. In several recent studies solid-phase microextraction was combined with multiple headspace extraction to enable quantitative determination of analytes in solid or complex aqueous matrixes. The theoretical principals of MHS–SPME under both equilibrium [14,15] and non-equilibrium [16] conditions were also presented. The main difference compared to conventional headspace extraction is the partitioning of the analytes between the fibre, sample matrix and headspace instead of just between sample matrix and headspace. Once the equilibrium is reached the mass of the analyte extracted by the fibre coating (mf) is related to the overall equilibrium of the analyte in the three phase system, and can be expressed as [17,18]: mf ¼
Kfs Vf dm0 Kfs Vf þ Khs Vh þ Vs
ð2Þ
Where the q′ value is equal to the slope of the linear regression line and ln A1 is given by the y-intercept. Applications for multiple headspace extraction include determination of volatiles in solid matrixes such as soil [5], polymers [3,6] and cellulose based packaging materials [7]. MHE has also been applied for determination of monomer solubilities in water [8], for automated determination of vapour-to-liquid partition co-
ð3Þ
Where m0 is the total mass of the analyte, Kfs and Khs are the fibre/sample and headspace/sample distribution constants and Vf, Vh and Vs are the volumes for fibre, headspace and sample. Under the non-equilibrium conditions the analyte mass extracted for a determined period of time (mf,t) can be defined as [16,19]: mf ;t ¼ mf ;e ð1−e−at Þ ¼
The sum of all peak areas can, thus, be calculated from two values: the peak area obtained in the first extraction, A1 and the exponent q′, which describes the exponential decline of the peak areas during the stepwise MHE procedure. A1 is a measured value and the exponent q′ is obtained from the linear regression analysis: ln Ai ¼ −qVði−1Þ þ ln A1
efficients [9,10] and for studying the process kinetics involving volatile species [11]. The amount of residual organic solvent in biodegradable microspheres was also determined by MHE to study the influence of preparation method on residual solvent content [12].
Kfs Vf dm0 ð1−e−at Þ Kfs Vf þ Khs Vh þ Vs
ð4Þ
Where mf,e is the mass extracted by the fibre when the equilibrium has been reached and a is a measure of how rapidly the partition equilibrium is reached. a can be obtained from the following expression [16,19]: a ¼ 2Am
kKfs Vf þ kKhs Vh þ kVs 2mkfh Vf Vs þ kKhs Vf Vh þ kVf Vs
ð5Þ
Where A is the surface area of SPME fibre, m the mass transfer coefficient of the analyte in the SPME phase, k the evaporation rate constant and Kfh the equilibrium partition constant for the analyte between the SPME fibre and headspace. Larger a value means faster partition equilibrium. For valid MHS–SPME the relationship between the peak area and the amount of analyte in the fibre coating must be linear over the
M. Hakkarainen / J. Biochem. Biophys. Methods 70 (2007) 229–233
studied range [15]. The volumes for sample, headspace and fibre and the distribution constants between the different phases must also remain constant. In the case of non-equilibrium MHS–SPME it is crucial that all the SPME parameter in each individual extraction are kept constant [16]. The Eqs. (1) and (2), derived for multiple headspace extraction, are applicable even for multiple headspace solid-phase microextraction. Solid-phase microextraction has rapidly become one of the most frequently used extraction techniques for different environmental analysis. It has e.g. in several studies been applied for extraction of contaminants from soil samples [20– 23]. A frequently used approach to enable quantitative determination of analytes in solid or complex matrixes is to combine SPME with another extraction technique. The analytes were, thus, first extracted from soil by for example solvent extraction, microwave assisted extraction or ultrasonic extraction [24–27]. These extracts were then further concentrated on the SPME fibre and quantified. The multiple headspace solidphase microextraction (MHS–SPME) technique could in many cases offer a more attractive alternative as it is both easily automated and eliminates the use of organic solvents. Recently MHS–SPME was successfully applied for determination of benzene, toluene, ethyl benzene and xylene isomer (BTEX) in contaminated soil [28]. The extractions were carried out from soil suspensions in water at 30 °C with carboxen-polydimethylsiloxane (CAR-PDMS) fibre. The obtained BTEX concentration values in certified soil samples were statistically equal to the reference ones proving that the MHS–SPME method removed the matrix effects. A direct quantitative MHS–SPME method has also been developed to monitor biogenic volatile organic compounds (BVOCs) released from a living leaf of Pelargonium hortorum in situ [29] and for determination of volatile compounds in antioxidant rosemary extracts [30]. In addition MHS–SPME has been applied for quantitative determination of volatile organic compounds in polymers [1,15], in cork stoppers [31] and in wine [32,33]. Determination of odour-causing volatile organic compounds in cork stoppers showed that the reproducibility of MHS–SPME was similar to soxhlet extraction, but the soxhlet extraction required a concentration step to obtain analyte concentrations over the detection limit [31]. Soxhlet extraction was also non-selective, took longer time, consumed large volumes of solvent and could not be used for the most volatile analytes. Compared to microwave assisted extraction, MHS–SPME offered higher sensitivity, solvent-free extraction, reduced labour-time and complete recovery of the analyte [1]. The MHS–SPME has, thus, several advantages over the liquid–solid extraction techniques. It also offers improvements to traditional multiple headspace extraction including increased sensitivity and selectivity. In addition SPME is better suited for the extraction of semivolatiles as several studies have shown that e.g. polymer additives that were not detected after traditional HS-sampling were easily extracted and detected by HS-SPME [34]. The SPME device is also easily portable enabling field sampling of environmental samples. A slightly different concept of multiple extraction, i.e. multiple solid-phase microextraction, has been presented by Koster
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Fig. 1. Headspace single-drop microextraction of analytes from solid sample matrix.
and de Jong [35]. This method was developed to reduce the extraction time and to increase the extraction yield and not for the quantitative determination of analytes. The SPME fibre was immersed directly into the buffer solution for non-equilibrium extraction. After the extraction the extracted analytes were statically desorbed in the desorption chamber, before a new extraction and desorption cycle. After the desired number of extractions, all the extracted analytes were injected into the LC system by one single injection. 4. Multiple headspace single-drop microextraction Traditional liquid–liquid (LLE) and liquid–solid (LSE) extractions require large amounts of highly purified solvent, and as a result of this, large amounts of hazardous waste are generated. Single-drop microextraction is a miniaturization of the traditional liquid–liquid extraction method, where the solvent to aqueous ratio is greatly reduced. The volatiles are extracted by a microdrop of non water-soluble organic solvent, which is immersed in an aqueous sample [36]. After extraction the drop is retracted into the syringe and the micro-drop is transferred to a GC for further analysis. Alternatively the micro syringe needle with the drop is suspended above the surface of sample phase to perform headspace single-drop microextraction (HS-SDME) [37,38]. HSSDME is an inexpensive and rapid technique, which uses practically no solvent and eliminates the possible memory effects, because new solvent drop is used every time. Immersion and headspace-SDME have successfully been applied for the extraction of volatiles from different liquid samples e.g. dialkyl phthalate in food simulants [39], organophosphorus insecticides in water [40], antifouling agents in water [41], amino acids in urine [42] and iodine pharmaceuticals [43]. MAE and HS-SDME were combined to enable quantitative determination of paeonol in traditional Chinese medicines [44]. Recently, the headspace-single drop microextraction technique was for the first time applied for the extraction of volatiles from solid samples [45]. Quantitative determination of volatiles became possible by combining the SDME and multiple headspace extraction techniques. Fig. 1 shows a schematic drawing over the HS-SDME of
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solid samples. Similar to the MHS-SPME, multiple headspace single-drop microextraction (MHS–SDME) system consists of three phases: sample matrix, headspace and micro-drop of organic solvent. After the equilibrium has been reached the mass of the analyte extracted by the micro-drop (md) can be calculated from Eq. (3), developed for three phase systems, if mf, Vf and Kfs in the equation are replaced by md, Vd (volume for micro-drop) and Kds (micro-drop/sample distribution constant), respectively [46]: md ¼
Kds Vd dm0 Kds Vd þ Khs Vh þ Vs
ð6Þ
Where m0 is the total mass of the analyte, Khs is the headspace/sample distribution constant and Vh and Vs are the volumes for headspace and sample. Similarly under the nonequilibrium conditions the analyte mass extracted for a determined period of time (md,t) can be defined as: md;t ¼ md;e ð1−e−at Þ ¼
Kds Vd dm0 ð1−e−at Þ Kds Vd þ Khs Vh þ Vs
ð7Þ
Where md,e is the mass extracted by the micro-drop when equilibrium has been reached and a is a measure of how rapidly partition equilibrium is reached. Soxhlet, microwave-assisted extraction, supercritical fluid extraction, headspace extraction and dissolution–precipitation techniques have earlier been evaluated and compared for determination the residual styrene content in polystyrene granules [47]. However, only the dissolution–precipitation method gave good results. Validation of the developed MHS–SDME method against the dissolution–precipitation method showed good agreement [45]. However, compared to the dissolution–precipitation method, the MHS–SDME method reduced labourtime and was practically solvent-free. 5. Adsorption systems Problems may arise in quantitation by MHE when both the analyte and the solid sample are polar. Adsorption of analytes is a well-known problem when extracting environmental pollutants from soil. It is also a problem during extraction of analytes from cardboard [7] and polar polymer samples [1]. These systems are referred to as adsorption systems, due to the adsorption of the analyte by the sample matrix. If the system is an adsorption system there will be no exponential decrease in peak area throughout the successive extractions. The system can be changed from an adsorption system into a partitioning system by adding a compound that has higher affinity towards the adsorbing sites in the matrix than the analyte of interest. Such a compound is called a displacer or modifier. The use of water as a displacer allowed quantitative determination of cyclohexanone in soil samples and increased the recovery from 4% to 99.4% [5]. BTEX (benzene, toluene, ethylbenzene and xylene isomers) in soil could also be analysed quantitatively by using MHS–SPME when water was added to the samples [28]. When water was added as a displacer, the amount of hexanal extracted from cardboard increased six times, clearly showing the higher affinity of the polar adsorption sites of the matrix towards water
[48]. The establishment of equilibrium between 2-cyclopentylcyclopentanone in polyamide-66 and the headspace required 12 h of incubation at 80 °C [49]. However, the addition of water as a displacer released the 2-cyclopentyl-cyclopentanone from polyamide-66 matrix and allowed rapid assessment of the analyte by MHS–SPME [1]. 6. Conclusions Combination of headspace solid-phase microextraction or headspace single-drop microextraction with multiple headspace extraction broadens the applicability of HS–SPME and HS– SDME to quantitative determination of volatiles in complex liquid and solid matrixes. MHS–SPME and MHS–SDME offer several advantages compared to both liquid–liquid and liquid– solid extraction techniques and traditional multiple headspace extraction. MHS–SPME is a solvent-free, easily automated, portable and sensitive technique. It is applicable for extraction of volatile and semivolatile compounds and the selectivity can be changed by changing the fibre material. MHS–SDME is a practically solvent-free, cheap and sensitive technique. It also eliminates possible memory effects as fresh solvent-drop is used for each extraction. The potential applications include extraction of volatiles and semivolatiles from e.g. environmental and physiological samples and from different polymer products such as medical and biomedical materials, food packaging and building materials. References [1] Gröning M, Hakkarainen M. Multiple headspace solid-phase microextraction of 2-cyclopentyl-cyclopentanone in polyamide 6.6 — possibilities and limitations in the headspace analysis of solid hydrogen-bonding matrices. J Chromatogr A 2004;1052:61–8. [2] Kolb B, Pospisil P. A gas chromatographic assay for quantitative analysis of volatiles in solid materials by discontinuous gas extraction. Chromatographia 1977;10:705–11. [3] Kolb B, Pospisil P, Auer M. Quantitative analysis of residual solvents in food packaging printed films by capillary gas chromatography with multiple headspace extraction. J Chromatogr A 1981;204:371–6. [4] Kolb B. Multiple headspace extraction — a procedure for eliminating the influence of the sample matrix in quantitative headspace gas chromatography. Chromatographia 1982;15:587–94. [5] Milana MR, Maggio A, Denaro M, Feliciani R, Gramiccioni L. Modern approach to the quantitative determination of volatiles in solid samples, Multiple headspace extraction gas chromatography for the determination of cyclohexanone residues in soil. J Chromatogr A 1991;552:205–11. [6] Tavss EA, Santalucia J, Robinson RS, Carroll DL. Analysis of flavor absorption into plastic packaging materials using multiple headspace extraction gas chromatography. J Chromatogr A 1988;438:281–9. [7] Wenzl T, Lankmayr EP. Reduction of adsorption phenomena of volatile aldehydes and aromatic compounds for static headspace analysis of cellulose based packaging materials. J Chromatogr A 2000;897:269–77. [8] Chai X-S, Schork FJ, DeCinque A. Simplified multiple headspace extraction gas chromatographic technique for determination of monomer solubility in water. J Chromatogr A 2005;1070:225–9. [9] Chai XS, Zhu JY. Simultaneous measurements of solute concentration and Henry's constant using multiple headspace extraction gas chromatography. Anal Chem 1998;70:3481–7. [10] Brachet A, Chaintreau A. Determination of air-to-water partition coefficients using automated multiple headspace extractions. Anal Chem 2005;77:3045–52.
M. Hakkarainen / J. Biochem. Biophys. Methods 70 (2007) 229–233 [11] Chai XS, Luo Q, Zhu JY. Multiple headspace extraction—gas chromatographic method for the study of process kinetics. J Chromatogr A 2002;946:177–83. [12] Bitz C, Doelker E. Influence of the preparation method on residual solvents in biodegradable microspheres. Int J Pharmacogn 1996;131:171–81. [13] Arthur CL, Pawliszyn. Solid phase microextraction with thermal desorption using fused silica optical fibres. Anal Chem 1990;62:2145–8. [14] De la Calle García D, Reichenbächer M, Danzer K, Hurlbeck C, Bartzsch C, Feller K-H. Use of solid-phase microextraction–capillary-gas chromatography (SPME–CGC) for the varietal characterization of wines by means of chemometrical methods. Fresenius J Anal Chem 1998;360:784–7. [15] Ezquerro Ó, Pons B, Tena MT. Multiple headspace solid-phase microextraction for the quantitative determination of volatile organic compounds in multilayer packaging. J Chromatogr A 2003;999:155–64. [16] Martínez-Uruñuela A, González-Saíz JM, Pizarron C. Multiple solid-phase microextraction in a non-equilibrium situation. Application in quantitative analysis of chlorophenols and chloroanisoles related to cork taint in wine. J Chromatogr A 2005;1089:31–8. [17] Zhang Z, Pawliszyn. Headspace solid-phase microextraction. J Anal Chem 1993;65:1843–52. [18] Pawliszyn J. Solid Phase Microextraction. Theory and practice. New York: Wiley-VCH; 1997. [19] Ai J. Solid-phase microextraction for quantitative analysis in nonequilibrium situations. Anal Chem 1997;69:1230–6. [20] Eriksson M, Fäldt J, Dalhammar G, Borg-Karlson A-K. Determination of hydrocarbons in old creosote contaminated soil using headspace solid phase microextraction and GC–MS. Chemosphere 2001;44:1641–8. [21] Hawthorne SB, Grabanski CB, Hageman KJ, Miller DJ. Simple method for estimating polychlorinated biphenyl concentrations on soils and sediments using subcritical water extraction coupled with solid-phase microextraction. J Chromatogr A 1998;814:151–60. [22] Zambonin CG, Palmisano F. Determination of triazines in soil leachates by solid-phase microextraction coupled to gas chromatography–mass spectrometry. J Chromatogr A 2000;874:247–55. [23] Doong R-A, Liao P-L. Determination of organochlorine pesticides and their metabolites in soil samples using headspace solid-phase microextraction. J Chromatogr A 2001;918:177–88. [24] Bouaid A, Ramos L, Gonzalez MJ, Fernández P, Cámara. Solid-phase microextraction method for the determination of atrazine and four organophosphorus pesticides in soil samples by gas chromatography. J Chromatogr A 2001;939:13–21. [25] Wennrich L, Popp P, Möder M. Determination of chlorophenols in soils using accelerated solvent extraction combined with solid-phase microextraction. Anal Chem 2000;72:546–51. [26] Hernandez F, Beltran J, Lopez FJ, Gaspar JV. Use of solid-phase microextraction for the quantitative determination of herbicides in soil and water samples. Anal Chem 2000;72:2313–22. [27] W. Ming-Chi, J. Jen-Fon. Determination of chlorophenols in soil samples by microwave-assisted extraction coupled to headspace solid-phase microextraction and gas chromatography–electron-capture detection. J Chromatogr A 2003;1012:111–8. [28] Ezquerro Ò, Ortiz G, Pons B, Tena MT. Determination of benzene, toluene, ethylbenzene and xylenes in soils by multiple headspace solid-phase microextraction. J Chromatogr A 2004;1035:17–22. [29] Deng X, Peng J, Luo B, Wei M, Hu W, Du J. A direct quantitative analysis method for monitoring biogenic volatile organic compounds released from leaves of Pelargonium hortorum in situ. Anal Bioanal Chem 2004;380:950–7. [30] Carrillo JD, Tena M. Determination of volatile compounds in antioxidant rosemary extracts by multiple headspace solid-phase microextraction and gas chromatography. Flavour Fragr J 2006;21:626–33.
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[31] Ezquerro Ò, Tena MT. Determination of odour-causing volatile organic compounds in cork stoppers by multiple headspace solid-phase microextraction. J Chromatogr A 2005;1068:201–8. [32] Gómez JL, García-Barrera T, Lorenzo F, Beltrán R. Use of multiple headspace solid-phase microextraction and pervaporation for the determination of off-flavours in wine. J Chromatogr A 2006;1112:133–40. [33] Carrillo JD, Tena MT. Determination of volatile oak compounds in aged wines by multiple headspace solid-phase microeextraction and gas chromatography–mass spectrometry (MHS–SPME–GC–MS). Anal Bioanal Chem 2006;385:937–43. [34] Hakkarainen M, Gröning M, Albertsson A-C. Solid-phase microextraction (SPME) in polymer characterization — long-term properties and quality control of polymeric materials. J Appl Polym Sci 2003;89:867–73. [35] Koster EHM, de Jong GJ. Multiple solid-phase microextraction. J Chromatogr A 2000;878:27–33. [36] Jeannot MA, Cantwell FF. Solvent microextraction as a speciation tool: determination of free progesterone in a protein solution. Anal Chem 1997;69:2935–40. [37] Theis AL, Waldack AJ, Hansen SM, Jeannot MA. Headspace solvent microextraction. Anal Chem 2001;73:5651–4. [38] Tankeviciute A, Kazlauskas R, Vickackaite V. Headspace extraction of alcohols into a single drop. Analyst 2001;126:1674–7. [39] Batlle R, Nerin C. Application of single-drop microextraction to the determination of dialkyl phthalate esters in food simulants. J Chromatogr A 2004;1045:29–35. [40] Lambropoulou DA, Psillakis E, Albanis TA, Kalogerakis N. Single-drop microextraction for the analysis of organophosphorus insecticides in water. Chim Acta 2004;516:205–11. [41] Lambropoulou DA, Albanis TA. Application of solvent microextraction in a single drop for the determination of new antifouling agents in waters. J Chromatogr A 2004;1049:17–23. [42] Fiamegos YC, Nanos CG, Stalikas CD. Ultrasonic-assisted derivatization reaction of amino acids prior to their determination in urine by using single-drop microextraction in conjunction with gas chromatography. J Chromatogr B 2004;813:89–94. [43] Das P, Gupta M, Jain A, Verma KK. Single drop microextraction or solid phase microextraction–gas chromatography–mass spectrometry for the determination of iodine in pharmaceuticals, iodized salt, milk powder and vegetables involving conversion into 4-iodo-N,N-dimethylaniline. J Chromatogr A 2004;1023:33–9. [44] Deng C, Yao N, Wang B, Zhang X. Development of microwave-assisted extraction followed by headspace single-drop microextraction for fast determination of paeonol in traditional Chinese medicines. J Chromatogr A 2006;1103:15–21. [45] Hansson E, Hakkarainen M. Multiple headspace single-drop microextraction — a new technique for quantitative determination of styrene in polystyrene. J Chromatogr A 2006;1102:91–5. [46] Zhang J, Su T, Kee Lee H. Headspace water-based liquid-phase microextraction. Anal Chem 2005;77:1988–92. [47] Garrigós MC, Marín ML, Canto A, Sánchez A. Determination of residual styrene monomer in polystyrene granules by gas chromatography–mass spectrometry. J Chromatogr A 2004;1061:211–6. [48] Wenzl T, Lankmayr EP. Effect of the water content of cardboard on the static headspace extraction of volatile analytes. J Sep Sci 2001;24:885–8. [49] Gröning M, Hakkarainen M. Correlation between emitted and total amount of 2-cyclopentyl-cyclopentanone in polyamide 6.6 allows the rapid assessment by HS and HS–SPME under non-equilibrium conditions. J Chromatogr A 2004;1052:151–9.
Review
Developments in multiple headspace extraction Minna Hakkarainen ⁎ Department of Fibre and Polymer Technology, School of Chemical Science and Engineering, Royal Institute of Technology, 100 44 Stockholm, Sweden Received 1 June 2006; accepted 23 August 2006
Abstract This paper reviews new developments in multiple headspace extraction (MHE), especially its combination with two miniaturized extraction techniques, solid-phase microextraction (SPME) and single-drop microextraction (SDME). The combination of the techniques broadens the applicability of SPME and SDME to quantitative determination of analytes in complex liquid and solid matrixes. These new methods offer several advantages over traditional liquid–solid, liquid–liquid and headspace extraction techniques. The potential applications include extraction of volatiles and semivolatiles from environmental and physiological samples and from different polymer products such as medical and biomedical materials, food packaging and building materials. The theoretical principals of the techniques are also briefly reviewed. © 2006 Elsevier B.V. All rights reserved. Keywords: Solid-phase microextraction; Single-drop microextraction; Multiple headspace extraction; Solid matrix
Contents 1. Introduction . . . . . . . . . . . . . . . . . . . 2. Multiple headspace extraction . . . . . . . . . 3. Multiple headspace solid-phase microextraction 4. Multiple headspace single-drop microextraction 5. Adsorption systems . . . . . . . . . . . . . . . 6. Conclusions. . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . .
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1. Introduction The extraction and quantitative analysis of volatiles and semivolatiles in solid or complex matrixes is an analytical challenge. Different liquid–solid extraction techniques, such as soxhlet extraction, microwave-assisted extraction (MAE), supercritical fluid extraction (SFE) and sonication are commonly used for the extraction of analytes from solid samples. However, these techniques are either expensive, time and labour intensive and/or use large amounts of toxic organic solvents. It is also difficult to perform accurate liquid–solid extraction in the case
⁎ Tel.: +46 8 790 8271; fax: +46 8 100 775. E-mail address: [email protected]. 0165-022X/$ - see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.jbbm.2006.08.012
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of volatile substances. If headspace techniques are used, partition of analytes from solid sample into the gaseous phase is often reduced, because of the interactions between the analytes and the matrix [1]. Internal and external calibration techniques rarely produce acceptable results due to the matrix effects, that cause considerable differences in the partition coefficients and release rates for different analytes. In 1977 Kolb and Pospisil presented a technique called discontinuous gas extraction [2]. The method was later renamed as multiple headspace extraction (MHE) [3]. The method removes the matrix effects and enables, thus, direct quantitative determination of analytes in solid matrixes by headspace techniques. This paper summarises new developments in multiple headspace extraction and especially its combination with solid-phase microextraction (SPME) and single-drop microextraction (SDME), which broadens the
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applicability of SPME and SDME to quantitative determination of analytes in solid and complex aqueous matrixes. 2. Multiple headspace extraction The multiple headspace extraction is a stepwise headspace extraction for the quantitative analysis of volatiles in solid or complex liquid samples [2]. The method theoretically calculates the total amount of analyte in a solid sample after only a few successive extractions and makes it possible to quantitatively determine volatile analytes in solid matrixes. The theoretical principals of multiple headspace extraction (MHE) were presented in 1981 by Kolb and Pospisil [3]. When a portion of the headspace is removed in the first extraction, the equilibrium between the analyte in the condensed sample and the headspace is disturbed. As the sample is allowed to re-equilibrate, more analyte migrates from the condensed phase into the headspace. The concentrations in the two phases will now be smaller than during the first extraction, but the ratio between the analyte concentration in the two phases will be the same. The second extraction and analysis, thus, results in a smaller peak and by continuing this procedure it is possible to strip off all the volatiles from the sample. If carried out ad infinitum, the various peak areas are summed up to get the total peak area, which corresponds to the total amount of the analyte in the sample. The influence of sample matrix is thus eliminated by the exhaustive extraction. As the MHE procedure follows a logarithmic function it is not required that the extractions are carried out until all the analyte is removed from the sample matrix. Instead, the logarithms of the various area values from the consecutive analyses are plotted versus the number of analyses in a linear scale and the total area value is obtained by regression calculation from the areas obtained in only a few extraction steps [4]. The total amount of a volatile analyte in a sample can be calculated by summarizing all individual peak areas (Ai), where i is the number of the extraction. As this is a converging geometrical progression, the sum can be derived as: iX ¼l i¼1
Ai ¼
A1 1−e−q
ð1Þ
3. Multiple headspace solid-phase microextraction Under the last decade there have been great improvements in the area of organic trace analysis and several new miniaturized extraction techniques have arised. Solid phase microextraction (SPME) was for the first time presented in 1990 by Pawliszyn and Arthur [13]. It is based on a thin fused silica fibre coated with polymeric absorbent or adsorbent. For sampling the SPME fibre is immersed directly into aqueous samples or in the headspace over the liquid or solid sample matrix. Several fibre materials with different selectivities are commercially available. In several recent studies solid-phase microextraction was combined with multiple headspace extraction to enable quantitative determination of analytes in solid or complex aqueous matrixes. The theoretical principals of MHS–SPME under both equilibrium [14,15] and non-equilibrium [16] conditions were also presented. The main difference compared to conventional headspace extraction is the partitioning of the analytes between the fibre, sample matrix and headspace instead of just between sample matrix and headspace. Once the equilibrium is reached the mass of the analyte extracted by the fibre coating (mf) is related to the overall equilibrium of the analyte in the three phase system, and can be expressed as [17,18]: mf ¼
Kfs Vf dm0 Kfs Vf þ Khs Vh þ Vs
ð2Þ
Where the q′ value is equal to the slope of the linear regression line and ln A1 is given by the y-intercept. Applications for multiple headspace extraction include determination of volatiles in solid matrixes such as soil [5], polymers [3,6] and cellulose based packaging materials [7]. MHE has also been applied for determination of monomer solubilities in water [8], for automated determination of vapour-to-liquid partition co-
ð3Þ
Where m0 is the total mass of the analyte, Kfs and Khs are the fibre/sample and headspace/sample distribution constants and Vf, Vh and Vs are the volumes for fibre, headspace and sample. Under the non-equilibrium conditions the analyte mass extracted for a determined period of time (mf,t) can be defined as [16,19]: mf ;t ¼ mf ;e ð1−e−at Þ ¼
The sum of all peak areas can, thus, be calculated from two values: the peak area obtained in the first extraction, A1 and the exponent q′, which describes the exponential decline of the peak areas during the stepwise MHE procedure. A1 is a measured value and the exponent q′ is obtained from the linear regression analysis: ln Ai ¼ −qVði−1Þ þ ln A1
efficients [9,10] and for studying the process kinetics involving volatile species [11]. The amount of residual organic solvent in biodegradable microspheres was also determined by MHE to study the influence of preparation method on residual solvent content [12].
Kfs Vf dm0 ð1−e−at Þ Kfs Vf þ Khs Vh þ Vs
ð4Þ
Where mf,e is the mass extracted by the fibre when the equilibrium has been reached and a is a measure of how rapidly the partition equilibrium is reached. a can be obtained from the following expression [16,19]: a ¼ 2Am
kKfs Vf þ kKhs Vh þ kVs 2mkfh Vf Vs þ kKhs Vf Vh þ kVf Vs
ð5Þ
Where A is the surface area of SPME fibre, m the mass transfer coefficient of the analyte in the SPME phase, k the evaporation rate constant and Kfh the equilibrium partition constant for the analyte between the SPME fibre and headspace. Larger a value means faster partition equilibrium. For valid MHS–SPME the relationship between the peak area and the amount of analyte in the fibre coating must be linear over the
M. Hakkarainen / J. Biochem. Biophys. Methods 70 (2007) 229–233
studied range [15]. The volumes for sample, headspace and fibre and the distribution constants between the different phases must also remain constant. In the case of non-equilibrium MHS–SPME it is crucial that all the SPME parameter in each individual extraction are kept constant [16]. The Eqs. (1) and (2), derived for multiple headspace extraction, are applicable even for multiple headspace solid-phase microextraction. Solid-phase microextraction has rapidly become one of the most frequently used extraction techniques for different environmental analysis. It has e.g. in several studies been applied for extraction of contaminants from soil samples [20– 23]. A frequently used approach to enable quantitative determination of analytes in solid or complex matrixes is to combine SPME with another extraction technique. The analytes were, thus, first extracted from soil by for example solvent extraction, microwave assisted extraction or ultrasonic extraction [24–27]. These extracts were then further concentrated on the SPME fibre and quantified. The multiple headspace solidphase microextraction (MHS–SPME) technique could in many cases offer a more attractive alternative as it is both easily automated and eliminates the use of organic solvents. Recently MHS–SPME was successfully applied for determination of benzene, toluene, ethyl benzene and xylene isomer (BTEX) in contaminated soil [28]. The extractions were carried out from soil suspensions in water at 30 °C with carboxen-polydimethylsiloxane (CAR-PDMS) fibre. The obtained BTEX concentration values in certified soil samples were statistically equal to the reference ones proving that the MHS–SPME method removed the matrix effects. A direct quantitative MHS–SPME method has also been developed to monitor biogenic volatile organic compounds (BVOCs) released from a living leaf of Pelargonium hortorum in situ [29] and for determination of volatile compounds in antioxidant rosemary extracts [30]. In addition MHS–SPME has been applied for quantitative determination of volatile organic compounds in polymers [1,15], in cork stoppers [31] and in wine [32,33]. Determination of odour-causing volatile organic compounds in cork stoppers showed that the reproducibility of MHS–SPME was similar to soxhlet extraction, but the soxhlet extraction required a concentration step to obtain analyte concentrations over the detection limit [31]. Soxhlet extraction was also non-selective, took longer time, consumed large volumes of solvent and could not be used for the most volatile analytes. Compared to microwave assisted extraction, MHS–SPME offered higher sensitivity, solvent-free extraction, reduced labour-time and complete recovery of the analyte [1]. The MHS–SPME has, thus, several advantages over the liquid–solid extraction techniques. It also offers improvements to traditional multiple headspace extraction including increased sensitivity and selectivity. In addition SPME is better suited for the extraction of semivolatiles as several studies have shown that e.g. polymer additives that were not detected after traditional HS-sampling were easily extracted and detected by HS-SPME [34]. The SPME device is also easily portable enabling field sampling of environmental samples. A slightly different concept of multiple extraction, i.e. multiple solid-phase microextraction, has been presented by Koster
231
Fig. 1. Headspace single-drop microextraction of analytes from solid sample matrix.
and de Jong [35]. This method was developed to reduce the extraction time and to increase the extraction yield and not for the quantitative determination of analytes. The SPME fibre was immersed directly into the buffer solution for non-equilibrium extraction. After the extraction the extracted analytes were statically desorbed in the desorption chamber, before a new extraction and desorption cycle. After the desired number of extractions, all the extracted analytes were injected into the LC system by one single injection. 4. Multiple headspace single-drop microextraction Traditional liquid–liquid (LLE) and liquid–solid (LSE) extractions require large amounts of highly purified solvent, and as a result of this, large amounts of hazardous waste are generated. Single-drop microextraction is a miniaturization of the traditional liquid–liquid extraction method, where the solvent to aqueous ratio is greatly reduced. The volatiles are extracted by a microdrop of non water-soluble organic solvent, which is immersed in an aqueous sample [36]. After extraction the drop is retracted into the syringe and the micro-drop is transferred to a GC for further analysis. Alternatively the micro syringe needle with the drop is suspended above the surface of sample phase to perform headspace single-drop microextraction (HS-SDME) [37,38]. HSSDME is an inexpensive and rapid technique, which uses practically no solvent and eliminates the possible memory effects, because new solvent drop is used every time. Immersion and headspace-SDME have successfully been applied for the extraction of volatiles from different liquid samples e.g. dialkyl phthalate in food simulants [39], organophosphorus insecticides in water [40], antifouling agents in water [41], amino acids in urine [42] and iodine pharmaceuticals [43]. MAE and HS-SDME were combined to enable quantitative determination of paeonol in traditional Chinese medicines [44]. Recently, the headspace-single drop microextraction technique was for the first time applied for the extraction of volatiles from solid samples [45]. Quantitative determination of volatiles became possible by combining the SDME and multiple headspace extraction techniques. Fig. 1 shows a schematic drawing over the HS-SDME of
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solid samples. Similar to the MHS-SPME, multiple headspace single-drop microextraction (MHS–SDME) system consists of three phases: sample matrix, headspace and micro-drop of organic solvent. After the equilibrium has been reached the mass of the analyte extracted by the micro-drop (md) can be calculated from Eq. (3), developed for three phase systems, if mf, Vf and Kfs in the equation are replaced by md, Vd (volume for micro-drop) and Kds (micro-drop/sample distribution constant), respectively [46]: md ¼
Kds Vd dm0 Kds Vd þ Khs Vh þ Vs
ð6Þ
Where m0 is the total mass of the analyte, Khs is the headspace/sample distribution constant and Vh and Vs are the volumes for headspace and sample. Similarly under the nonequilibrium conditions the analyte mass extracted for a determined period of time (md,t) can be defined as: md;t ¼ md;e ð1−e−at Þ ¼
Kds Vd dm0 ð1−e−at Þ Kds Vd þ Khs Vh þ Vs
ð7Þ
Where md,e is the mass extracted by the micro-drop when equilibrium has been reached and a is a measure of how rapidly partition equilibrium is reached. Soxhlet, microwave-assisted extraction, supercritical fluid extraction, headspace extraction and dissolution–precipitation techniques have earlier been evaluated and compared for determination the residual styrene content in polystyrene granules [47]. However, only the dissolution–precipitation method gave good results. Validation of the developed MHS–SDME method against the dissolution–precipitation method showed good agreement [45]. However, compared to the dissolution–precipitation method, the MHS–SDME method reduced labourtime and was practically solvent-free. 5. Adsorption systems Problems may arise in quantitation by MHE when both the analyte and the solid sample are polar. Adsorption of analytes is a well-known problem when extracting environmental pollutants from soil. It is also a problem during extraction of analytes from cardboard [7] and polar polymer samples [1]. These systems are referred to as adsorption systems, due to the adsorption of the analyte by the sample matrix. If the system is an adsorption system there will be no exponential decrease in peak area throughout the successive extractions. The system can be changed from an adsorption system into a partitioning system by adding a compound that has higher affinity towards the adsorbing sites in the matrix than the analyte of interest. Such a compound is called a displacer or modifier. The use of water as a displacer allowed quantitative determination of cyclohexanone in soil samples and increased the recovery from 4% to 99.4% [5]. BTEX (benzene, toluene, ethylbenzene and xylene isomers) in soil could also be analysed quantitatively by using MHS–SPME when water was added to the samples [28]. When water was added as a displacer, the amount of hexanal extracted from cardboard increased six times, clearly showing the higher affinity of the polar adsorption sites of the matrix towards water
[48]. The establishment of equilibrium between 2-cyclopentylcyclopentanone in polyamide-66 and the headspace required 12 h of incubation at 80 °C [49]. However, the addition of water as a displacer released the 2-cyclopentyl-cyclopentanone from polyamide-66 matrix and allowed rapid assessment of the analyte by MHS–SPME [1]. 6. Conclusions Combination of headspace solid-phase microextraction or headspace single-drop microextraction with multiple headspace extraction broadens the applicability of HS–SPME and HS– SDME to quantitative determination of volatiles in complex liquid and solid matrixes. MHS–SPME and MHS–SDME offer several advantages compared to both liquid–liquid and liquid– solid extraction techniques and traditional multiple headspace extraction. MHS–SPME is a solvent-free, easily automated, portable and sensitive technique. It is applicable for extraction of volatile and semivolatile compounds and the selectivity can be changed by changing the fibre material. MHS–SDME is a practically solvent-free, cheap and sensitive technique. It also eliminates possible memory effects as fresh solvent-drop is used for each extraction. The potential applications include extraction of volatiles and semivolatiles from e.g. environmental and physiological samples and from different polymer products such as medical and biomedical materials, food packaging and building materials. References [1] Gröning M, Hakkarainen M. Multiple headspace solid-phase microextraction of 2-cyclopentyl-cyclopentanone in polyamide 6.6 — possibilities and limitations in the headspace analysis of solid hydrogen-bonding matrices. J Chromatogr A 2004;1052:61–8. [2] Kolb B, Pospisil P. A gas chromatographic assay for quantitative analysis of volatiles in solid materials by discontinuous gas extraction. Chromatographia 1977;10:705–11. [3] Kolb B, Pospisil P, Auer M. Quantitative analysis of residual solvents in food packaging printed films by capillary gas chromatography with multiple headspace extraction. J Chromatogr A 1981;204:371–6. [4] Kolb B. Multiple headspace extraction — a procedure for eliminating the influence of the sample matrix in quantitative headspace gas chromatography. Chromatographia 1982;15:587–94. [5] Milana MR, Maggio A, Denaro M, Feliciani R, Gramiccioni L. Modern approach to the quantitative determination of volatiles in solid samples, Multiple headspace extraction gas chromatography for the determination of cyclohexanone residues in soil. J Chromatogr A 1991;552:205–11. [6] Tavss EA, Santalucia J, Robinson RS, Carroll DL. Analysis of flavor absorption into plastic packaging materials using multiple headspace extraction gas chromatography. J Chromatogr A 1988;438:281–9. [7] Wenzl T, Lankmayr EP. Reduction of adsorption phenomena of volatile aldehydes and aromatic compounds for static headspace analysis of cellulose based packaging materials. J Chromatogr A 2000;897:269–77. [8] Chai X-S, Schork FJ, DeCinque A. Simplified multiple headspace extraction gas chromatographic technique for determination of monomer solubility in water. J Chromatogr A 2005;1070:225–9. [9] Chai XS, Zhu JY. Simultaneous measurements of solute concentration and Henry's constant using multiple headspace extraction gas chromatography. Anal Chem 1998;70:3481–7. [10] Brachet A, Chaintreau A. Determination of air-to-water partition coefficients using automated multiple headspace extractions. Anal Chem 2005;77:3045–52.
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