Element selective detection for supercritical-fluid chromatography

J. Biochem. Biophys. Methods 43 (2000) 45–58 www.elsevier.com / locate / jbbm

Review

Element selective detection for supercritical-fluid chromatography a b, Nohora P. Vela , Joseph A. Caruso * a

US Food and Drug Administration, Forensic Chemistry Center, 6751 Steger Drive, Cincinnati, OH 45237, USA b University of Cincinnati, Department of Chemistry, Room 137, McMicken Hall, Cincinnati, OH 45221 -0172, USA

Abstract This manuscript describes the use of Supercritical-Fluid Chromatography (SFC) with plasma spectrometric detection for the analysis of organometallics. An introduction on the principles and characteristics of Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP-AES) and Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) is included, along with a discussion about requirements for coupling SFC to plasma detection and the different approaches for interfacing SFC to ICP. The last part of this review paper provides a comprehensive description of SFC–ICP applications for the analysis of organometallics containing iron, silicon, tin, chromium, arsenic, lead, mercury and antimony.  2000 Published by Elsevier Science B.V. All rights reserved. Keywords: Supercritical-fluid chromatography; Inductively coupled plasma-atomic emission spectrometry; Inductively coupled plasma-mass spectrometry; Speciation; Organometallics

1. Introduction Over the past decades, there has been a marked interest in the development and improvement of spectroscopic instrumentation for metals determination. This effort has been driven by the findings of environmental, and forensic studies showing that certain metals are toxic at concentrations much lower than initially believed. Additionally, toxicity of a metal containing compound is not only a function of the metal itself but is highly dependent on the metal’s oxidation state as well as the identity of the functional *Corresponding author. Tel.: 1 1-513-556-5858; fax: 1 1-513-556-0142. E-mail address: [email protected] (J.A. Caruso) 0165-022X / 00 / $ – see front matter  2000 Published by Elsevier Science B.V. All rights reserved. PII: S0165-022X( 00 )00091-9

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groups attached to the metal [1]. The process of separation and quantification of metal species is known as ‘Metal Speciation’. In this article ‘metal’ is used loosely to include metalloids. The process of metal speciation often requires the combination of two complementary techniques. The first technique allows for the efficient separation of the species of interest, while the second is responsible for the detection and quantitation. Compatibility between these two techniques is also required. This may refer to the compatibility between the chromatographic mobile phase and its flow-rate to the response and tolerance of the detector to it. The capability of the detector to acquire ‘real time’ data is not mandatory but certainly an advantage when coupling two techniques. The most common separation techniques are gas chromatography (GC) and liquid chromatography (LC). However, only few metal containing compounds are volatile and preparation of derivatives is not only time consuming but not always practical in speciation studies. LC offers several separation modes including reversed-phase, ionpair, ion-exchange, micellar and size exclusion chromatography and several reviews have been published covering these topics [2–6]. A list of element selective detectors include atomic absorption spectroscopy (AAS), graphite furnace atomic absorption spectroscopy (GF-AAS), flame / laser-excited atomic fluorescence spectroscopy, ICPAES and ICP-MS. Among these detectors only ICP-AES and ICP-MS allow practical on line, ‘real time’ detection for both gaseous and liquid samples. Typical LC flow-rates are compatible with ICP uptake flows, but the ICP tolerance to LC mobile phases containing organic solvents or more than 0.2% dissolved solids is limited. This problem is even more severe when using ICP-MS as a detector since salt deposition occurs not only in the nebulizer and in the inner tube of the ICP torch, but also in the sampler and skimmer orifices. Advantages and characteristics of SFC suggest that this would be the method of choice for compounds that are not easily amenable to GC or LC separation. One of the advantages of coupling SFC with plasma detection is the ability to use the separating power of liquid-like supercritical-fluids with the selectivity and sensitivity of plasma spectrometry for gaseous samples. Therefore, the combined effects of SFC and plasma spectrometry suggests the potential for an efficient and rapid method for metal speciation.

2. Plasma spectrometry A plasma is a gas or mixture of gases in which a fraction of their atoms or molecules is ionized [7–9]. The ICP is initiated and sustained by induction from a high-frequency magnetic field. The most common gases used in ICPs are argon and helium. A plasma is formed in a torch that consists of an assembly of three concentric tubes [7–9]. Plasma gas is passed through the three tubes of the torch at different flow-rates and each one has a specific function. The flow carried between the intermediate and the outer tube is known as the support flow or cooling gas. This plasma gas is introduced tangentially and forms a vortex flow to center, which provides a toroidal shape plasma. Another function of this flow is to cool the inside walls of the torch, preventing it from melting. Typical coolant flows are between 10 and 16 l / min. The gas flowing through the inner tube of

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the torch is the nebulizer gas which functions to transport the sample as an aerosol to the plasma. Nebulizer gas flows are usually between 0.8 and 1.5 l / min. The third flow, called auxiliary flow comes between the inner and the middle tube. Its function is to position the plasma and keep the plasma away from the torch. Power is coupled from the radio frequency (rf) generator to the torch via a water-cooled load coil. This coil, made of copper, has three or four turns and surrounds the upper part of the torch. The ICP high frequency generator operates typically at 27.12 or 40.68 MHz with output levels of 0.5–2.5 kW. The process of ionization, for the supporting argon gas is initiated by seeding electrons in the load coil space using a Tesla discharge coil [7–9]. Argon ions are formed in the plasma, and by absorbing sufficient energy from the rf generator, they maintain temperatures ranging from 6000 to 9000 K, sustaining the plasma indefinitely. After a plasma is initiated and stabilized, the nebulizing gas, which produces the aerosol from the sample, is introduced. The auxiliary flow is optional and depends on the instrumentation and particular conditions of analysis. Penetration of the sample as an aerosol occurs into the center of the plasma. Atomization, ionization and excitation of the sample occur in the central channel of the plasma through energy transfer, mainly by thermal conduction. The excited species may be determined by atomic emission spectrometry. One of the advantages of ICP-AES is the capability for simultaneous or rapid sequential determination of the elements at different concentrations ranging from major to trace amounts without changing experimental parameters. Linearity is found over five orders of magnitude and detection limits in the ppb range are common. Argon plasmas are also used as ion sources for mass spectrometry because of their efficiency and stability on the excitation / ionization process. A combination of the ionizing power of the ICP, with the sensitivity and specificity of mass spectrometry results in a highly sensitive and element-selective technique known as ICP-MS [9]. The ICP yields primarily single charged ions, M 1 , and the presence of doubly charged ions is expected only from elements like Ba 21 that have low second ionization potential. A disadvantage in ICP-MS is the formation of polyatomic species in the plasma. This is due to the interaction between matrix elements and the atmospheric and plasma gases (N 2 , O 2 , Ar 2 , Xe). Isobaric elemental interferences in ICP-MS also take place when isotopes of different elements combine to form an ion with the same nominal m /z as the element of interest. A typical example of isobaric interference is the 40 Ar 35 Cl 1 formation that interferes with the monoisotopic arsenic determination at m /z 75. The use of a quadruple mass analyzer gives better than unit mass resolution over a mass range up to m /z 5 300 amu. ICP-MS is considered a sequential multielement analyzer, with scan time less than 20 ms for one sweep from m /z 5 1 to 200 a.m.u. ICP-MS is characterized by high sensitivity (detection limits often less than the ppt level), wide linear dynamic range, and isotope analysis capability with a precision of 0.1–3% relative standard deviation.

3. Considerations in coupling SFC and ICP The main factors to consider when coupling SFC to plasma detection are analyte

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transport efficiency and plasma response to supercritical-fluids. Carbon dioxide is so far the most common mobile phase. Advantages of using CO 2 as mobile phase for SFC include availability and convenient critical point (31.18C and 72.8 atm) for chromatographic purposes [10]. Additionally CO 2 is non-toxic, non-flammable, inexpensive and commercially available at high purity. However, the introduction of CO 2 to the plasma should be limited due to possible carbon deposition on the sampler and skimmer. Thus, there is a marked preference in selecting either packed microcolumns [11,12,26,30,32] or capillary columns [13–24] over packed SFC columns. Shen et al. [13] have noted carbon deposition on the sampler of the ICP-MS system after running for 10 min CO 2 (as auxiliary flow) at 10 ml / min [13]. Other authors reported no appreciable carbon deposition on the sampler when introducing CO 2 at flow-rates ranging from 1 to 3 ml / min. These gas flow-rates are typically obtained with a 50 mm i.d. capillary column when using a pressure program ramping from 80 to 400 atm [14].

3.1. SFC–ICP interface In theory interfacing SFC with ICP is simple since after exiting the restrictor, the sample is a gas, therefore nebulizer and spray chamber typically used for liquid sample introduction to the plasma can be eliminated and replaced with the SFC–ICP interface. Coolant and auxiliary argon flows are kept the same as in regular plasma operation. Nebulizer gas is necessary to penetrate the central channel of the plasma while carrying the eluent. In summary, no other modifications are necessary to interface SFC to ICP. However the interface must provide enough heat to the tip of the restrictor because that point is where the supercritical-fluid expands to a gas at atmospheric pressure. This process of changing from a SF to a gas, is subject to the Joule–Thomson effect and a net cooling is the result. Thus, sufficient heat should be provided to the restrictor in order to avoid cluster formation and wall condensation [10,15]. There are two basic designs for coupling SFC to ICP. One has been proposed for interfacing packed microcolumn SFC to ICP-AES [11,12]. The other design with slight modifications among authors has been used for coupling capillary SFC to either plasma optical emission or plasma mass spectroscopy [13–24]. In the first interface used for packed microcolumns, the outlet of the restrictor is located in a heated cross flow nebulizer as indicated in Fig. 1A. For capillary columns, the restrictor is inserted into the central tube of the ICP torch (Fig. 1B and C). A regular ICP torch (Fig. 1B) or a direct injector nebulizer (DIN) torch (Fig. 1C) have been used for coupling capillary SFC to ICP. Olesik et al. [16] published the first paper dealing with the introduction of supercritical-fluids to ICP-AES. In that report, the restrictor was made at the end of the capillary tube that was used to transport the SF. This capillary was inserted in a copper tube where preheated argon is also flowing. The copper tube is heated by a resistive heater wire and the temperature of the argon flowing inside the copper tube is monitored and controlled by a variable transformer. For an estimated supercritical-fluid rate between 20 and 300 ml / min, Olesik et al. [16] found satisfactory results using a low flow ICP torch and operating the plasma at 1.25 kW. The paper published by Forbes et al. [15] describes the use of a DIN torch for

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Fig. 1. SFC–ICP interfaces. (a) Interface design for packed microcolumns, (b) interface for capillary SFC using a regular ICP torch and (c) interface for capillary SFC using a DIN torch. Reprinted from Carey et al. [19] with permission of Elsevier Science Publishers.

capillary SFC coupled to plasma atomic emission spectrometry. A heated transfer line is used to maintain the column temperature between the outlet of the SFC oven and the base of the ICP torch. A butt connector is installed to join the capillary column to the restrictor. In this case, the nebulizer gas that enters by the third arm of the DIN torch is also used to push the column effluent into the plasma. The interface design by Shen et al. [13] uses also a heated transfer line and a butt connector to couple the capillary column to the frit restrictor. The interface has a series of Swagelock unions to force the flow of the preheated argon along with the column effluent to enter the central channel of the plasma. The tip of the restrictor is positioned flush with the end of the central tube of the ICP torch. The argon is preheated by passing it through a copper coil wrapped in heating tape and insulated with glass fiber material. Separate temperature controls are installed for the transfer line and for the interface. This interface was employed for several applications of SFC coupled to ICP-MS [17–22].

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Other authors report the use of a tapered restrictor and a restrictor heater positioned inside the torch [14,23,24]. The restrictor heater consists of a Nichrome wire tightly wound around a melting point tube and insulated with a heat resistant polyimide resin. The ends of the wire were also insulated and attached to a low voltage direct current power supply. The restrictor was passed through the glass tube and positioned flush with the end of the restrictor heater. The temperature of the restrictor was increased from 200 to 3508C by varying the applied voltage between 6 and 10 V.

4. Applications of SFC to metal speciation A summary of the metal containing species analyzed by SFC with plasma spectrometric detection is presented in Table 1, and a detailed description is presented in the next paragraphs.

4.1. Speciation of iron containing compounds Ferrocenes were the first compounds used to demonstrate the capabilities of ICP-AES detection for supercritical-fluid-based sample introduction. The work conducted by Olesik and Olesik [16] describes the 0.2 ml injection of a cyclohexane solution of ferrocene in the capillary tube that carries the supercritical-fluid to the plasma. A detection limit of 60 pg of Fe was reported in this paper. A mixture of ferrocene, acetylferrocene and benzoylferrocene was separated by SFC using CO 2 as mobile phase and a stainless steel microcolumn (25 cm 3 1 mm i.d.) packed with Shimpak Diol-150. The column temperature was maintained at 408C and the inlet column pressure at 160 atm. The iron emission at 259.94 nm indicates that for a Table 1 Organometallics analyzed by SFC with plasma spectrometric detection Compounds Ferrocenes Ferrocenes Ferrocenes Organosilicon Tetraalkyltin Tri and tetraorganotins Tri and tetrabutyltins Acetyl acetone chromium complex Ketone chromium Organoarsines Triphenylarsine Organolead Diethylmercury Diphenylmercury Organoantimony a

n.s.: Not specified.

ICP torch design

Detector

Linear range a

Detection limit

Ref.

Low-flow torch Regular torch Regular torch Direct Inject Neb. Regular torch Regular ICP torch Regular ICP torch Regular ICP torch

ICP-AES ICP-AES ICP-MS ICP-AES ICP-MS ICP-MS ICP-MS ICP-AES

n.s. 7.5–375 ng n.s.a 7.24–145 ng 1–1000 pg 0.05–5 ng 0.1–10 ppm n.s.a

60 pg 7.5 ng n.s.a 5.8 ng 0.034–0.047 pg 0.2–0.8pg 0.07–6.7 pg n.s.a

[16] [11] [24] [15] [13] [17] [23] [12]

Regular ICP torch Regular ICP torch Regular ICP torch Regular ICP torch Regular ICP torch Regular ICP torch Regular ICP torch

ICP-MS ICP-MS ICP-MS ICP-MS ICP-MS ICP-MS ICP-MS

0.1–100 ng n.s.a n.s.a 0.5–50 ng 0.05–50 ng n.s.a n.s.a

0.9–3 pg 0.48–4.8 pg 100 ppb 0.5–10 pg 3 pg 0.05 pg 0.01 pg

[20] [21] [24] [22] [21] [21] [21]

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0.5 ml ferrocene injection, the lowest detectable concentration is 50 ng / ml for ferrocene (15 ng / ml iron) at a signal-to-noise ratio of 4 [11]. Blake [24] reports the use of SFC–ICP-MS for the analysis of ferrocene solutions. However, they found elevated background signal for iron determination at its major isotopes (m /z 56 and m /z 54) due to the presence of 40 Ar 16 O 1 and 40 Ar 14 N 1 polyatomic species in the plasma. A very small peak was observed for an injection of a 100 ppm solution and this might be due to the ArFe 1 formation. This assumption was made by the authors since a scan between 92 and 100 amu shows peaks at m /z 94, 96 and 97.

4.2. Speciation of silicon containing compounds Forbes et al. [15] describe the use of SFC–ICP-AES for the separation and detection of two tetrasiloxanes (octamethylcyclotetrasiloxane and 1,7-dichlorooctamethyltetrasiloxane) at the Si I 251.6 nm emission line. A fused-silica capillary column coated with poly(dimethylsiloxane) and maintained at 608C was used for the separation along with CO 2 as mobile phase. The temperature of the transfer line was 708C, while the base of the torch was kept between 80 and 908C. Isobaric pressures of 80 and 100 atm were tested for the separation of the tetrasiloxanes, however, better resolution was obtained when the pressure was manually ramped from 80 to 100 atm. The linear dynamic range for octamethylcyclotetrasiloxane was between 7.24 and 145.0 ng of Si injected for a 0.1 ml injection.

4.3. Speciation of organotin compounds The potential of ICP-MS as an element selective detector for SFC was initially demonstrated with tetraorganotin compounds [13]. The separation of tetrabutyltin and tetraphenyltin was obtained using carbon dioxide and a SB Octyl 50 capillary column (50 mm i.d., 195 mm o.d., 0.25 mm film thickness and 2.5 m length). The chromatographic conditions for the separation were optimized in the univariant mode by evaluating the effect of isobaric pressure, pressure ramp and hold time in the pressure program. Base-line resolution for the mixture of tetraorganotins was obtained when the initial pressure of 100 atm was held for a minute followed by a pressure ramp of 80 atm / min to a final pressure of 200 atm. This paper also illustrates the capability of ICP-MS to obtain multielement chromatograms by monitoring the same element at different isotopes. Results presented in this paper indicate that by comparing the actual ratio with the experimental ratio for m /z 120 / 118 and 120 / 116, the error varies from 1.9 to 2.6%. Vela and Caruso [17] used SFC–ICP-MS for the speciation of tetra and triorganotin compounds utilizing a SB biphenyl capillary column and carbon dioxide mobile phase. This paper describes in detail the effect of carbon dioxide in the argon plasma and the procedure followed to optimize the ICP-MS operating conditions. Parameters in SFC such as hold time, carbon dioxide pressure program, mobile phase composition and column length are optimized for a mixture containing tetrabutyl tin (TBT), tributyltin chloride (TrBT-Cl), triphenyltin chloride (TrPT-Cl), and tetraphenyltin (TPT). Detection

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limits for these organotin are 0.26, 0.80, 0.57 and 0.20 pg, respectively, with linear dynamic ranges of three orders of magnitude. A separate paper published by the same authors [18] compares SFC–FID with SFC–ICP-MS for the speciation of TBT, TrBT-Cl, TrPT-Cl and TPT. The effect of the SFC–ICP interface temperature in peak shape and peak area for these organotins is also presented and compared to the results obtained with SFC–FID [18]. Results obtained by SFC–ICP-MS indicate that retention time, peak area and peak width of phenyltin compounds (TrPT-Cl and TPT) are more susceptible to changes as a function of the interface temperatures (215–3508C) as compared to the butyltin compounds (TBT and TrBT-Cl). However, when using SFC–FID, no variations in peak area or peak shape are observed when varying the detector temperature over the same temperature range. Other parameters compared in this paper include resolution, detection limits, linear dynamic range and reproducibility. Results indicate better chromatographic resolution when using SFC–FID as compared to SFC–ICP-MS. However, sensitivity is one order of magnitude better with SFC–ICP-MS than with SFC–FID. Fig. 2 illustrates the chromatograms obtained for a mixture of tri and tetraorganotins analyzed by SFC–FID and by SFC–ICP-MS. Low cost and ease of operation are evident characteristics of SFC–FID but sensitivity and selectivity are the predominant advantages of ICP-MS for the analysis of real samples by SFC–ICP-MS. The paper published by Blake [23] describes the use of a new SFC–ICP-MS interface for the speciation of organotin compounds. According to their results on background scans this interface presents several advantages over the interface design by Shen [13] and used by Vela and Caruso for the speciation of tri and tetraorganotin compounds [17]. Blake and co-authors used the same column as Vela [17] and initially mention TBT, TrBT-Cl, TrPT-Cl and TPT as the organotins selected to demonstrate the advantages of the new interface. However, the study is restricted only to TBT and TrBT-Cl and the authors claim significant memory effect for TrPT-Cl at the injector. As previously mentioned, the study conducted by Vela and Caruso indicates that TrPT-Cl and TPT are more sensitive than TBT and TrBT-Cl to variations in the interface temperature. Blake et al. did not address this problem.

4.4. Chromium containing compounds by SFC–ICP-MS Stability of chromium acetyl acetate complexes in supercritical-fluid CO 2 was initially evaluated by Jinno [12] using photodiode array UV and ICP-AES. The work presented by Carey et al. [20] demonstrates base-line resolution for the separation of chromium(III) 2,2,6,6-tetramethyl-3,5 heptanedionate (MHDC), chromium(III) 2,4 pentanedionate (PDC) and pentamethyl cyclopentadienylchromium dicarbonyl dimer (MCCD) using SFC with flame ionization detection. This separation was obtained using a 4 m long capillary (50 mm i.d.) SB biphenyl 30 column with supercritical CO 2 and a pressure program started constant at 85 atm for 4 min, followed by a pressure ramp of 30 atm / min to a final pressure of 360 atm. The oven temperature was maintained at 708C. However, the chromatographic conditions optimized for SFC–FID required some modification when coupling SFC with ICP-MS. First, due to the isobaric interference at m /z 5 52 ( 40 Ar 12 C 1 ) for chromium determination at its most abundant isotope, it was

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Fig. 2. (a) SFC–FID for organotins using the following conditions: oven temperature: 758C, initial pressure 70 atm held for 1 min, pressure ramp: 60 atm / min, final pressure: 300 atm. (b) SFC–ICP-MS chromatogram for an oven temperature of 508C and the same pressure program as described for (a). Peaks: 1: TBT, 2: TrBT-Cl, 3: TrPT-Cl and 4: TPT. Reprinted from Vela et al. [18] with permission of Elsevier Science Publishers.

necessary to use NO 2 instead of CO 2 as mobile phase. Second, the ramp rate in the pressure program was increased up to 70 atm / min and finally the peak corresponding to MCCD was not observed, possibly due to its thermal decomposition and / or adsorption to the capillary walls. A comparison in analytical figures of merit indicates an improvement of at least one order of magnitude in sensitivity when using ICP-MS detection as compared to FID.

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4.5. Speciation of organoarsenic compounds Kumar and co-authors [21] illustrate the use of SFC–ICP-MS for the separation of trimethyl arsine (TMAs), triphenyl arsine (TPAs) and triphenyl arsine oxide (TrPAsO) all dissolved in methanol and injected in a 2.5 m 3 50 mm i.d. SB biphenyl 30 column. The same mixture was injected in a SFC–FID system and the TrMAs peak overlaps with the solvent peak, making it impossible to detect (refer to Fig. 3). The pressure program used for the separation of organoarsenicals is the same for both FID and ICP-MS detection. However, better results are observed for an oven temperature of 508C for SFC–ICP-MS and 908C for SFC–FID. Fig. 3 compares the SFC–FID and SFC–ICP-MS chromatograms obtained for a mixture of organo–arsenicals. Detection limits for TMAs, TPAs, and TPAsO are 4.8, 3.1 and 0.43 pg, respectively. Determination of TrPAs by SFC–ICP-MS has also been reported by Blake and co-authors [24], who describe a high background signal at m /z 75 due to the presence of 40 Ar 35 Cl 1 in the plasma. Since arsenic is monoisotopic, there is no alternative mass for quantitation. The minimum TrPAs concentration that can be detected in the system designed by Blake is 100 ppb.

4.6. Speciation of organolead compounds The first paper mentioning the detection of organoleads by SFC–ICP-MS was published by Vela and Caruso [17]. In that paper the authors demonstrate the capability of SFC to separate two organometallics containing the same organic substituents but different central atoms. The compounds were tetraphenyl tin and tetraphenyl lead. In another publication [18] the same authors emphasize on the advantages of ICP-MS selective detection as compared to FID for the same mixture of tetraphenyl metallics. Carey [22] describes the use of SFC–ICP-MS for the separation and detection of tetraethyl lead (TTEL) and tributyl lead acetate (TRBL). However, the apparent instability of TRBL under SF conditions and the presence of an additional peak when the mixture of TTEL and TRBL is injected, does not favor the determination of TRBL by SFC–ICP-MS. This paper also describes the use of SFC–ICP-MS for the quantification of TTEL in a SRM NIST 2715 ‘Lead in Reference Fuel’. The results obtained by this method are in excellent agreement with the certified value.

4.7. Organomercury and antimony compounds by SFC–ICP-MS Analysis of diethyl mercury (DIEM) by SFC–ICP-MS has been described [19]. In this paper the authors compare detection limits for DIEM using two different acquisition modes. The usual mode for collecting chromatographic information from ICP-MS is by Single Ion Monitoring (SIM) mode. In this case the instrument is set at a specific m /z and the signal is recorded as a function of time. However, some instruments are also equipped with a ‘Time Resolved Acquisition’ (TRA) software that allows for a single injection, the capability to monitor several isotopes of the same element or different elements at their characteristic m /z. Calculations indicated detection limits in the low pg range for both SIM and TRA modes.

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Fig. 3. Chromatograms obtained by (A) SFC–ICP-MS for arsenic, antimony and mercury compounds (B) SFC–FID for organoarsenicals. Adapted with permission from Ref. [21].

The paper published by Kumar et al. [21] also describes the use of the TRA software for the analysis of diphenyl mercury (DPHg) and triphenyl antimony (TPSb) by SFC–ICP-MS. Fig. 3 displays the chromatograms obtained for DPHg and TPSb.

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Mercury measurements were made at m /z 202, 200, 199 and 201 while antimony was monitored at m /z 121 and 123. Good agreement was found when comparing the natural abundance with the calculated abundance for the different antimony and mercury isotope monitored.

4.8. Stability of acetylacetone compounds The paper published by Jinno [12] describes the results obtained for the stability study under supercritical conditions of acetylacetone complexes. A microbore packed SFC column, a photodiodide array UV detection system and an ICP atomic emission spectrometer were used for the study. The dimensions and packing material of the microbore column are the same as in the paper published by Fujimoto [11] and used for the separation of ferrocenes. The acetylacetate complexes of Al, Co, Cr, Cu, Fe, Mg, Mn, Ni and Zn were evaluated. The photodiode UV spectra of these complexes were taken in three mobile phases. The mobile phases used were: dichloromethane, and supercritical carbon dioxide at 508C and 140 atm with and without modifier (5% mol methanol). Comparison of the results obtained with the two detectors indicate that from the nine complexes evaluated, only the Al, Co, Cr and Cu acetylacetate complexes are stable under supercritical conditions.

5. Conclusions The potential of SFC with plasma spectrometric detection for elemental speciation studies has been described in this paper. A list of additional review articles and manuscripts describing SFC–ICP studies that does not necessary involve speciation is also included in the reference section [25–33]. However, it is important to note that the number of publications and applications in this field has been limited in part due to the popularity and the number of approved methods and technical notes based on LC and GC separations. Another factor to consider is the ‘mismatching’ between polarity of most organometallics as compared to the non-polar characteristics of the CO 2 that is the most common mobile phase for SFC. However, it is important to note that plasma detection has the advantage over universal detectors since it is ‘transparent’ to solvents, so detection of organometals is not obscured by the presence of modifiers. Formation of non-polar metal complexes from polar organometallics is another option to overcome problems related with polarity. The sensitivity and selectivity of plasma detection are a definite advantage in the analysis of ‘complex samples’. In the analysis of such samples, matrix interferences and ‘non-ideal chromatographic separations’ can be tolerated to a certain degree that enough information can be obtained about the sample. The ICP-MS potential of collecting information at different isotopes of an element serves to provide not only selective but also conclusive results about the presence of certain elements in a sample. These data can be used for quantitative analysis as well as for comparison purposes when obtaining the elemental profile of certain samples.

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Acknowledgements The authors acknowledge the collaboration of Claudia Ponce de Leon for providing a preliminary bibliographic review and a copy of some of the articles included in this manuscript.

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