Anal. Chem. 1989, 61, 236-240
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relative standard deviation to be 2.34%. Determination of Selenium in Standard Samples. The results for contents of selenium in standard samples determined by the present method are shown in Table I, which refers to the certified values and the results obtained by DAN fluorometrywith tellurium coprecipitation (19) and by atomic absorption spectrometry with hydride generation (HG-AAS) (19). These standard samples contained some coexistent metals with higher contents than selenium. Since the formation of PenSTS proceeds in strongly acid solution, only a limited sort of metal ions such as copper, which is known to form stable chelate with penicillamine, may interfere with the selenotrisulfide formation reaction. Such ions and other unknown substances may still cause deposit of precipitates during derivatization procedure, lowering of recovery and poor separation of fluorophoreon HPLC. To eliminate these fears, we added EDTA to the penicillamine solution and used a large excess of penicillamine for the formation of penicillamine selenotrisulfide. When 5 pg of Se(1V) was spiked to the digested solution of each standard sample, 95-110% of the spiked Se(1V) was recovered as shown in Table I. Thus, it is concluded that no significant interferences with the determination of selenium were not observed, though the standard samples used in this work contained about 2-70-fold higher amount of copper than selenium. Student's paired t tests (p < 5%) were carried out with respect to the selenium contents (pg/g) given as the certified value and obtained by the present method and by the previously established method. The results indicated there was no significant difference, except for the pair of the results for SRM-1645 obtained by the present method and by HG-AAS. The present method, though it takes about 21 min for a HPLC analysis, allows determination of a low level of selenium in a small sample size with easy operation. The applicability
of the present method can be expanded to include the samples with much lower contents of selenium and different sorts of interfering substances, when it is combined with the selective preconcentration method of selenium using anion exchange resin loaded with bismuthiol-I1sulfonate, which the authors developed (20).
LITERATURE CITED (1) Schwartr, K.; Foitz, C. M. J . Am. Chem. SOC. 1957,79, 3292. (2) Das, N. P.; Ma, C. W.; Salman, Y. M. Blol. Trace Elem. Res. 1988,
IO, 215. (3) Schrauzer, G. N.; White, D. A.; Schneider, C. J. Bloinorg. Chem.
1977, 7 , 23. (4) Greeder, G. A.; Milner, J. A. Science 1980,209, 825. (5) Hiraki, K.; Yoshii, 0.; Hirayama, H.; Nlshlkawa, Y.; Shlgematsu, T. Bunseki Kagaku 1973,2 2 , 713. (6) Shlbata, Y.; Morita, M.; Fuwa, K. Anal. Chem. 1084,5 6 , 1527. (7) Narasaki, H.; Ikeda, M. Anal. Chem. 1984,56, 2059. (8) Campbell, A. D. Pure Appl. Chem. 1984,56. 645. (9) Kimberly, M. M.; Paschal, D. C. Anal. Chlm. Acte 1985, 174, 203. (10) Maenhaut, W.; Dereu, L.; Tomza, U. Anal. Chim. Acta 1982, 136, 301. (1 1) Dllli, S.;Sutikno, I.J . Chromatogr. 1884,300, 265. (12) Tamarl, Y.; Ohmori, S.;Hiraki, K. Clin. Chem. 1986,32, 1464. (13) Ganther, H. E. Biochemistry l988?7 , 2899. (14) Nakagawa, T.; Hasegawa, Y.; Yamaguchi, Y.; Chikuma, M.; Sakurai, H.; Nakayama, M.; Tanaka, H. Biochem. Biophys. Res. Common. 1988, 135, 183. (15) Nakagawa, T.; Aoyama, E.; Kobayashi, N.; Tanaka, H.; Chlkuma. M.; Sakurai, H.; Nakayama, M. Biochem. Blophys . Res. Commun. 150, 1149. (16) Imai, K.; Watanabe, Y. Anal. Chim. Acta 1881. 130, 377. (17) Watanabe, Y.; Imai, K. J . Chromatogr. 1882,239, 723. (18) Itoh, K.; Nakayama. M.; Chikuma, M.; Tanaka, H. Fresenius' 2.Anal. Cbem. 1086,325, 539. (19) Itoh, K.; Chikuma, M.; Tanaka, H. Fresenius' 2.Anal. Chem. 1988, 330, 800. (20) Nakayama, M.; Tanaka, T.; Tanaka. M.; Chikuma, M.; Itoh. K.; Sakurai, H.; Tanaka, H.; Nakagawa, T. Tahnta 1987,34, 435.
RECEIVED for review December 31,1987. Resubmitted August 16, 1988. Accepted October 31, 1988.
Thermal Gradient Liquid Chromatography: Application to Selective Element Detection by Inductively Coupled Plasma Atomic Emission Spectrometry Wilton R. Biggs* and John C. Fetzer Chevron Research Company, Richmond, California 94802-0627
The use of temperature gradlents to achleve reversed-phase llquld chromatographlc Separations for systems detected by plasma technlques is demonstrated. Aqueous (detecting S) and nonaqueous (detectlng SI) systems are studied. The technique poves usekd for soMng the general eluHon problem while anowlng detectlon on an elemental basis by Inductively coupled plasma atomic emlsslon spectrometry ( ICP-AES).
INTRODUCTION This work describes the use of temperature programming in high-performance liquid chromatographic (HPLC) separations detected by inductively coupled plasma atomic emission spectrometry (ICP-AES). Although temperature programming is a widely used technique in gas chromatography, it has only recently begun to receive attention as a 0003-2700/89/0361-0236$01.50/0
means of controlling liquid chromatographic retention. Several workers (1-10) have demonstrated successful temperature-programmed separations, and the theory of temperature effects in HPLC has been developed (11,12),as well as important selectivity considerations (13) and a systematic optimization strategy (14). This work directly impacts efforts to employ plasma emission devices as specific element detectors in liquid chromatography because temperature programming allows the matching of operational requirements for optimum performance of each component in a coupled HPLC/ICP system. Specific element detection of HPLC separations offers several potential advantages; among these are the simplification of complex sample matrices, the use of elemental response factors which are independent of the chemical form of the element, and the detection of compounds having no molecular property which can be suitably exploited for de0 1989 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 61, NO. 3, FEBRUARY 1, 1989
tection purposes. Complex samples often contain components of interest that have widely varying retention behavior. The analyst must address this "general elution problem" or the full range of HPLC separation capability cannot be made available, To do so requires some form of gradient separation while attempting the specific detection experiment. Historically, the answer has been the use of solvent gradients because of the ready availability of appropriate hardware and the focus of detection in HPLC being on a molecular rather than elemental basis. A solvent gradient approach has been successfully employed (15) for the separation of various silicon compounds by HPLC and detection by direct current plasma (DCP) AES; but, in our hands, the ICP has been a much less forgiving detector. Solvent gradients identical with those successfully used with the DCP produced unacceptable background emission shifts and high reflected power readings and occasionally extinguished the ICP plasma. This behavior reflects the ICP's low tolerance (relative to the DCP) for both injection of large amounts of organic solvent vapor and changes in the nature of solvent supplied to the plasma. T o achieve optimum performance from the ICP, it must be supplied a solvent of reasonably constant composition. Gradient elutions generated by thermal means rather than through solvent changes represent an almost ideal compromise for coupled HPLC/ICP. In essence, an isocratic mobile phase can be used to produce a gradient separation. This study demonstrates the successful application of thermal gradient chromatography coupled with specific element detection using ICP-AES. EXPERIMENTAL SECTION Apparatus. The oven of a Hewlett-Packard Model 5710A gas chromatograph was used to generate thermal gradients. The HPLC column was placed inside the oven with a loop of 0.009-in. i.d. ('/le in. 0.d.) tubing connecting it to a Rheodyne Model 7125 injection valve located outside the oven and fitted with appropriate sample loops for introduction of the sample. Subambient temperatures were maintained by cooling the oven with dry ice. An Altex (Beckman)Model 110 HPLC pump supplied isocratic mobile phase to the column. Column effluent was directed to a Meinhard concentric glass nebulizer by using first 0.009-in.-i.d. stainless steel tubing from the column exit to a ZDV coupling (Alltech) and then Teflon '/le-in.-o.d. X 0.009-in.4.d. tubing from the ZDV coupling to the nebulizer. The Teflon line was fitted to the nebulizer, and then heat shrink tubing was used to seal the joint. An intentionally overtightened Teflon ferrule fitted to the Teflon line at the ZDV coupling was used to generate a back pressure on the column and stainless steel exit line. Although microconcentricnebulizer/torch assemblies have been developed (16)specifically for HPLC/ICP application,a standard Meinhard concentric glass nebulizer, Plasma-Therm torch, and spray chamber were used in this study. An RF Plasma Products Model HFP-2500F ICP source was used in conjunction with a Minuteman Model 310 scanning monochromator driven by a Superior Electric translator and purged with argon. The monochromator phototube (Thorn EM1 9781B) was driven by a Keithley Model 247 high-voltage power supply. The output of the photomultiplier tube was fed to a Keithley Model 427 variable gain current-to-voltage amplifier. The ampwied signale were then sent to a Nelson Analytical Seriea 760 interface box previously downloaded with a suitable analysis protocol. The analysis protocol was developed from options provided by Nelson Analytical chromatographysoftware (Revision 6.22). At the completion of a run, the time versus element response chromatogram was uploaded to a Hewlett-Packard Model 9816 computer fitted with an additional 0.75-Mbyte RAM board. Chromatograms were processed using the Nelson software run by a modified Basic 3.0 operating system. Raw data was stored on 3.5411. microfloppy disks by use of a Hewlett-Packard Model 9122 disk drive. Plots of raw data were obtained by using a modified version of a Nelson Analytical plotting routine (XY-
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Table I. Typical Operating Conditions Conditions ICP Source 1400 incident power, W
