Anal. Chem. 1985, 57, 2219-2222 (5) Armstrong, D. W.; Stine, G. Y. J . Am. Chem. SOC. 1983, 105, 2962-2984. (6) FuJimura, K.; Ueda, T.; Ando, T. Anal. Chem. 1983, 55, 446-450. (7) Kawaguchi, Y.; Tanaka, M.; Nakae, M.; Funazo, K.; Shono, T. Anal. Chem. 1983, 5 5 , 1852-1857. (8) Tanaka, M.; Kawaguchi, Y.; Nakae, M.; Mlzobuchi, Y.; Shono, T. J . Chromatogr. 1984, 299, 341-350. (9) Tanaka, M.; Kawaguchi, Y.; Shono, T.; Uebori, M.; Kuge, Y. J . Chromatogf. 1984, 301, 345-353. (10) Armstrong, D. W.; DeMond, W. J . Chromatogr. Scl. 1984, 2 2 , 411-415. (11) Sybilska, D.; Lipkowski, J.; WBycikowski, J. J . Chromatogr. 1982, 253, 95-100. (12) Sybilska, D.; Dgbowski, J.; Jurczak, J.; hkowski, J. J . Chfomatogf. 1984, 288, 163-170. (13) Sybilska, D. Proceedings of the VIth International Symposium on Chromatography and Electrophoresis: Presses Acad6miques Europ6en: Brussels, 1971; pp 212-221.
2219
(14) Sybilska, D.; SmolkovB-Kulemansovi, E. In "Inclusion Compounds"; Academic Press: London, 1984; Volume 3, pp 173-243. (15) Jinno, K. Chromatogfaphia 1983, 17, 367-369. (18) Uekama, K.; Hirayama, F.; Nasu, S.; Matsuo, N.; Irie, T. Chem. Pharm. Bull. 1978, 2 6 , 3477-3484. (17) HONath, C.; Meiander, W.; Nahum, A. J . Chromatogr. 1979, 186, 371-403. (18) Matsui, Y.; Mochida, K. Bull. Chem. SOC.Jpn. 1979, 52, 2808-2814. (19) Armstrong, D. W.; DeMond, W.; Alak, A,; Hinze, W. L.; Riehl, T. E.; Bui, K. H. Anal. Chem. 1985, 5 7 , 234-237. (20) Hinze, W. L.; Riehl, T. E.; Armstrong, D. W.; DeMond, W.; Ward, T. Anal. Chem. 1985, 5 7 , 237-242.
RECEIVED for review March 1,1985. Accepted June 3,1985. This study was supported within the Polish Academy of Sciences Project 03.10.
Simultaneous Determination of Nickel, Lead, Zinc, and Copper in Citrus Leaves and Rice Flour by Liquid Chromatography with Hexamethylenedithiocarbamate Extraction Susumu Ichinoki* and Mitsuru Yamazaki School of Pharmacy, Hokuriku University, Kanazawa 920-1 1, Japan
Reversed-phase llquld chromatography followed by solvent extraction with hexamethyleneammonlum hexamethylenedlthlocarbamate (HMA-HMDC) was carrkd out to determlne NI, Pb, Zn, and Cu In standard reference citrus leaves and rice flour. These samples (250 mg) were ashed wlth nltrlc acid and perchloric acid. The metals In the ash were extracted into chloroform as HMDC chelates which were then separated on a C 18 column and rnonltored at 260 nm. This method permitted Separation and determlnatlon of Cd-, NI-, Pb-, Zn-, Cu-, Hg-, Co-, and BI-HMDC chelates. Cd, Hg, Co, and BI could not be detected, but the microgram per gram levels of NI, Pb, Zn, and Cu In the standard blologlcal materials were simultaneously determined wlthln 25 min.
In recent years, high-performance liquid chromatography (HPLC) has been applied to the separation and determination of various metals. In principle, HPLC enables the simultaneous determination of several metals to an extent comparable to that of atomic absorption spectrometry (AAS) and spectrophotometry. Some reviews (1-3) and many papers on this subject have been published. A number of papers dealing with the separation of metal chelates have been published, but not many with the determination of metals in real samples, except for water. The following reports have been published: determination of Cd, Co, Cu, Pb, Hg, and Ni in zinc sulfate plant electrolyte (4); Mn, Fe, Co, Ni, Cu, Zn, and P b in steel ( 5 ) , Ni-Cr-Fe alloys, zirconium, and uranium (6);Cu, Ni, Pb, and Mn in a standard kale and fish meal (7). A common procedure is to separate the metals as metal chelates, and the most commonly used chelating agents are dithiocarbamates since they react with many heavy metals and the metal chelates have large molar absorptivity in the ultraviolet (UV) region. Consequently, the simultaneous and sensitive determination of heavy metals is possible with a 0003-2700/85/0357-2219$0 1.50/0
conventional HPLC equipped with a UV detector. On-column formation of dithiocarbamato chelates has been carried out by some workers (8-11). This method is simple, but the sensitivity is less than that by precolumn formation (10). Solvent extraction permits preconcentration of metals as chelates and provides relatively sharp peaks (good resolution). In previous work, we attempted the simultaneous determination of heavy metals by precolumn formation (solvent extraction) and reversed-phase HPLC with diethyldithiocarbamate (DDTC) (12), tetramethylenedithiocarbamate (13, 14), and HMA-HMDC (15,16) and published our results on the separation and determination of heavy metals (Cd, Ni, Pb, Zn, Cu, Hg, Co, Bi) in water (12,13,15), standard orchard leaves (14), and standard bovine liver and oyster tissue (16). The standard biological samples (14, 16) could be ashed by the dry ashing method (450-550 "C) without any loss of the metals to be determined. However, not all biological samples could be ashed successfully by dry ashing because of various matrices present in the samples. Actually, the standard citrus leaves and rice flour could not be ashed successfully with a muffle furnace. In this paper, wet ashing has been investigated for its application to the HPLC determination of heavy metals in plant samples. This method is based on a wet ashing method (HN03 + HC104) and precolumn formation, separation, and UV detection of the metal-HMDC chelates. A new eluent composition was employed so as to take advantage of a longer column.
EXPERIMENTAL SECTION Reagents. All reagents used were of analytical grade unless otherwise stated. Standard reference materials (citrus leaves and rice flour) were obtained from National Bureau of Standards (NBS, Washington DC). Nitric acid, perchloric acid, and ammonia water were of special grade; for example, the content of Zn was 10 ng/mL (ppb) and 0 1985 American Chemical Soclety
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 12, OCTOBER 1985
that of Pb, 1 ppb in the perchloric acid. Ammonium citrate solution (pH 9) was shaken with a dithizone-chloroform solution to remove heavy metals. The mixed standard solutions for the analysis of citrus leaves (A) and rice flour (B) were prepared as follows. Standard solution (A) contained 0.1 M HC1,30 ppb of Ni, 665 ppb of Pb, 1.45 pg/mL (ppm) of Zn, and 825 ppb of Cu. Standard solution (B) contained 8 ppb of Ni, 970 ppb of Zn, 110 ppb of Cu, and 0.1 M HC1. Five milliliters of each solution contained equal amounts of heavy metals to 250 mg of the sample as calculated from certified NBS values. The column packing used was Cosmosil 5 C 18 (5 pm ODs, Nakarai Chemicals, Ltd., Kyoto, Japan). The Cosmosil5 C 18 was a spherical silica based C 18 bonded packing (pore size, 110 A; surface area, 330 m2/g; carbon %, about 22%). All solvents used were of LC grade. HMA-HMDC was synthesized with hexamethylenimine and carbon disulfide. The detailed procedure, slightly different from Busev's method (17), is given in a previous work (16). Apparatus. A liquid chromatograph consisting of a Model KHP-010 pump (Kyowa Seimitsu Co., Ltd., Tokyo, Japan), a Model 7125 Rheodyne injector (Rheodyne, Inc., Contati, CA), a Model UVILOG-7 variable wavelength detector (Oyobunko Kiki Co., Ltd., Tokyo, Japan), and a Chromatopac C-R1B data processor (Shimadzu Co., Kyoto, Japan) was used. Eluted metal chelates were monitored at 260 nm. A schematic diagram of the HPLC system is given elsewhere (15). A Cosmosil5 C 18 column (Nakarai Chemicals) was immersed in a water thermostat. A Model PR-151 plasma reactor (Yamato Scientific Co., Ltd., Tokyo, Japan) was also used for ashing the plant samples. All glassware was immersed in a mixture of 6 M HNOBand 30% HzOz(9 + 1) overnight before use. Extractioin Procedure for Determination of Cd, Ni, Pb, Zn,Cu, Hg, Co, and Bi. After the sample was wet ashed, the heavy metals in a 50-mL conical beaker were dissolved in 5 mL of 1M HN03 followed by the addition of 35 mL of water and 10 mL of 1.0 M ammonium citrate. This solution was adjusted to pH 8.0 (by using a pH meter) with 20% ammonia water. After transferring the solution to a 100-mL glass separatory funnel, the conical beaker was washed with 10 mL of water which was also added to the separatory funnel. Three milliliters of 0.01 M HMA-HMDC and 1.0 mL of chloroform were added to the separatory funnel. The contents were shaken for 15 rnin with an autoshaker. Ten microliters of the chloroform layer was injected into the C 18 column described above. The peak area of each metal chelate was constant for at least 8 h following phase separation. HPLC Conditions for Determination of Heavy Metals in Plant Samples. Conditions were as follows: column, Cosmosil 5 C 18,4.6 i.d. x 250 mm, 40.0 & 0.1 "C; mobile phase, methanol/water/chloroform/O.Ol M HMA-HMDC 76/16.5/6/1.5; flow rate, 0.8 mL/min; injection volume, 10 pL; detection wavelength, 260 nm. The mobile phase reservoir was cooled in an ice bath as discussed in a previous work (16). Forty milliliters of methanol/ water/cliloroform (76/18/6) was made to flow through the column after use each day to remove any chelating agent (HMA-HMDC) from the HPLC apparatus. Wet Ashing Procedure for Citrus Leaves and Rice Flour. (A) Citrus Leaues (NBS,SRM-1572). The sample (250 mg) was placed in a 50-mL conical beaker, followed by the addition of 7 mL of "OB, and covered with a watch glass. After the beaker was allowed to stand overnight, the contents in the beaker were heated on a hot plate (100 "C 15 rnin 150 "C 15 rnin 200 "C 15 min) to the temperature of the hot plate. After the sample was cooled, 8 mL of HC104was added and the mixture was heated again at 200 "C until the solution became light yellow (about 1 h). The watch glass was removed and the acid evaporated to dryness at 150 "C. A white residue was obtained. Five milliliters of 1 M "OB was added to the beaker and the contents were heated at 150 "C for 1 min. The clear solution obtained was subjected to solvent extraction. A blank test was also carried out by the same procedure. ( B )Rice Flour (NBS, SRM-1568). The ashing procedure for the rice flour was essentially the same as that for the citrus leaves. That is, 5 mL of HN03-HC104 mixture (2 + 1)was added and
-
-+
-
the solution was allowed to stand overnight. The contents were heated (100 "C 15 min 150 "C 10 min). After cooling, 3 mL of HC104was added and the solution was heated again at 200 "C until the solution became light yellow (about 30 min). When the acid evaporated, a white residue was obtained. Working Curves. Blank. Five milliliters of 1M HN03 was added to a blank beaker containing a residue equal in amount to the acid used for each sample. Standard. To the blank beaker, 5 mL of 1 M HN03 and 5 mL of mixed metal standard solution (see Reagents) were added. Three blanks and three s h d a r d solutions were used to prepare working curves for the heavy metals since the working curves of HMDC chelates showed good straight lines, as reported previously (15).
The volume of the aqueous phase was made to equal to that of the sample solution and extraction was carried out by the same procedure. The linear calibration range of the HMDC chelates was about 1-1000 ppb as metals in 50 mL of aqueous solution (15). This concentration range corresponded to 0.2-200 pg/g (ppm) in biological samples for a 250-mg sample.
RESULTS AND DISCUSSION Extraction Behavior of Metal-HMDC Chelates. Some reports on the extraction of metal-HMDC chelates have been published (17-19). Busev et al. (17)reported the extraction of 14 heavy metals in chloroform but did not indicate the concentration of metals. Dornemann et al. (18)also reported the extraction of HMDC chelates using methyl isobutyl ketone and diisopropyl ketone (DIBK) as organic solvents. The parts-per-billion levels of Cd, Ni, Zn, Co, Mn, Cu (10 ppb) and P b (40 ppb) have been reported by Tao et al. (19). Unfortunately, they also used DIBK as the organic solvent. Thus the effects of the concentration of HMA-HMDC and shaking time on extraction of heavy metals in chloroform were investigated in a previous work (16). In the present work, the effect of pH on the extraction of heavy metals in chloroform was investigated a t pH 7.5-10.0. The concentrations of the heavy metals were 100 ppb for Cd, Zn, Cu, and Co, 200 ppb for Ni, Pb, and Bi, and 400 ppb for Hg. Under these conditions, all metals except for Zn were extracted completely. The peak area (height) of Zn chelate was inversely proportional to pH but was reproducible when the extraction was carried out at constant pH (7.5-9.5). Iron is sometimes present at high concentration levels in the biological samples. When the extraction was carried out from an acidic solution, some of the Fe(II1) was extracted in spite of the presence of citrate and the poor peak shapes of the Fe chelate overlapped with that of Co chelate. These results show pH 8-9 to be suitable, and we used pH 8.0 in our work. Effects of Eluent Composition on Elution Behavior of HMDC Chelates. Dithiocarbamates of Hg, Cu, Co, Ni, and Cd are relatively stable in the mobile phase of methanol-water; those of Bi, Pb, Zn, Mn, and Fe are unstable as described by Thorn and Ludwig (20) and Hulanicki (21). Drash et al. (22)reported the efficient separation of DDTC chelates of Cd, Ni, Cu, Hg, and Co, using a mobile phase of methanol-water-chloroform, and the addition of chloroform made the DDTC chelates stable. In a previous work (16), we used a complex mixture of ethanol-methanol-water-buffer solution (pH 7~5)-10-~M HMA-HMDC (43/32/13/12/3) as the mobile phase. The ethanol was required for separation of P b and Ni, and the buffer solution and HMA-HMDC for preventing on-column dissociation of certain chelates, However, this mobile phase (ethanol-methanol-water) caused high back pressure of the column, compared with that of methanol-water. With use of a longer column to obtain better resolution of peaks, this mobile phase was not suitable. Thus, methanol-water-chloroform (22)was tested for the separation of HMDC chelates of Cd, Ni, Pb, Zn, Cu, Hg, Co,
ANALYTICAL CHEMISTRY, VOL. 57, NO. 12, OCTOBER 1985
2221
Table I. Analytical Results of Heavy Metals in Citrus Leaves and Rice Flour
samplea
0.5 0
2
a
6
4
10
Content of CHCl3 ('1.)
Flgure 1. Effect of chloroform content on elution behavior of metal HMDC chelates: D.P., peak of decomposition product of HMA-HMDC; mobile phase, methanol/water/chloroform/O.Ol M HMA-HMDC 82 x/16.5/x/1.5(x,content % of CHCI,). The other conditions are glven in the text.
I
0
10
I
I
I
20
30
40
T I M E ( rnin)
Flgure 2. Typical chromatogram of metal HMDC chelates (metal amount injected, ng): Cd (50),Ni (loo),Pb (loo),Zn (50), Cu (50),Hg (200),Co (50),Bi (100). Asterisk indicates peak of decomposition product of HMA-HMDC. Extraction and HPLC conditions are given in the text.
and Bi. The Zn and Bi chelates did not appear on the chromatogram owing to on-column dissociation. Secondly, a methanol-water401 M HMA-HMDC mixture was tested. The Pb peak overlapped with the Ni peak, and peak shape of Zn was poor (tailing). Thus, neither chloroform nor HMA-HMDC could prevent singly on-column dissociation of some HMDC chelates. Methanol-water-chloroform401 M HMA-HMDC was then examined as a mobile phase. The content of HMA-HMDC was 04.0% and that of chloroform, 0-10% (Figure 1). When a mobile phase of methanol-water-chloroform-0.01 M HMA-HMDC (76/16.5/6/1.5) was used, all chelates were separated completely (Figure 2). The peak shape and peak area reproducibility were good. From these results and Figure 1,chloroform appears usable as a stabilizer of some HMDC chelates and is effective for the separation of HMDC chelates. The maximum absorption wavelength of each HMDC chelate was in the range of 246-278 nm (16). Although the addition of HMA-HMDC in the mobile phase was required to prevent on-column dissociation of some chelates (particularly Zn and Bi), a high concentration of HMA-HMDC (more than 2%) made chelates detection difficult at around 260 nm. Thus, the above eluent composition (76/16.5/6/1.5) was selected and used for further work. Mn and Fe chelates dissociated on the column even when this mobile phase was used. Mn chelate was unstable even in chloroform. The column back pressure was reduced, compared with a previous work (16). Since a long column (5 Km particle, 4.6 i.d. X 250 mm) was used in this study to obtain better resolution, the column back pressure was still high at room temperature. The column temperature was thus maintained at 40.0 h 0.1 "C,since the column back pressure increased with mobile phase viscosity. This temperature control resulted in stable base line and good retention time and peak area reproducibility. A typical chromatogram of eight metal-HMDC chelates is given in Figure 2. The decomposition product was probably
citrus leaves (W.A.) rice (W.A.) flour (P.R.)
metal
amt found,b coeff of pg/g variation, %
certified values, pg/g
Ni Pb Zn Cu
0.83 i 0.19 13.2 f 0.5 29.7 f 0.5 15.9 f 0.2
23 3.8 1.2
0.6 f 0.3 13.3 f 2.4 29 f 2 16.5 f 1.0
Zn Cu Zn Cu
19.5 f 0.6 1.95 f 0.09 19.5 f 0.5 1.87 f 0.11
3.1 4.6 2.6 5.9
19.4 2.2 19.4 2.2
1.7
f 1.0 f 0.3 f 1.0 f 0.3
"W.A., wet ashing; P.R., plasma reactor. bAverage value f standard deviation (based on six replicate determination including ashing, extraction, and HPLC steps).
(HMDC)2 formed from free HMA-HMDC in the aqueous phase during extraction. Ashing of Plant Samples. Dry ashing with a muffle furnace was tested at first. However, recovery (accuracy) of Zn was not satisfactory in the range of 400-800 "C. Secondly, a plasma reactor was used (one chamber; 150 W; oxygen, 50 mL/min). Rice flour was ashed successfully but not citrus leaves. Wet ashing was finally investigated with hydrogen peroxide, nitric acid, sulfuric acid, and perchloric acid. The sample amount in each case was 250 mg. In the AAS method, an ashed sample solution obtained by wet ashing procedure is sometimes subjected to analysis directly. However, the present HPLC method requires solvent extraction at pH 8.0 before HPLC analysis. The effects of the above acids and hydrogen peroxide on the extraction of HMDC chelates were thus investigated. It was found that 5 mL of nitric, sulfuric, and perchloric acids reduced the extraction percent of some heavy metals (particularly Zn and Co) and a small amount of hydrogen peroxide interfered greatly with the extraction of all eight metals. Hydrogen peroxide probably decomposes (oxidizes) HMA-HMDC. Different concentrations of the acids changed the peak areas (heights) of the metal chelates. Recovery of chloroform after extraction varied with the chloroform amount dissolved in water (aqueous phase), when extraction was performed at a high ratio of aqueous to organic phase (about 65/1) in this work. Thus, concentration of chelates varied with chloroform recovery since it was effected by volume and temperature of aqueous phase. The amount of ammonia water neccessary for pH adjustment changed with acid concentration in the aqueous phase. Consequently, the peak area (height) of the chelates changed with volume and temperature (heat of nutralization) of the aqueous phase. The addition of hydrogen peroxide caused vigorous boiling and sulfuric acid could not be evaporated under the conditions used. Thus, nitric acid and perchloric acid may be suitable to wet ashing for HPLC analysis with HMDC extraction, but the acids used must be evaporated. When only nitric acid was used, the ashing required a long time and large amount of acid. Thus, perchloric acid was also used. The ashing temperature and amount of acid were investigated. Successful ashing procedures were given in the ashing procedure section (see Experimental Section). Use of a greater amount of acid made the evaporation time long. In the ashing procedure for rice flour, nitric acid could be used instead of a mixture of HN03-HC104, but ashing with a mixture of acids was faster than when using only nitric acid. Determination of H e a v y Metals in C i t r u s Leaves and Rice Flour. Analytical results of the plant sample are summarized in Table I. Ni, Pb, Zn, and Co chelates were sepa-
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 12, OCTOBER 1985
of heavy metals in biological samples.
Zn
ACKNOWLEDGMENT The authors appreciate the assistance of M. Konishi for preparing the column and K. Nakata for investigating the eluent composition. We also wish to thank K. Matsumoto (Kanazawa University) for his advice in carrying out the wet ashing. Registry No. HMDC, 874-56-6;Ni, 7440-02-0;Pb, 7439-92-1; Zn, 7440-66-6; Cu, 7440-50-8; Cd, 7440-43-9; Hg, 7439-97-6; Co, 7440-48-4; Bi, 7440-69-9; chloroform, 67-66-3.
cu
!
Pb
LITERATURE CITED I
Zn
I, 0
5
I
10
I
15 TIME (min)
I
I
20
25
Flgure 3. Analytical chromatogram of citrus leaves (A) and rice flour (B): (---) blank chromatogram; *, peak of decomposition product of HMA-HMDC. Conditlons of ashing, extraction, and HPLC are given in the text.
rated completely (Figure 3). Analytical results with a plasma reactor are also given in Table I. The analytical results are in good agreement with the certified values of NBS and satisfactory relative standard deviations were obtained. Cd, Hg, Co, and Bi in citrus leaves and Cd, Ni, Pb, Hg, Co, and Bi in rice flour could not be determined because of their very low concentrations (less than 0.16 ppm). Had these metals been present in detectable amounts, they could have been separated and determined accurately. Our results show that the present HPLC method with HMA-HMDC extraction after wet ashing is very useful for the simultaneous determination of the parts-per-million levels
(1) Wiiieford, Bennet R.; Veenlng, Hans J . Chromatogr. 1983, 251, 61-88. (2) O’Laughiin, Jerome W. J . Llq. Chromatogr. 1984, 7 , 127-204. (3) Nickless, 0. J. Chromatogr. 1985, 313, 129-159. (4) Bond, A. M.; Wallace, G. G. J . Liq. Chromatogr. 1983, 6 , 1799-1822. (5) Edward-Inatlmi, E. B.; Dalzlel, J. A. W. Anal. R o c . (London) 1980, 17, 40-42. (6) Cassldy, Richard M.; Eichuck, Steven J. Liq. Chromatogr. 1981, 4 , 379-398. (7) Edward-Inatlml E. B. J . Chromatogr. 1983, 256, 253-286. (8) Smith, Roger M.; Yankey, Lawrence E. Analyst (London) 1982, 107, 744-748. (9) Bond, A. M.; Wallace, 0. 0. Anal. Chem. lW4, 56, 2085-2090. (IO) Bond, A. M.; Wallace, G. G. Anal. Chim. Acta 1984, 164, 223-232. (11) Smith, Roger M.; Butt, Arif M.; Thakur, Arun Analyst (London) 1985, 110 35-37. (12) Yamazaki, Mltsuru; Ichlnoki, Susumu; Igarashi, Rleko Bunseki Kagaku 1981, 3 0 , 40-44. (13) Ichlnoki, Susumu; Yamazakl, Mltsuru Bunseki Kagaku 1982, 31, E319-E326. (14) Ichlnokl, Susumu; Yamazakl, Mltsuru; Morlta, Toshlhiro Bunsekl Kaga kU 1983, 32, 285-287. (15) Ichlnoki, Susumu; Morlta, Toshihlro; Yamazaki, Mltsuru J. Liq. Chromatogr. 1983, 6.2079-2093. (16) Ichinoki, Susumu; Morita, Toshlhlro; Yamazaki, Mltsuru J . Llq. Chromatogr. 1984, 7 , 2467-2482. (17) Busev. A. I.; Byr’ko, V. M.; Tereshenko, A. P.;Novlkova, N. N.; Naldina, V. P.; Terent’ev, P. B. Zh. Anal. Khim. 1970, 2 5 , 665-669. (18) Dornemann, A.; Kleist, H. Analyst (London) 1979, 104, 1030-1036. (19) Tao, Hlroakl; Mlyazakl, Aklra; Bansho, Kenji; Umezakl, Yoshlml Anal. Chlm. Acta 1984, 156, 159-168. (20) Thorn, G. D.; Ludwig, R. A. “The Dithiocarbamates and Related Compounds”; Elsevler: Amsterdam, 1962. (21) Huianickl, Adam Talanta 1887, 14, 1371-1392. (22) Drasch, Gustav; Kauert, Gerold; Meyer, Ludwig von Trace Elem. Anal. Chem. Med. Blol. 1983, 2 , 1109-1117. I
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RECEIVED for review March 5,1985. Accepted May 21,1985.
