1839
Anal. Chem. 1983, 55, 1839-1840
flections apparent. The column was wasshedovernight with EDTA solution followed by DMF. The chromatograph in Figure 2B demonstrates5 that aliquots of the sample injected into this washed column now show improved, sharper peak shape as well as increased resolution. We have found that DMF also helps at times to improve column back pressure, probably by removing lipid substances trapped on the column. It is possible to incorporate EDTA (up to 0.05%) into the mobile phase used for the chromatography of leukotrienes. This continual presence of a chelator in the mobile phase has helped to improve and istabilize the chromatography of leukotrienes, especially those containing sulfur, possibly by removing metal cations trapped in the stationary phase. However, care must be exercised when using EDTA in this manner due to its very llow solubility in organic solvents such as methanol and acetonlitrile. This is especially critical when using gradient elution reverse-phase HPLC. Registry No. EDTA, 60-00-4; DMF, 68-12-2; EDTAqZNa, 139-33-3;LTC4, 72025-60-6; LTB4, 71160-24-2;A6-trans-LTB4,
71652-82-9;A, 86161-62-8;B, 86161-63-9.
LITERATURE CITED (1) Lewis, R. A,; Austen, K. F. Nature (London) 1981, 293, 103-105. (2) Ford-Hutchinuon, A. W.; Bray, M. A,; Borg, M. V.; Shipley, M. E.; Smith, M. J. H. Nature (London) 1980, 286, 264-265. (3) Murphy, R. C.; Hammarstrom, S.;Sammuelsson, B. Proc. Natl. Acad. Sci. U . S . A . 1979, 7 6 , 4275-4279. (4) Mathews, W. R.; Rokach, J.; Murphy, R. C. Anal. Biochem. 1981, 118, 96-101. (5) Murphy, R. C.; Mathews, W. R. "Methods In Enzymology"; Lands, W. E. M., Smith, W. L., Eds.; Academic Press: New York. 1982; Vol. 86, pp 91-96. (6) Peters, D. G.; Hayes, J. M.; Hieftje, G. M. "Chemical Separations and Measurements"; W. B. Saunders: Philadelphia, PA, 1974; Chapter 6. (7) Snyder, L. R.; Kirkland, J. J. "Introduction to Modern Liquid Chromatography"; Wlley: New York, 1979; pp 266-267.
RECfor review January 17,1983. Accepted May 16,1983. This work was supported in part through grants from the National Institutes of Health (HL 25785 and NS 09919), a fellowship for J.Y.W. (AA 05193), and Boehringer Ingelheim Ltd. (U.S.A.).
Preparation of n-Alkyl Trichloroacetates and Their Use as Retention Index Standards in Gas Chromat ogriaphy Ted R. Schwartz* and Jimmie D. Petty
U S . Department of the Interior, Fish a,nd Wildlife Service, Columbia National Fisheries Research Laboratory, Route 1, Columbia, Missouri 65201 Edwin M. Kaiser Department of Chemistry, University of Missouri-Columbia,
Columbia, Missouri 65201
High-resolution gas chromatography has proved to be an effective means of quantifying single components in the presence of complex mibrtures (1-3). When this analytical technique is used in conjunction with the Kovats or other retention index systems (4,5),data generabed in any laboratory will have a common reference and can be directly compared. An integral part of the llovats retention index system is a series of homologous reference compounds employed as the retention index standards, the most common being normal alkanes used with flame ionization detection. Although the normal alkanes are the most widely used, their practical application to most environmental analyses cannot be realized due to the limited response of the electron capture detector (ECD) to the normal alkanes. Other homologous series have been proposed as substitutes for the n-alkanes in a retention index system that would be compatible with the ECD. Among these are the n-bromoalkames (6) and n-alkyl trichloroacetates (7). In a study of the distribution of polychlorinated biphenyl (PCB) isomers in environmental samples, the n-alkyl trichloroacetates were selected as the retention index standards, because of both their thermal stability amd their adequate response in an ECD at picogram concentrations. The utility of n-alkyl trichloroacetate for retention indexing was first recognized by Neu and co-workers (7) and they have been used for such purposes in the gas chromatographic analysis of PCBs (8,9). Unfortunately, the synthetic route for obtaining these compounds has not previously been described in the chemical
literature. Thus the synthesis and purification of such n-alkyl trichloroacetates are described below. The procedure involves adding trichloroacetyl chloride to the alcohol in the presence of pyridine and warming the mixture overnight to produce the corresponding esters (4545%). Attempts were not made to maximize yields. This procedure appears to be generally applicable to the synthesis of these types of esters.
0003-2700/63/0355- 1639$01.50/0
C13CCOC1 + ROH
C6HsN
C13CC02R + HCl
EXPERIMENTAL SECTION A slight excess of trichloroacetyl chloride was slowly added through a dropping funnel to the desired alcohol (all chemicals were obtained from Aldrich Chemical Co., Milwaukee, WI) and 0.05 mL of pyridine in a round-bottom flask fitted with a reflux condenser. The reaction mixture was gently warmed (90 O C ) overnight in a fume hood allowing HC1 gas to escape. The amber-yellow crude product was transferred to a separatory funnel and washed several times with distilled water until the washings had a pH of 7. The crude product (5 mL) was transferred to the top of a 13 mm i.d. X 600 mm silica gel column (EM-60 70-230 mesh ASTM from E. Merck, Darmstadt, Germany) that had been activated at 130 'C overnight. The sample was allowed to drain into a 2 cm bed of' Na2S04above the silica gel. Elution with 550 mL of 2% methyl tert-butyl ether in hexane was begun. The first 50 mL was discarded and the remainder was collected in a round-bottom flask. Solvent was removed by rotary evaporation. Compound purity was determined on a gas chromatograph. 0 1983 American Chemical Society
1840
.
ANALYTICAL CHEMISTRY, VOL 55, NO. 11, SEPTEMBER 1983
chromatographed. The mass spectra of these compounds in either electron impact or negative chemical ionization did not exhibit a molecular ion but rather a fragmentation indicative of the loss of C1. All 12 esters exhibited the same major IR cm-* 2938 (CH); 1743 (C=O ester); peaks. IR (neat): ,,v 1242 (C-0-C); 828 and 678 (C-Cl). The utilitity of these compounds as retention index standards is demonstrated in Figure 1, where the n-alkyl trichloroacetates were first chromatographed as a mixture and subsequently with an Aroclor (polychlorinated biphenyl) mixture. In summary, although the alkyl trichloroacetates have been employed as retention index compounds, their systhesis has not previously been described in the chemical literature. The procedure described here is simple and materials are relatively inexpensive and commercially available. The response of the ECD to these compounds makes them an ideal retention index standard.
A
1
L 10
ACKNOWLEDGMENT The authors thank James L. Johnson for his assistance in GC/MS analyses.
id---20
30
IC_L
40
60
L
eo
ELUTION TIME, MIN
Figure 1. n-Alkyl trichloroacetates added to a mixture of Aroclor standards (A). n-Alkyl trichloroacetates chromatographed as a standard mixture (B). The concentrations of the n -alkyl trichloro-
acetates (ng/FL)are as follows: nonyl, 0.15; decyl, 0.13; undecyl, 0.15; dodecyl, 0.15; tridecyl, 0.15; tetradecyl, 0.34; pentadecyl, 0.51; hexadecyl, 1.14; octadecyl, 4.4. the gas chromatograph (Varian Model 3700, Varian Instrument, Sunnyvale, CA) was equipped with a e3Ni ECD and a 0.25 mm i.d. X 30 m glass capillary column coated with Apolane-87 (C-87) (QuadrexCrop., New Haven, CT). Hydrogen, linear velocity of 32 cmlmin, was used as the carrier gas, and nitrogen delivered at 15 mL/min was used as the detector makeup gas. The inlector and detector temperatures were 220 OC and 300 OC, respectively. After injection the oven temperature was programmed from 120 O C to 250 O C at 2 OC/min.
Registry No. Cl3CC0C1, 76-02-8; C1,CC02R (R = nonyl), 65611-32-7; C13CC02R(R = decyl), 65611-33-8; C1,CCO2R (R = undecyl), 74339-49-4; C13CC02R (R = dodecyl), 74339-50-7; C13CC02R(R = tridecyl),74339-51-8;C13CC02R(R = tetradecyl), 74339-52-9; C13CC02R(R = pentadecyl), 74339-53-0; C13CC02R (R = hexadecyl), 74339-54-1;C13CC02R(R = octadecyl), 3542517-3;nonanol, 143-08-8;decanol, 112-30-1;undecanol, 112-42-5; dodecanol, 112-53-8;tridecanol, 112-70-9;tetradecanol, 112-72-1; pentadecanol, 629-76-5; hexadecanol, 36653-82-4; octadecanol, 112-92-5.
LITERATURE CITED (1) Zell, M.; Neu, H. J.; Ballschmiter, K. Z . Anal. Chem. 1978, 292,97. (2) Rlbick, M. A.; Dubay, G. R.; Petty, J. D.; Stalling, D. L.; Schmitt, C. J. Environ. Sci. Techno/. 1982, 76, 310. (3) Vassilaros, D. L.; Stoker, P. W.; Booth, P. W.; Lee, M. L. Anal. Chem. 1982, 5 4 , 106. (4) Kovats, E. Helv. Chim. Acta 1858, 4 7 , 1915. (5) vanDen Dool, H.; Kratz, P. D. J . Chromafogr. 1963, 7 7 , 463. (6) Pacholec, F.; Poole, C. F. Anal. Chem. 1982, 5 4 , 1019. (7) Neu, H. J.; Zell, M.; Ballschmlter, K. Z . Anal. Chem. 1978, 293, 192. (8) Ballschmiter, K.; Zell, M. Z . Anal. Chem. 1980, 302, 20. (9) Ballschmlter, K.; Unglert, Ch.; Nev, H. J. Chemosphere 1977, 6 , 51.
Confirmation of structural identity was accomplished by infrared analysis and mass spectrometry.
RESULTS AND DISCUSSION A high degree of purity was demonstrated by the presence of only one major peak when the esters were individually
RECEIVED for review April 11, 1983. Accepted June 9, 1983. References to trade names do not imply government endorsement of commercial products.
