ANALYTICAL CHEMISTRY, VOL. 51, NO. 11, SEPTEMBER 1979
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If Equation 4 is plotted on log-log paper, the result is a series 5 0
1
r(
of straight lines intersecting a t a point (1,l)with slope of -n. On this plot, (1 - E ) decreases as KD increases. But as (1E ) decreases from 1to 0, its complement E is increasing from 0 to 1. Thus, E may be plotted as a linear function of [(l KD)(VE/ Vs)] on the inverted log scale, but this plot is awkward because it is in a negative quadrant, Le., E increases ad KD increases but the slopes are negative. This relationship may, however, be transformed to positive coordinates by reflecting through the plane where E = 0. With this transformation, one obtains the set of lines with slope of +n and intercept a t (l,O), (Note, however, that %E is plotted on the inverted log scale obtained by turning a sheet of loglog paper over from top to bottom.) This relation is then off-set by -1 so that the abscissa now becomes (KD)(VE/ V s ) . In this plot, a series of straight lines with slope of n (the number of extractions) relates the %E on an inverse log scale in the product of K,(VE/ Vs) on a log scale. With this plot, it is possible to: (1)make rapid comparisons between systems with known KD’s, (2) select volume ratios necessary to use with systems with known KD’s, (3) chose a system with an adequate KD to give a desired % recovery ( % E ) ,or (4)determine the number of steps necessary for “complete” (99%) extraction. In past practice, the analyst generally varied (VE/ Vs)and n but selected the extraction pair somewhat arbitrarily on the basis of experience. However, with the plot given, selection of extraction pairs and conditions can be made systematically from KD values.
+
30 20 10
0 0
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1
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DISTRI6LTI0\1 RATIO K D
Flgure 1. Graph of
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YO extracted vs. the distribution coefficient
selected on the basis of experience or semi-empirical considerations. Even workers who have measured distribution coefficients have sought alternate expressions for comparing different extractions. For example, Bowman and Beroza (2-4) have defined a “p-value” for several pesticide related extraction systems which in effect is the percent extracted under a fixed set of conditions. Suffet and Faust (6--8)have extended this p-value approach to calculate an (F,) value (total fraction extracted) as a more general term. These new terms were created because it is difficult to visualize the relationship between the distribution coefficient (ITD),the volume ratio (VE/Vs), and the number of extractions ( n ) on the percent efficiency ( % E ) when plotted in normal coordinates. The difficulty in relating the effect of extraction parameters on percent efficiency has been overcome by the derivation of the plot shown in Figure 1. The plot was derived in the following topological exercise. Equation 2 may be re-written in the form,
LITERATURE CITED (1) A. Leo, C. Hansch, and D. Elkins, Chem. Rev, 71, 525 (1971). (2) M. C. Bowman and M. Beroza, J . Assoc. Off. Anal. Chem., 48, 943 i,i a f i m “VI,.
(3) M. C. Bowman and M. Beroza. J . Assoc. Off. Anal. Chem., 48, 358 (1965). (4) M. C. Bowman and M. Beroza. Anal. Chem., 38, 1544 (1966). (5) V. G. Berezkin, A. G. Pankov, and V. D. Losbshilova, C:hromatographia, 9, 490 (1976). (6) I. H. Suffet and S. D.Faust, “Chapter 2, Fate of Organic Pesticides in the Aquatic Environment”, Adv. Chem. Ser., 111, 1972. (7) 1. H. Suffet, J . Agric. Food Chem., 21, 288 (1973). (8) I. H. Suffet, J . Agric. food Chem., 21. 591 (1973).
RECEIVED for review March 9,1979. Accepted May 10, 1979.
Sample Cleanup and Concentration Apparatus for the Determination of Chlorinated Hydrocarbon Residues in Environmental Samples John Solomon Department of Fisheries and Oceans, Freshwater Institute, 50 1 University Crescent, Winnipeg, Manitoba, Canada R3T 2N6
In toxicological studies of pesticides in the aquatic environment there is often a need to analyze large numbers of small animal tissue samples (fish organs, insects). Rapid methods of extraction of small samples for the determination of chlorinated pesticide residues using a specially designed 0003-2700/79/0351-1861$01.OO/O
ball-mill have been developed ( I , 2) but the cleanup and concentration of the extract for gas chromatographic (GLC) analysis has remained somewhat time-consuming. Recommended multiresidue procedures for cleanup of extracts containing chlorinated hydrocarbons (3,,4) use large quantities 1979 American Chemical Society
1882
ANALYTICAL CHEMISTRY, VOL. 51, NO. 11, SEPTEMBER 1979
Figure 3. Glass Filament Concentrator
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Figure 1. Six unit chromatographic and concentration apparatus ~
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Figure 4. Manifold head construction detail
EXPERIMENTAL
Figure 2. Chromarographic microcolumn construction detail
of chromatographic materials and solvents as well as requiring separate evaporation of the eluant and transfer to a test tube for GLC analysis. A six-unit apparatus has been designed (six microcolumns a n d six glass filament concentrators) t o permit the cleanup of six sample extracts and the simultaneous concentration of t h e eluting solvent.
Apparatus. The six-unit apparatus is shown in Figure 1. Individual components of the apparatus consist of: (I) Microcolumn (Figure 2). A small glass chromatographic column with a Luer fitting is fitted to a Teflon "mininert" syringe valve (Supelco) and a standard hypodermic needle. Column flow rates can be varied by use of different gauge needles ($26 X 10 mm is used routinely). The syringe valve is used to stop the elution so that the chromatographic column does not go dry. A permanent filter plug of glass wool topped with tissue paper is inserted at the base of the tube and is washed with acetone and hexane before use. The microcolumn is supported on top of the glass filament concentrator with a Teflon butt joint. (2) Glass Filament Concentrator (Figure 3). This consists of a jacketed glass filament (1-mm diameter) bonded (epoxy) to an aluminum head. The glass filament was hand-drawn from solid glass tubing. Helium enters through an inlet in the aluminum head at the top of the concentrator. The microcolumn eluate drops from the hypodermic needle to the top of the glass filament and flows through pinholes to the outside of the filament where the solvent is evaporated by the gas. Helium and solvent vapors escape through vent holes at the bottom of the concentrator. The sample concentrate collects in receiving tubes. The rate of flow of helium is adjusted with individual flow valves on the manifold head to which each concentrator is attached. The overall rate of concentration of the eluting solvent is controlled by the main metering valve on the helium flow interrupter. (3)Helium Flow Interrupter. This apparatus permits the flow of helium gas to be stopped for a short interval each minute to allow the residue on the glass filament to be washed into the receiving tube to reduce the evaporative losses of volatile compounds. It consists of an adjustable 1-min interval timer (Cramer, Old Saybrook, Conn.), a solenoid gas valve (Sporland Valve Co., tA3S1), a gas needle valve and an on/off switch all mounted in a metal chassis. The unit is mounted on the helium tank regulator. (4) Manifold Head. (Figure 4). The brass head has an inner and outer manifold. The outer manifold is connected to six needle
ANALYTICAL CHEMISTRY, VOL. 51, NO. 11, SEPTEMBER 1979
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Table I. Recoveriesa (%) of Ten Chlorinated Pesticides after Elution through the Cleanup-Concentrator Apparatus and Evaporation of the Eluting Solvent to Various Final Volumes quantity added, ng
pesticide HCB
1.25 2.50 5.00
oxychlordane p,p'-DDE p,p'-DDD p,p'-DDT
a
5.00 5.00
Final Volume, m L 0.23
0.60
0.70
2.0
2.1
35 83 89 98 94 86 79 100 98 100
54 102 98 90 99 89 81 98 10 5 10 2
76 101 99 102 105 89 90 100 100 94
69 98 92 94 87 88 89 96 92 97
89 92 95 93 98
93 94 94 95
lindane 1.25 aldrin 1.25 1.25 heptachlor epoxide cis-chlordane 1.25 dieldrin 1.25 One determination of each compound at each final volume.
Table 11. Recoveries and Relative Standard Deviations of 5 Chlorinated Pesticides after Concentration of t h e Column Eluate to 0 . 5 m L recovery,a rel. std. pesticide ngadded % dev., % HCB 0.38 57.3 21.1 87.3 6.1 oxychlordane 1.25 p ,p ' -DDE 1.25 95.4 3.7 p,p'-DDD p,p'-DDT
1.25
96.3
1.25 100.7 Average of 4 replicate analyses.
-
0.20
3.6 4.4
valves that are equdy spaced on the outer edge. The needle valves are connected to the helium inlets of the concentrator tubes via coiled Teflon tubing (2-mm 0.d.). The inner manifold is connected to six equally spaced syringe needles and to a vacuum source. The chromatographic packing in the microcolumns can be dried after use by attaching the columns to the syringe fittings with vacuum aspirator on. The manifold head is mounted on a laboratory stand. It can be rotated 360' with the receiving tubes supported on a wooden base plate. The concentrator tubes (spring-clip mounted to the manifold) can be moved up or down to permit the removal of the receiving tubes. A flow rotameter is mounted on the upright support, between the manifold and the base plate. Six slotted holes on the outside of the manifold are used for the storage of chromatographic columns. The apparatus is used in a fume hood. Application to Chlorinated Pesticide Analysis. The microcolumn is filled with 1.0 g Florisil (80/100 mesh, 5% water, w/w) with gentle tapping and topped with Na2S04(2-3 mm). The column is prewashed with hexane (4 mL elution with the vacuum line on the manifold). An aliquot of a hexane extract (0.5 mL) of tissue, is applied to the column with the syringe valve open. When the extract has completely entered the top of the column, 1.0 mL of hexane/diethyl ether (96:4) is added. The initial eluate (1.5mL) is discarded. Chlorinated pesticides are separated from lipid co-extractives by elution of the Florisil with 6.0 mL hexane/diethyl ether (96:4). A constant flow rate from the chromatographic column is acheived by use of a narrow-necked eluting vial (10-mL volume) which when filled with the eluant and inverted in the reservoir above the Florisil will maintain a constant head at the reservoir. The eluate is concentrated as it flows down the glass filament and is collected in the receiving tube. To study the efficiency of recovery of chlorinated hydrocarbons in the cleanup-concentrator apparatus, nanogram quantities of 10 compounds in hexane solution (0.50 mL) were chromatographed and concentrated to various final volumes. The concentrated eluates were analyzed with a Tracor Model 220 GC equipped with a linearized 63Nielectron-capture detector. The GLC column was 1.8 m X 6 mm 0.d. 5% OV-101 on Chromosorb W-HP (80-100 mesh). Carrier flow (N,) was 60 m l i m i n . Oven, detector, and inlet temperatures were 200, 350, and 230 "C, respectively.
-
102
-
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-
RESULTS AND D1SCUSSIO:N Table I gives the recoveries of low levels of 110 chlorinated pesticides after concentration of t h e Florisil column eluate to final volumes of from 0.2 to 2.1 m L (30- to 2.9-fold concentration). Different final volumes were achieved by altering the flow rate of t h e He with the main metering valve o n t h e gas flow interrupter. A 30-fold concentration of t h e eluting solvent had little effect ( < 5 % ) on t h e recoveries of p,p'-DDD, p,p'-DDT, dieldrin, cis-chlordane, or heptachlor epoxide. Losses of aldrin, lindane, oxychlordane, and p,p'-DDE at the final volume of 0.20 mL ranged from 21.5 to 11.5%. An increase in the final solvent volume t o 0.60 m L reduced t h e losses of these compounds to less than 10%. Table I1 shows the recoveries of 5 chlorinated pesticides a t 2- to 4-fold lower levels than those shown in Table I after concentration of t h e eluting solvent to 0.5 mL. Relative standard deviations were (10% except for HCB. Problems were encountered with evaporative losses of HCB because of the high volatility of this compound. The losses were reduced to 11% or less by collecting a separate sample for HCB analysis, which was concentrated only 3-fold. Since, HCB has a greater electron-capturing response than t h e other compounds that were studied, great concentration of t h e eluting solvent was not necessary for the analysis of 1.25 ng applied to the Florisil column, Application of larger quantities of chlorinated pesticides to the column (10-100 ng) also reduced volatilization losses of all compounds: including HCB, to less than 1 0 % . Sample cleanup and concentration was rapid. Six samples could be processed in 10 to 15 min. Considerable savings of solvents as well as reduction in health and fire hazards are also achieved when using the apparatus. Good cleanup could be achieved with sample extracts (0.5 mL aliquot of a hexane extract) containing up t o 15 mg lipids. T h e apparatus is relatively simple for a skilled glass blower and a machinist to make.
LITERATURE CITED (1) Grussendorf, 0. W.; McGinnis, A. J.; Solomon, J. J . A.jsoc. Off. Anal. Chem. 1970, 5 3 , 1048-1054. (2) Solomon, J.; Lockhart, W. L. J . Assoc. Off. Anal. Chem. 1977, 6 0 , 690-695. (3) "Pesticide Analytical Manual", Volume 'I. U.S. Food and Drug Administration, Washington, D.C., 1975. (4) "Analyfil Methods for Pesticide R e s a w in Foods", Canada Dept. N a h I Health and Welfare, Ottawa, 1973.
RECEIVED for review February 5,1979. .Accepted May 18,1979.
