An a Iys is of Pesticide Resid ues Using Mic roc0uIometric Tempera tu re-Programmed Gas Chroma tog ra phy W. A. BOSlN Research and Development Department, The Pillsbury
b The principles developed for the microcoulometric gas chromatographic analysis of pesticide residues have been incorporated intcl modified equipment. The major intent of this modification was to acquire greater flexibility in column conditions-namely, temperature programming. A secondary advantage is that a dual detector system is provided for the gas chromatograph. The column effluents may b e directed through either a microcoulometric cell or a thermal conductivity detector. Therefore, this instrument provides improved utilization of the microcoulometric detecior for pesticide residue analysis in addition to broad applications of gas chromatographic analysis. The addition of temperature programming gives better resolution and greater sensitivity for multicon;ponent pesticide residue mixtures thar isothermal operation. A new colurnn also achieves these advantages, The advantages are illustrated b y chr'3matograms derived from the analysis of pesticide mixtures under both tsemperafure programming and isothermal conditions.
A
and universally applicable technique for the analysis of all the halogenated or sulfur-containing pesticides wa? desired for application to a wide variety of samrlles of unknown spray history. Utilization of the commercially available microcoulometric gas chromatograph ( 5 , 8 )appeared to be the best approach. However, thi, equipment 11 ould have more extended application if its oper.tting parametera rould be made more versatile. Specifically the ability to temperature-program the gas chromatographic column would be a definite athantage. Temperature programming would improve the qualitative specificity of this technique through increa-ed resolution in of a multicomponent peaticaide re4due mixture. Quantitative vniitivity would al-o be enhanced berai1.e of the incremed peak sharpness for pchticides having longer retention time,. Therefore, conimercially available in5truments were modified to in(-liidetemperature programnnnq of the gn- c.lirornntogr:i1,Ilic coluniri, RAPID
Co., Minneapolis 7 4,
Minn.
I
TOP VIEW
r
k
TERMINAL
I
STRIP
WITH COVER REMOVED
17"
INJECTION +LL=K
%q
2"
+3" + + 2"
m
t
INSULATING
2"
XIT
(DRAWN TO SCALE1
Figure 1 .
Modified detector assembly
APPARATUS
Gas Chromatograph. A Model 500-A linear programmed temperature gas chromatograph (F&M Scientific Corp., Avondale, Pa.) was used t o separate the various pesticides through temperature programming. T h e detector assembly was modified and enlarged to include a special aluminum block through which t h e column effluents can be passed. Figure 1 shons the position of the original thermal conductivity detector and the special aluminum block. The effluents leaving the column through the column evit block are directed through either the thermal condueti1 ity detector or the aluminum block by simple interchange of connecting tubing at the column exit block. Initially a two-way valve n as used to direct the column effluents through either dctector, but this developed leaks a t high temperature operation. However, it is believed that this valve deficiency can be corrected. The special aluminum block (Figure 2) was acquired from Dohrmann In5triiments Co , Palo .ilto, Calif.; its dcsign i- similar to that used in the microcoillometric ga\ chromatograph ( 5 ) . The column efflucnts enter this block and a second carrier gas qul)ply *\I w.yis them through a platinum tuhc into a quartz combustion tube. The qiini t z rombustion t i i h (Dohimnnn
Instruments) contains a platinum gauze plug and is partially packed with 10to 20-mesh quartz chips ( 5 ) . It is connected to the aluminum block by a clamped ball and socket arrangement. The socket is a stainless steel fitting threaded into the aluminum block. Oxygen also enters this block, passes around the platinum tube, and a t its end mixes with the column effluents, thereby supporting combustion. The burned effluents pass into the microcoulometric cell, where the resulting hydrogen halide or sulfide is automatically titrated. The injection block assembly was modified t o include a removable quartz glass tube, as described by Cassil ( 2 ) (Figure 3). In addition to serving as a n easily cleaned trap for nonvolatile materials from uncleaned or cleaned samples, this tube also eliminates the degradation and partial loss of certain pesticides because of a probable dphalogenation reaction with the metal. Combustion Furnace. A Hoskins combustion furnace, Type FH-303-A, mas used. T h e supporting stand was removed and the furnace butted directly against the end of the detector assembly with a n insulating asbestos plate inserted between them (Figure 2 ) . T h e combustion furnace temperature is controlled by a per cent time-on input controller. The outer surface of t h e furnace has a copper VOL. 35, NO. 7,JUNE 1963
833
1 ,,/
/
I
l
I
COMBUSTION FURNACE ,INSULATING PLATE ,INSULATED BOX ,HEATING U N I T
,,/
l
1 I
t
c-- 2" -+
(DRAWN TO SCALE)
Figure 2.
Aluminum block
cooling coil to minimize heat transfer to the room's atmosphere. Microcoulometric Cell. The chloride-sensitive titration cell (Model T-100) and its associated coulometer (Model C-100) control were obtained from Dohrmann Instruments Co. The principles of operation are described in the literature (6). The titration cell was directly connected t o the exit end of the combustion tube by a clamped ball and socket fitting. Recorder. The recorder used is a Leeds and Northrup N o . 64101-51ClO-F5-Ql-R16-951 Speedomax Type G recorder for gas chromatography, equipped with a Model 203 disk chart integrator. This recorder has a variable span (1 to 50 niv.) and a zero offset adjustment, which are desirable characteristics for this operation. Although the microcoulometric cell as designed requires a 10-mv. recorder, addition of a resistor will permit satisfactory use of n l-mv. recorder for both the microcoulometric cell and the thermal conductivity detector. Column 1. A 191/2-inch length of S/le-inch 0.d. (0.144-inch i.d.) aluminum tubing was packed with 1.70 grams of 15'% Dow Corning 200 fluid (12,500 cs.) and 1.7y0 of Tween 80 (Atlas Chemical Co.) on 60- to 80-mesh Gas Chrom Z (Applied Science Laboratories, Inc.). The column packing was prepared by dissolving 5.0 grams of D C 200 fluid and 0.57 gram of Tween 80 in 150 ml. of chloroform; 28.0 grams of Gas Chrom 2 were added. The solution was placed in a shallow tray and stirred continuously as the chloroform was evaporated with an air stream. When the chloroform was completely removed, the 19'/2-inch length of aluminum tubing was packed, using vacuum. The tubing was tapped along its sides until all of the 1.70 grams of packing was within the tube. The ends of the tube were plugged with glass wool. Column 2. The second column, used for comparison purposes only, consisted of a &foot length of 1/4-inch 0.d. aluminum tubing packed with 17.5% Dow Corning high vacuum sili-
834
ANALYTICAL CHEMISTRY
cone grease on 30- to 60-mesh acidmashe8 Chromosorb P. This column has been used for much of the previous work on the gas chromatographic analysis of pesticide residues. A procedure for its preparation is found in the literature ( 7 ) . RESULTS A N D DISCUSSION
Numerous variations in column parameters and operating conditions were investigated for the most efficient separation of the many pesticides through temperature programming of the column. After unsatisfactory results with other liquid phases, all of the following studies utilized Dow Corning 200 fluid (12,500 cs.) as suggested by Smith ( 0 ) . An additional suggestion (8) about the advantages in separating pesticides through the use of Tween 80 in combination with silicone greases or fluids, started investigations which resulted in the column recommended here. The advantages derived from
combinations of nonpolar and polar liquid phases have been demonstrated by Averill (1) and they also apply to this column. Completely desirable separations were not attained until a l/dinch 0.d. (0.154-inch i.d.) glass column was used instead of the usual '/&-inch0.d. aluminum tubing. However, it was not possible to make the glass-to-metal fitting completely leak-free, and low recoveries for pesticide standards resulted. Since the desirable increased resolution resulting from the glass column was related to its smaller inside diameter, aluminum tubing of 2/jla-inch0.d. (0.144-inch i.d.) was employed. This column, packed as described above (column l), provides a very satisfactory separation for the pesticides studied. The most efficient operating conditions for Column 1 were isothermal operation a t 150' C. for the first 7 minutes after sample injection and then programming at the rate of 2.1' C. per minute to a maximum temperature of 235' C. The isothermal operation a t the start adequately separates the early eluting pesticides with a sufficient time spread between the elution of lindane and aldrin. Programming a t this rate produces sharp peaks with good resolution for the late-eluting pesticides and, with the exception of CoRal, all of the pesticides studied elute within 45 minutes. If the maximum temperature of 235' C. is exceeded consistently, Tween 80 is lost by bleeding and the column efficiency is no longer satisfactory. The column has a long life, if operated under the conditions described above. Comparisons are made of three chromatograms resulting from a mixture of pesticide standards. Figure 4 illustrates the results obtained when a mixture of chlorinated pesticides (lindane, heptachlor, aldrin, dieldrin, per-
TO COLUMN
t
CONE R U B B E R G A S K E T
(DRAWN T O SCALE)
Figure 3.
Modified injection block
CHROMATOGRAM A
%FULL SCALE
PESTICIE
CHROMATOGRAM E
1
120
- - _0- _ _ _ -_ _ _ ___. iio
lb0
io
u9 PESTICIDE
Ypw
0
LINDANE
2.75
2 01
@ @
HEPTACHLOR
4.07
I 95
ALDRIN
2.72
157
@
DIELDRIN
3 78
2 09
@
PERTHANE
729
I69
@
~LDDT
8.15
4 08
@
TEDION
5.96
2 38
40
CHROMATOGRAM
C
- 20
Figure 4.
Column Helium flow, ml./min. Column temp., O C.
Mixture
Microcoulometric cell, M o d e l T-1 00 Resistance, 6 4 ohms Block temp., 2 5 0 ' C. Injection block temp., 28OoC. Recorder, 10 mv. a n d 0.5 inch/min. Chromatoaram A Chromatoaram B Is% D.c. 2 0 0 a n d 1.7% Tween 80-
130
130
60
1 7 0 isothermal
2 4 0 isothermal
%
FULL SCALE
- 80 -
60
- 80
A
(4).
Chromatogram A , Figure 4,shows the improved resolution and sensitivity gained by temperature programming with the column spctcially developed for this purpose. Comparison of chromatogram A with chromatogram C, which was acquired under the usual gas chromatographic conditions for pesticide residue an zlysis ( 7 ) , very effectively illustrates 1 hese advantages. Dieldrin and perthane are satisfactorily separated by temperature programming, but are not resolved in chromatogram C. Temperature pl-ogramming improved the over-all recolution of all the peqticides studied. Later eluting com-
Chromatogram C 1 7 . 5 7 0 D.c. high vacuum grease
Programmed
thane, p,p'-DDT, and Tedion) was analyzed under three different sets of conditions. Exactly the same quantity of pesticides was used for each chromatogram (Figure 4). The operating conditions used for each chromatogram are also listed in Figure 4. The chloride-sensitive microcoulometric titration cell (Model T-1013) has since been redesigned, increasing sensitivity eight to 12 times ( 3 ) . The cell was operated a t 64-ohm resistance, a setting indicative of its sensitivity. The operating principles of this cell have been published
Column Helium flow rate, ml./min. Column temp., O C. Injection block temp., C.
of chlorinated pesticides
-
60
-
40
-
80
- 60
AL - 4
4
---- - _ - _ _ - _ - _r ---_- - - - - - 70
Figure 5.
60
- 40 - 20 BASE
JO+TIME+45
Chromatograms of
295 pg.
of CoRal Microcaulometric cell, M o d e l T-1 00 Resistance, 6 4 ohms Block temp., 2 5 0 ' C. Recorder, 10 mv. a n d 0.5 inch/min.
Chromatogram A Chromatogram B 1 5 % D.c. 2 0 0 a n d 1.7% Tween 80 130 130 Programmed to 2 3 5 max. Programmed to 2 5 5 mox.
280
280
ponents have sharp symmetrical peaks providing a much higher degree of sensitivity for these components than shown in chromatogram C. The sensitivity to heptachlor under both sets of conditions is about equal, with temperature programming providing greater sensitivity to pesticides eluting later than heptachlor and slightly less sensitivity to those eluting before heptachlor. Only one peak is acquired for p,p'-DDT, indicating that no degradation occurs under these temperature-programming conditions. I n this respect, column 1 has never indicated D D T degradation and it is easily and rapidly prepared without additional processing of the solid support. Of the pesticides studied (Table I), only CoRal elutes later than Tedion. Thus, with temperature programming, most pesticides except CoRal elute within 45 minutes, the time a t which the 235' C. maximum temperature for best column life is reached. A comparison of chromatogram B of Figure 4, the 170' C. isothermal operation of the column used for chromatogram A , shows that this column also has advantages even when operated isothermally. Dieldrin and perthane are again clearly separated with reason-
17.5%
Chromatogram C D.c. high vacuum grease 60 2 4 0 isothermal
250
VOL. 35, NO. 7, JUNE 1963
835
CHROMATOGRAM A
% F U L L SCALE
- 60 - 20
PEAK PESTICIDE
0
Figure 6.
- 80 - 60
NO PESTICIDE
&
0
THIYET
1.28
0.46
@
DI SYSTON
1.16
0 . 4I
0
vc-I3
8.93
0.91
@ @ @ @ @
MALATHION
6.03
1.17
MITOX
6.25
0 74
OVEX
7 46
0 79
ETHION
2.54
0 85
EPN
7.50
0.74
Mixture of sulfur-containing pesticides Microcoulometric cell, M o d e l T-200-P Resistance, 64 ohms Block temp., 2 5 0 ' C. injection block temp., 280' C. Recorder, 10 mv. and 0.5 inch/min.
Column Helium flow rate, ml./min. Column temp., O C.
Chromatogram A Chromatogram 15% D.c. 200 and 1.75% Tween 80
130
Chromatogram C 17.5% D.c. high vacuum grease
B
60
130 170 isothermal
Programmed
240 isothermal
able sensitivity, as are heptachlor and aldrin. Later-eluting components show poor sensitivity, however, and CoRal did not elute before 3 l / 2 hours. The isothermal operation of column 1 shows definite advantages in resolution for components eluting before perthane, compared to the column of chromstogram C . Figure 5 indicates the peaks acquired from exactly the same quantity of CoRal under the three conditions as listed. For chromatogram A , column 1 was programmed to 235' C. maximum and held; for chromatogram B, column 1 was programmed to 255' C. maximum and held. Chromatogram C is of the usual 240' C. isothermal operation of column 2. Chromatograms A and B indicate the same or greater sensitivity for CoRal than chromatogram C, but with nonsymmetrical peaks, while the latter produced a symmetrical peak. Total time for elution under a n y of these three conditions is approximately the same. Figure 6 illustrates the results when a mixture of sulfur-containing pesticides (Thimet, Di-Syston, VC-13, hfalathion, hlitox, Ovex, Ethion, and EPK) was analyzed under the conditions used for Figure 4, b u t using the sulfur-sensitive microcoulometric cell instead of the chloride-sensitive cell. This cell, Nodel T-200-P, is of the new design (3) with increased sensitivity. The difference
836
ANALYTICAL CHEMISTRY
Table I.
Relative Retention Times of Chlorinated and/or Sulfur-Containing Pesticides
Pesticidr
13 etention tinip, min.
8ysto.c (1st peak):
0.28
'l'liinietfi bystox (2nd pe:i!i)" Isopropyl estri, 2,4-D Diazinono Lindane IX-Syston" VC-13b Isopropyl ester, 2,4,5-T Heptachlor Aldrin Methvl Darathionn Alalathihn" Parathiona Chlorothionb Thiodan (1st peak)h lllitoxb Dieldrin Endrin (1st peak) Butoxpethanol ester, 2,4-D DhfC Ethyl hexyl ester, 2,411 Ovexb o,p-I)DT Perthnne A4ramiteb
0.40 0 , 50 0 .i 1 ,
0.62 0 . GY 0 . GO 0.86 0.88
Pesticide Thiodan (2nd peak)* Endrin (2nd peak) Ethiona Chlorobenzilnte Ethyl hexyl ester, 2,4,5-T p,p'-DDT Trithionb Uethoxychlor
EPX.
0.90 1 .00 1.17 1.26 1 .:34 1.42
Tedionb CoRalb Chlortinn
1.88
Tosnplienr
1 .fLj
1.88 1 Rti 1 98 2.02
Others rontnin oniy c.hIoritie.
Retrntion time, min. 2.04 2.10 2.12 2.17 2.23 2.26 2.28 2 i0 2 T8 2.92 3,80 Continuuni with peaks indicated at, 0.43, 0.72, 0.83, 1.03, l . I $ l j 1.28, 1.41, l.qN,l . G , and 1.92 Continuum with peaks indicated n t 1.63, 1.87, 2.08, and 2.28 Continuum with peaks indicated a t 1.6.5, 1.77, 1.90, 2.09, 2.24, 2.31, 2.35, 2.52, and 2.61
iii sciisitivity betwec !I the chloridcsensitive cell of origina.1 design and the newer model sulfur-sensitive eel1 is seen by comparing t.ie chloride and sulfur quantities indicated for the respective peaks on Figures 4 and 6. The same conclusion: can be drawn from Figure 6-that temperature programming with this improved column provides increased resolution and greater sensitivit’y for pesticide analysis than was previously attainable by isothermal conditions. Table I lists the r’dat’ive retention times of the pesticide3 studied. This list includes chlorinated pesticides, sulfur-containing pesticides, and some pesticides that contain both chlorine and sulfur. All retention times are calculated relative to aldrin as unity. The relative retention times of pesticides that contain wlfur only were related to aldrin by the use of hlitox (RZ‘ = 1.56) as an intermediate standard. These values art: specific to col-
1 uiider tlie conditions of temperature programming described.
uniii
ACKNOWLEDGEMENT
The author thanks the folloniiig for their advice and cooperation in this project: R. L. Ferin of The Pilltbury Co., L. A. Cavanagh of Stanford Research Institute, Dohrmann Instruments Co., F and 1I Scientific Corp., and the PilLbury Mechanical Development Department. LITERATURE CITED
(1) Averill, W., “Columns with ;2Iinimuin Liquid Phase Concentration for Use in Gas-Liquid Chromatography,’’ ISA International Gas Chromatography S j m posium, Michigan State University, East Lansing, Mich., June 1961. (2) Cassil, C. C., Stanford Research Institute, Pesticide Res. Bull. 1, No. 1 (1961). ( 3 ) Cavanagh, L. A., Coulson, D. ll., RlcCarthy, E. A I . , Salas, L. J., iT7ilton, V., “Improvenients in Micmcoulometric Gas Chromatography,” Pitts-
burgh Conference on Analytical Chcmistry and Applied Spect,roscopy, Pittsburgh, Pa., &larch 1962. (4) Coulson, D. >I., Cavanagh, L. A., A s . 4 ~ .CHEW32, 1245 (1960). ( 5 ) Coulson, D. AI., Cavanagh, L. A., ~‘R.licrocouloinetric Iletection in Gas Chromatography,” Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Pittsburgh, Pa., February 1961. ( 6 ) Coulson, D. M., Cavanagh, I d . A., DeVries, J. E., Walther, Barbara, J . .4qr. Food Chern. 8, 399 (1960). ( 7 ) Coulson, 1). XI., IleVries, J. E., Walther, B. h.,“Pesticide Residues on Fresh Vegetables,” Stanford Research Institute, Menlo Park, Calif., Tech. X e p t . I V on SRI Project N o . S-2360, p. 11 (1960). (8) Scliwecke, W, RI., (kneral Mills, Inc., Minneapolis, lrinn., private communication. ( 9 ) Smith, E. D., A N ~ LC. m x 33, 1626 (1961j. RECEIVED for review October 3, 1962. Accepted March 15, 1963. Division of ;~griculturaland Food Chemistrj-, 142nd hIeet.ing, ACS, Atlantic City, S . J., September 1962.
Gas Chromatographic Behavior of CI-C4 Saturated Alcohols and Water on Polyethylene Glycol Substrate Effect of Solid Support Treatment PAUL URONE Department of Chemisrry, University o f Colorado, Boulder, Colo.
ROBERT L. PECSOK Department o f Chemistry, University o f Califarnia, 10s Angeles 24, Calif,
b Twelve columns coated with a p proximately 20% polyethylene glycol400 were used to study the effect of solid support treatment upon the gas chromatographic behavior of seven Cl-C4 saturated alcohols and water. The treatment affectgd the activity of the solute and resulted in large variations of specific reiention volumes, distribution coefficients, and heats of solution. Clausius-Clapeyron type plots showed linear iemperature dependence for all solutes on all columns. Relative retention volumes, hence relative activity coefficients, were largely independent of the treatment of the solid support a t the approximately 20% liquid phase loading studied. Log-log plots of specific retention volumes vs. the vapor pressures of the alcohols and water a t 70” C. for each column showed a remarkably repetitive pattern. The pattern:; showed a vertical displacement of th,3 lines due to increases or decreases of retention volumes caused b y the treatment or subcoating of the support particles and
a parallelism of the lines which gives a higher degree of confidence that the relative activity coefficients are representative of the liquid substrate.
T
HE STDY OF POL‘4R solutes in gasliquid partition chromatography cannot proceed without a more intimate knowledge of the effect of the solid support upon the activity, the partition coefficient, and the retention volume of the solute. .!. relatively large number of studies concerned with solid support effects has becn reported (I, 2, 4-8, 10-20, 22, 23). Earlier 3tudies were generally concerned with the problem of eliminating tailing (,$, ‘7, is). However, it has become preponderantly apparent that the surface area and activity of the support, the amount of liquid phase loading, m d the sample size significantly contribute to tlie over-all gas clirorriatographic behavior of polar solutes. Scholz and Brandt (I?‘, 18) have observed clianges in rctcntiori voluiiies
with change> in sample size, injcction order, and amount of liquid phase coating Martin ( I $ ) has attempted to explain these anomalies by taking into account adsorption on the liquid phase surface. Keller and Stewart (11) have described gas-liquid partition chromatography as essentially a threephase system with contributions to thc resultant partition coefficient from all three phases. The purpose of this work was to study eyperimentally the effect of subcoating and treating various solid supports on the activity of low molecular weight alcohol.: in a polar liquid phase. Efforts were made to have the columns identical in all respects except the type of solid support surface. Diatomaceous earth (siliceous) supports of 60,430 mesh, coated with a conitant amount of polyethylene glycol-400, were used becausc~ of their relatively high surface area> ( 5 ) . The >upport materials n ere acid-\$ ashed firebrick, firebrick Iegular, Chromoiorb P, and Chromosorb It‘. 1rcatni~iitor siil,Loating of tlic sui)r 7
VOL. 35, NO. 7, JUNE 1963
837
