851
V O L U M E 2 4 , NO. 5, M A Y 1 9 5 2 The expression used in calculating experimental hydrogenation values is:
H.V.
=
V p - p, 100 X ___ _ _ 10-3117 T
273.1 X 0.0898 X 760
-
where
V = volume absorbed, cc. IT’ = weight of sample, mg. 1‘ = absolute temperature p = pressure p , = partial presssure of solvent a t T o 0.0898 X 10-3 = specific gravity of hydrogen NTP Hydrogenation Rate Curves. Pure methyl oleate (6) was hydrogenated with W5 Raney nickel as catalyst ( 3 ) and 95% ethyl alcohol as solvent. Table I1 shows, in column one, the time of the readings (7’ minutes) and in column two, the concentration, C, of methyl oleate present a t the tixqe of the reading. This concentration is expressed by the volume of hydrogen still t o be absorbed. I n column three are indicated the values of the ratio In C,/C where
C, is the initial concentration-Le., the volume absorbed a t the completion of the reaction. The value of K = 1/T In CJC, shown in column four is comprised within very narrow limits, indicating that the reaction is of the first order in respect to the concentration of methyl oleate. The rate curve is this case is a straight line (rate us. C) passing through the origin (slope = K ) . The remarkable consistency of the K value is indicative of the precision attainable. LITERATURE CITED
( 1 ) Adams, R., Voorhees, V., and Shriner, R. L., “Organic Syntheses,” Collective Vol. I, p. 463, New York, John Wiley & Sons, 1948. (2) Gruen and Halden, 2. deut. OZ. u. Felt Ind., 44, 2 (1924). (3) Hadkins, H., and Billika, H. R., J . Am. Chem. Soc., 70, 695 (1948). (4) Kuhn, K., and Moller, E. F., 2. angew. Chem., 47, 145 (1934). (5) Manegold, Erich von, and Peters, F., KoZZoid Z., 85, 310 (1938). (6) Mattil, K. F., and Longenecker, H. E., Oil & Soap, 21,16 (1934).
( 7 ) Pregl, Fritz, “Quantitative Organic Microanalysis,” 3rd English ed.. p. 216, editor, H. Roth, London, J. & A. Churchill, Ltd.. 1937.
(8) Smith, H.A.,Fuzek, J. F., and hferiweather, H. T., J. Am. Chem. Soc., 71, 3765 (1949).
RECEIVED for review August 27, 1951. Accepted February 5 , 1952.
Microdetermination of Aldrin and Dieldrin by Infrared M. D. GARHART, F. J. WITMER, AND Y. A. TAJIMA Physical and Physico-chemical Laboratories, Julius Hyman & Co., Denver, Colo. Toxicological and field residue studies that were prerequisites to the marketing of the new insecticides, aldrin and dieldrin, necessitated the development of analytical micromethods applicable for such studies. Infrared spectrophotometry was chosen as the analytical tool most likely to have the necessary selectivity and sensitivity. Procedures were thus developed with sensitivities of 0.0005 and 0.0007% for aldrin and dieldrin, respectively, with a probable error of &O.OOl%. Accuracy was attained by refinements of usual infrared spectrophotometric techniques. Absorption microcells were used to attain sensitivity. By the use of these methods of analysis, and techniques for preparations of samples described elsewhere in the literature, it has been possible to carry out a large scale program of toxicological and field residue studies on aldrin and dieldrin.
T
HE new alkali-stable polychloro organic insecticides, aldrin (Compound 118) and dieldrin (Compound 497) were reported in 1949 by Lidov et al. ( 5 ) . Dieldrin is the epoxy derivative of aldrin (Figure 1). Aldrin is used here in reference to the active ingredient 1J2J3J4J10J10-hexachloro-1,4,4a,5,8,8a-hexahydro-lJ4,5,8-dimethanonaphthalene.Dieldrin is used in reference to the active ingredient 1,2,3,4,10,10-hexachloro-6,7-epoxy1,4,4a,5,6,7,8,8a-octahydro-1,4,5,8-dimethanonaphtha~ene. The widespread use of these insect toxicants in toxicological studies and field residue studies with sprays and dusts necessitated the development of sensitive and specificanalytical methods. A colorimetric method for the determination of aldrin, described by Danish and Lidov (3), involves the reaction of aldrin with phenyl azide to form aldrinphenyldihydrotriazole. The triazole is then coupled with diazotized 2,4-dinitroaniline, producing a highly colored product. The intensity of the color is determined a t 515 mp. An analytical method was required for dieldrin and a complementary method for the determination of aldrin was also desired. The infrared spectrophotometric methods described here were devised to fulfill these needs, and have been found sensitive t o 1 t o 2 micrograms of dieldrin or aldrin. The spectra of aldrin and dieldrin are shown in Figures 2 and
ALDRIN
DIELDRIN
Figure 1. Planar Representation of Aldrin and Dieldrin
3. The 8.48-micron absorption peak of aldrin and the 10.98 micron absorption peak of dieldrin were chosen for the determinations. It is possible that aldrin and dieldrin would interfere with each other if present in the same solution. Aldrin and dieldrin were not used together in the entomological and toxicological tests. The quantity of insecticide in a sample is related to the height (intensity) of the absorption peak. The essence of this project was the determination of this relationship with the
852
ANALYTICAL CHEMISTRY
added conditions of high sensitivity and an accuracy of the same order as the sensitivity.
use the 8.49-micron absorption peak of dieldrin, if aldrin were known to be absent. Toxaphene and Chlordan. The infrared absorption spect,ra of toxaphene and chlordan are reproduced in Figures 6 and 7. I t is apparent that these materials are not ell defined entities and are probably mixtures. Because of the nature of these pesticides, it would be difficult to predict how they would be affected by the sample preparation techniques and how they would affect the determination of aldrin and dieldrin if present in the final prepared sample. Neither toxaphene nor chlordan would be expected to exhibit spectral interference unless it was present, in the spectroscopic sample, in concentrations greatly in excess of aldrin or dieldrin. If the latter condition were to exist, one might still be able to use the 8.48- and 10.98-micron absorption peaks of aldrin and dieldrin, respectively, or if only toxaphene were present in large qumtity one might use the 12.02- or 12.93-micron absorption peaks of aldrin and the 11.82- or 12.37-micron absorption peaks of dieldrin.
SPECTRAL INTERFERENCES BY PESTICIDES OF CHLORINATED HYDROCARBOK TYPE
Possible spectral interferences by closely related pesticides such as chlordan or by any foreign impurities do not pose special problems unique to the infrared spectrophotometric methods here described for aldrin and dieldrin. Nost of the foreign matter and possibly other pesticides will be removed during the preparation of samples. If other pesticides are present in the final prepared sample, the usual techniques of spectrophotometry can be used to elininate errors that might be caused by spectral interferences. Aldrin and Dieldrin. If aldrin and dieldrin are present together in the final prepared sample, one can expect mutual interferences. However, it was known in the cases where the described methods were used that aldrin and dieldrin were not coesistent,. If these two pesticides are suspected of being present together in a spectroscopic sample, one can use, for example, the 12.93-micron absorption peak of aldrin and the 11.82-micron absorption peak of dieldrin and be assured that no mutual spectral interference will occur. DDT and Lindane. The infrared absorption spectra of these pesticides are reproduced in Figures 4 and 5. It is apparent that these materials are well defined compounds giving rather clean epectra. Only trial would determine whether the methods of sample preparation ( 2 ) would eliminate these pesticides. D D T present with aldrin or dieldrin in the spectroscopic sample would probably cause no difficulty. Lindane would not interfere with the determination of aldrin, but would interfere with the determination of dieldrin if i t coexisted in concentrations as high as or higher than dieldrin. In such a case, the 9.60-, 9.96-, or 12.37-micron absorption peak of dieldrin might be used. As a further example of the versatility of infrared spectrometric methods, one could
100
I
75
-
50
-
UI 25 v
I
APPARATUS
,
Perkin-Elmer Model 12-C infrared spectrometers were used. Sealed liquid cells with sodium chloride windows, also of Perkin-Elmer manufacture, were used in recording the spectra of aldrin and dieldrin. Perkin-Elmer S o . 12-099 microcells, of a design described by Colthup et al. (1 ), were used in the analytical procedures, The nominal thickness of these cells is 3 mm., with total volume of 0.05 ml. These microcells are provided with two openings. One opening consists of a hypodermic needle without the tapered female joint. The exposed beveled tip of the needle is curved back along a U-bend. The other opening is fitted with the tapered female joint of a hypodermic needle. Tuberculin syringes (0.25-ml.) were used to measure out, the predetermined volumes of solvent and also to transfer the sample solution into the microcell if the volume of the solution was 0.2 ml. or greater. If the volume of solution was less, it was necessary to use microsyringes, which may be obtained from the Micro-Metric Instrument Co. The microsyringes were obtained with the tips ground to fit the tapered filling holes of the microcells.
1
I
'
l,-0.025rnm Cell c CI4
-
-
Z 4:
E
-
Solvent Corbon Telrochloride Corbon D i s u l I i d e Cell- S e o l e d L i q u i d Sodium Chloride Concentration-
29.02% (.EM)
-
Instrument Perkin-Elmer
25
t
0 9.0
Model 12
COMPOUND "118"
I
I
I
I
I
95
100
105
I10
115
Figure 2.
I
I
I
I
130 135 WAVE LENGTH, MICRONS 120
125
I
140
,
I
145
150
155
160
Infrared 4bsorption Spectra of AIdrin (Compound 118) over Range 2.5 to 15 Microns
I
165
V O L U M E 2 4 , NO. 5, M A Y 1 9 5 2 100
I
15
-
50
-
25
-
z
I
1
853 - 1
I
I
I
1
I
I
I
I
I
i
\
5
5
100
I-
3
on Disulfide on Telrachloride
U
15
S e a l e d Liquid C e l l Sodium Chloride 0 . 0 2 5 mm.
L Y
50
lnslrumentP e r k i n - E l m e r Model 12C
C OM P 0 U N 0
25
"
497"
Sof'd. Sol'n In CC14
9
I
I
9.5
10.0
I
10.5
I
I
I
I
11.0
11.5
12.0
12.8
I
13.0
I
I
13.5
14.0
I
I
14.5
15.0
I
1
16.0
15.5
I
5
WAVE LENGTH, MICRONS
Figure 3.
Infrared Absorption Spectra of Dieldrin (Compound 497)
Carbon disulfide of analytical reagent grade was used ITithout further purification. However, each batch of solvent was checked t o make certain that extraIleous absorptions by the carbon disulfide were not presentat the same wave lengths as the absorption peaks of aldrin and dieldrin-that is, solvent only should analyze 0% concentration of aldrin or dieldrin.
were recorded in solution, usillg automatic wave length and slit drives, The absorption of aldrin at 8.48 microns and of dieldrin .at 10.98 microns was measured as a function of the concentration.
SPECTRA OF ALDRIN AhD DIELDRIN
Preparation of Sample. The extraction of aldrin or dieldrin and the subsequent treatment of the extract are described by Danish et a!. ( 2 ) .
PROCEDURE
The spectra of aldrin (Compound 118) and dieldrin (Compound -497) are shown in Figures 2 and 3. The spectra of the compounds
Y
I 110
I
I
140
I3 0
I
la0
II0
tZ I
I
I
I20 1
1 90
I SOLVENTC A W MSULFIOE CARBOW TETRAWILORIDE
_Li BO
ANALYTICAL CHEMISTRY
854
s
Y
m w
I
I
1
I
I
I
I
SOLVENTCARBON OLWCFIDE CARBON TETRACHLORIOL SEALED LlOUlD CELL SODIUM CHLORIDE 0.100 MY INSTRUY ENTR R K I N - E U E R YOOELILC COI(RWJI(0-LIIOIIL
SBX
PURITY-
L I I I L . so
7.0
4.0
1.0
I
SO
WAVE LENGTH, MICRONS
Figure 5.
Infrared Absorption Spectra of Lindane
The final treated sample is submitted for infrared spectroscopic analysis as an almost invisible residue in a small glass stoppered vial or Erlenmeyer flask. A predetermined volume of carbon disulfide is accurately measured into the vial or flask containing the residue. A concentration of aldrin or dieldrin of 0.05% (w./v.) falls in the middle of the calibration curve. The concentration unit is defined as the weight in grams of insecticide per 100 ml. of solution-for example, 0.05% aldrin refers to a solution containing 0.05 gram of aldrin in 100 ml. of solution. If the residue contains 100 micrograms of insect toxicant, 0.20 ml. of carbon disulfide is added to obtain a concentration of 0.05%. The volume of the
6
m w
I
I
residue was found to be negligible, so that the final volume of solution was taken to be equal to the volume of solvent used. The container is thoroughly shaken in order to dissolve the residue completely. The soldtion is then traneferred to an absprption microcell by means of a tuberculin syringe or microsyringe. The microcell should be held in a vertical position, so that the tapered fitting is a t the bottom. The solution is then introduced through the tapered fitting, and the cell is filled, until a drop of solution is exuded out of the curved needle. The microcell is then stoppered with small rubber plugs on the needle and Teflon plugs in the tapered fittings. These plugs are cut with regular cork borers from soft gray rubber stoppers and
I
I
I
I
i
I
SOLVENTCARBON DlSULflDE TETRACHLORDE
INSTRUMENTR R K l N - E W E R WD€L CWFWND-
le 0
TOXARIENE
PURIlY- TECHfflCAL GRAOE
I
I
I
I
I
I
(Spectra of Solvents)
I ~
'
V O L U M E 24, NO. 5, M A Y 1 9 5 2
855 I
>.
B Y
Z
Y
I
1 16.0
I
14.0
13.0
I
I
12.0
11.0
I 10.0
0.0
8.0
SEALED L I Q U I D C E L L SQOlUM CHLORIDE 0.100 M M INSTRUMENTPERKIN-ELMER
MODEL 12C
COMPOUND-CHLORDAN PURITY- 98%
m
N. 0.000 Solvent on1 0.005 %
0.020 %
0.040 %
0.060 %
o.oe0
a/$
/ /
I
1/
t W a v e Length
4 Z e r o Radiation Line Z e r o R a d i a t i o n Line
Figure 8. Analytical Spectra of Aldrin for Various Concentrations in Carbon Disulfide
Figure 9. Analytical Spectra of Dieldrin for Various Concentrations in Carbon Disulfide
-
ANALYTICAL CHEMISTRY
856 Table I. Concn.,
%
0.1000 0.0960 0,0920 0.0900
0.0816 0.0800 0.0780 0.0740 0.0700 0.0600
Calibration Data for Determination of Aldrin AV. No of Readin Readings Ratio A?B 9 3 3 3 3 6 3 3 6 6
Concn.,
%
0.0500 0,0408 0.0400 0.0300 0,0204
0.0200
0.0100 0.0050
0.0000
Av. No of Readin Readings Ratio ATB 9 3 6 6 3 6 9 6 12
Standard deviation = 0,0016
Typical spectra for various concentrations of aldrin in carbon disulfide are shown in Figure 8. The spectrum is recorded in the same manner between 10.82 and 11.18 microns for the determination of dieldrin. Typical spectra for a range of concentrations of dieldrin in carbon disulfide are reproduced in Figure 9. BASE-LINE TECHNIQUES
Base-line techniques as described by Heigl et al. ( 4 ) and Wright (6) were developed. The base line was constructed so that the spectrum of the solvent corresponded to 0% concentration of aldrin or dieldrin. A ratio A / B , corresponding to the true ratio I o / I of incident light intensity to fmal light intensity, is calculated by the base-line technique. Distance A corresponds to I Oand distance B to I . The ratios A / B are correlated t o concentrations in the same manner t h a t the ratio of intensities Ia/I are related to concentration. Aldrin. The method of constructing the base line for the calcu-
lation of the ratio A4/L?is illustrated in Figure 10. The base linc is drawn from the 8.38-micron point of the spectrum to the 8.57micron point. Distance 9 is measured at 8.48 microns from the zero radiation line to the base line. Distance B is also measured a t 8.48 microns from the zero radiation line to the absorption peak. The ratios A / B are correlated for various concentrations of aldrin by means of a calibration curve. .4 typical calibration curve is reproduced in Figure 10. Dieldrin. The method of drawing the base line for the det,ermination of dieldrin is complicated. The solvent, spectrum through the aldrin range is straight, while the solvent spectrum through the dieldrin range is curved. This curvature does not permit one to draw a simple base line if it is required that ratio A / B be unity for the solvent. The method of drawing the base line for the calibration of ratio B I B is illustrated in Figure 11. Line CD is constructed through the 11.04- and 11.11-micron points. Line EF is constructed between the 10.82- and 10.90micron points. The base line is then drawn from the 11.02micron point on line CD to the 10.91-micron point on line EF. Distance A is measured at 10.98 microns from the zero radiation line to the base line and distance B is measured from the zero radiation line to the absorption peak. The ratios A / B and various concentrations of dieldrin are correlated by means of a calibration curve, such as that shown in Figure 11. DISCUSSION
The calibration curves are prepared by measuring ratios A I B for a range of concentrations of the pure compounds and then plotting ratios *4/Bagainst per cent concentration. The data collected in the above process were later analyzed ta determine the accuracy and sensitivity of the particular method. Sumerous solutions of pure insecticide over a range of concen-
I ~
Wave L e n g t h
2 era R a d i a t i o n Line 0
-
Per C e n t C o n c e n t r a t i o n 01 .02 .03 . 0 4 . 0 5 .06 .07 .08 ,OS
v
Figure 10. Construction of Base Line in Determination of Aldrin
'igure 11. Construction of Base Line in Determination of Dieldrin
857
V O L U M E 2 4 , NO. 5, M A Y 1 9 5 2 Table 11. Calibration Data for Determination of Dieldrin Concn.,
%
0,094 0,090 0.080 0.060 0 050 0.045 0,030
Av. Av. No. of Reading, Concn.. K O . of Reading, Readings Ratio A I R Readings Ratio A / B 3 1.234 0,025 3 1,058 3 1,223 0.0228 3 1.053 3 1,196 0 016 3 1.038 0.010 3 1 023 3 1.144 10 1.119 0 005 6 1.012 3 1.107 0.000 19 1,000 3 1.070 Standard deviation = 0.0037
In general, when a number of analyses are carried out in routine fashion three readings are taken per analysis. The probable error should be within &O.OOl% under routine conditions of analysis. ACKNOWLEDGMENT
The authors wish to thank John Wirth, John Streich, and
W. R. Miller for their assistance in collecting the expeGmental data. LITERATURE CITED
trations were prepared and a number of readings taken for each concentration. The accumulated data were then treated by the method of least squares and the probable error was calculated from the standard deviation. Aldrin. The calibration data pertinent to aldrin are tabulated in Table I, including the number of readings and the average value for each concentration. The individual readings rather than the averages were used in the actual statistical treatment. The standard deviation was 0.0016. The sensitivity was 0.000570 aldrin. Dieldrin. The (data collected for the calibration of dieldrin are presented in Table 11. Only the number of readings and the averages are listed. The standard deviation was 0.0037. The sensitivity was 0.0007c7, dieldrin.
Colthup et al., Rev. Sci Instruments, 18, 931 (1947). Danish, A. A,, Koenig, N., and Kuderna, J., “Treatment for Photometric Analysis of Biological Materials Containing Micro Quantities of Aldrin and Dieldrin,” Division of Agricultural and Food Chemistry, Symposium on Methods of Analysis for Micro Quantities of Pesticides, 119th Meeting of AMERICAN CHEMICAL SOCIETY, Boston, April 4, 1951. (3) Danish, A. A., and Lidov, R. E., Advances in Chem Series, No. 1, 190 (1950). (4) Heigl, J. J., Bell, >!I.F., and White, J. U.,ANAL. CHEM.,19, 292 (1947). (5) Lidov, R E., Bluestone, H., Soloway, 8.B., and Kearns, C. W , Advances in Chem. Series, KO.1, 175 (1950). ( 6 ) Wright, N., IND. ENG.CHEW., ANAL.ED.,13, 1 (1941). (1) (3)
RECEIVED for review April 26, 1951. -4coepted January 29, 1952. Presented before the Division of Agricultural and Food Chemistry, Symposium on Methods of Analysis for Micro Quantities of Peaticides, a t the 119th Meeting of the AMERICAN CHEMICAL SOCIETY, Boston, Mass.
Colorimetric Microdiffusion Determination of Chloride Application to Chlorinated Insecticides H. T. GORDON Division of En tornology and Parasitology, University of California, Berkeley 4, Calif.
.
This method is a simpler, faster, and more sensitive colorimetric modification of the Conw-ayniicrodiffusion analysis for chloride (0.3 to 3 micrograms of chloride, in 0.1 to 0.4 ml. of solution) and bromide. It is applicable to many organic compounds containing chlorine or bromine. Chlorine is liberated from organic compounds by direct permanganate oxidation, or by reaction with sodium-n-propoxide to form chloride. Chloride is oxidized to chlorine by permanganate. The chlorine formed diffuses into and quantitatively decolorizes a solution of the dye, Fast Green. Residues of 1 to 10 p.p.m. of chlorinated insecticides (such as chlordan, heptachlor, lindane, toxaphene, methoxychlor, and DDT) in biological material can be determined as alkali-labile chlorine.
08T micromethods for chloride ion are adaptations of argentometric macrotitration ( 13, 24), potentiometry (&?.),or turbidimetry (11). The Conway microdiffusion cell has also been used for chloride determination (4.). The chloride reacts with acid permanganate in the outer chamber of the cell, and chlorine diffuses into the inner chamber containing iodide solution; for 1 to 7 micrograms of chlorine, the method is colorimetric (starch-iodine complex.). X nioie sensitive colorimptiic micromethod for chlorine or bromine is now available, using the tiiphenylmethane dye, Fast Green (8). This paper describes the adaptation of Fast Green colorimetry to the Conway diffusion procedures for chloride and bromide, and the application of this new chloride method to the microanalysis of insecticides having alknli-labile or oxidant-labile chlorine. REAGE3TS
Fast Green FCF, 0.0167% in 0.05 -TI sulfuric acid. The voncentration is not critical, but designed so that 0.15 ml. will contnin 25 micrograms of dye. These are convenient values for the Connay cells and colorimeter tubes used in this n o r k .
Potassium permanganate, 67, in water. Sulfuric acid, c P. conrentrated, low in chloride. This is prepared by adding about 1 gram of C.P. chromic acid to 200 ml. of concentrated sulfuric acid and slon-ly heating to the boiling point in a fume hood, Sulfuric acid, 1 JI, prepared from the low-chloride concentrated acid by diluting approximately 5.4 ml. in water to 100 ml. Standard solutions roiitaining 5, 10, 15, and 20 micrograms of sodium chloride per nil in water. Standard solutions caontaining 10, 20, 30, and 40 micrograms of sodium bromide per nil. in water. Chromic acid, c.P., 20% (m./v.) in 30% (v./v.) sulfuric acid (low-chloride). The chloride is removed by slowly heating to the boiling point in a fume hood. Sodium n-propoxide, approximately 2 M in 1-propanol, is prepared by dissolving clean sodium metal in anhydrous 1-propanol (redistilled over sodium from Eastman Kodak No. 848). The solution contains chloride; most of this ill settle out on standing. The clear supernatant is siphoned off; this has a very low chloride blank. The solution gradually turns yellow, and becomes cloudy after repeated exposure to air, but this does not interfere with its analytical use. Perchloric acid, c.P., 72%. Toxaphene (chlorinated camphene), technical, 67 to 69% chlorine, from the Hercules Yon-der Co., JVilmington, Del.
