Determination of Organophosphorus insecticide Residues Using the Emission Spectrometric Detector C. A. BACHE and D. J. LlSK Pesticide Residue laboratory, Cornell University, Ithaca,
b The emission spectrometric detector is sensitive and specific for determination of organophosphorus insecticide residues. Measuring the intensity of the 2535.65A. atomic phosphorus line following gas chromatography provides a sensitive and highly selective method for these phosphorus compounds. Typical injections of 5 PI. of concentrated solvent extracts of samples, opaque with colored plant extractives, result in chromatograms seldom showing peaks other than the solvent and insecticide. Recoveries of organophosphorus insecticides from 0.03 to 0.6 p.p.m. from potatoes, lettuce, grapes, soil, timothy, alfalfa, milk, urine, chicken, eggs, bees, and halibut are reported.
M C C O R M A C KTong, , and Cooke ( 7 ) used a microwave powered argon discharge for excitation and emission detection of organic compounds emerging from a gas chromatographic column. .I sensitivity of 1 x lo-" gram per second was found for organo1 ~ 1 i o s ~ ~ hcompounds o~us bascd on photoelectric measurement of the 2535.65 A. atomic 1)hospholus line. The possibility of its application as a sensitive, sl)ec+ic phosphorus detector for the deteimination of organophosphorus insecticide residues \vas suggested (6). I n the work reported, the determination of several organop1ios;l)horusinsecticides \vas studied in a variety of foods and biological samples using the detector teni.
N. Y.
the retention time for most of the compounds within about 10 minutes, column temperatures between 165" and 200" C. and argon flow rates from 20 to 115 cc./minute were used. The discharge tube consisted of 0.8mm. bore, 3-mm. 0.d. quartz capillary tubing. The portion of the tube between the column exit and discharge was heated using an outer glass chimney and heating tape. A Raytheon ;\lode1 PGM-10 85-watt microwave generator was used and a filter system was installed in it to reduce ripple to below 1%. Line voltage to the generator was stabilized through a Sola CVS transformer. A spectrometer slit width of 75 microns and a height of 20 mm. was used in virtually all analyses. Procedure. A typical analysis consisted of a column injection of 3 t o 5 pl. of the sample in acetone or diethyl ether with the discharge off. After 1 to 2 minutes to allow escape of the solvent, the microwave power generator was turned on and, a t a power setting of about 8070, the discharge was started with a Tesla coil. The power was then readjusted to the desired setting. Recording of the chromatogram a t 2535.65 then followed. The interval between sample injection and ignition of the discharge prevented the solvent from extinguishing the discharge and eliminated deposition of carbon in the quartz tube. By following this procedure the tube was used daily for 3 months with no visible carbon deposits. Complete fragmentation of organic compounds in the discharge permitted repetitive sample injection and rapid analysis.
Table 1. EXPERIMENTAL
Apparatus. The equipment used was identical t o t h a t described by RlcCormick, Tong, and Cooke (7) with several esceptions. A Research Specialties ;\lode1 601-1 column oven and proportional temperature controller were used. The oven cover was replaced with a piece of 0.5-inch transite upon which the tapered cavity, discharge tube, and lens were mounted. The column \vas borosilicate glass, Eshaped, 5-mm. i.d., and 2 feet long. The packing was either 5% Doiv Corning high vacuum silicone grease (the ethyl acetate soluble fraction) or S.E. 30 silicone gum rubber on 80-100 mesh acid-washed Chromosorb W. To keep
Compound Diazinon Dimethoate
Disyston Ethion Parathion Ronnel
Solvents such as diethyl ether, acetone, chloroform, or hesane were all suitable for injection of the sample. Benzene tailed badly unless about 4 minutes was allowed before ignition. It' also imparted a green color to the discharge. Injection of solvents containing water enhanced solvent tailing. Ignition of the discharge with the coil was immediate. Difficulty was only encountered if air was present in the column or discharge tube. The argon gas was therefore kept flowing overnight. The precise wavelength setting for optimum sensitivity was dekrmined by first locating the approximate 2535.65 A. region using the 2536.52 A. mercury line. The same amount of a given compound was then repeatedly injected while varying the wavelength about 0.2 A. each time in the 2535.65 A. region using a slit width of about 25 microns. The setting showing t,he greatest peak height was used. The effect of ambient temperature changes and normal laboratory vibrations on the spectrometer grating made periodic refocusing essential. Permanent mounting of the discharge tube, lens, and spectrometer on an optical bench and operation in a constant temperature room would be very advantageous. The 2535.65 A. region was free of other strong interfering lines such as carboncarbon or carbon-nitrogen bands. X wide slit opening (75 microns) could therefore be used to increase sensitivity. References to the procedures used for analysis of insecticides in agricultural samples are cited in Table I. Modifications to these procedures were as follows:
Recovery of Insecticides from Agricultural Samples
Sample Grapes Alfalfa Timothy Lettuce Milk Cow urine Potatoes Grapes Soil Bees Halibut Letture Whole chicken Eggs
Isolation procedure (1) (9) (9) (9) (9) (9) (9)
(1) (1) (1)
(4,8) (4,8) (1)
(4,8 ) (4,8)
Added, p.p.m.
Recovery,
0.30 0.06 0.19 0.20 0.03 0.20 0.20 0.18 0.40 0.20 0.60 0.20 0.20 0.25 0.25
79 106 88 91 113 115 98 81 72 73 83 105 82 71 90
VOL. 37, NO. 12, NOVEMBER 1965
%
1477
DlSY STON (6NG.)
(8 NG.)
Figure 1. Gas chromatograms of Disyston, diazinon, and reagent blank
Anderson-MacDougall Procedure (1). After processing the samples by this procedure which included blending with acetone and column chromatography on charcoal, the eluted insecticide fraction was evaporated in a 25-ml. volumetric flask. One milliliter of an equal volume mixture of diethyl ether and n-hexane was added and the flask was made t o volume with saturated sodium sulfate and shaken for 1 minute. The organic layer was injected for analysis. Niessen-Frehse Procedure (9). I n this procedure, samples were blended with acetone and interferences were precipitated with phosphoric acid. After neutralization and chloroform
extraction of the filtrate, the chloroform was evaporated in a 50-ml. volumetric flask. The residue was dissolved in 2 ml. of diethyl ether, the flask was made to volume with 48 ml. of saturated sodium sulfate and shaken. The ether layer was injected for gas chromatography. Kelson (8) and Frear-Enos Procedure (4). This procedure involved blending the sample with acetonitrile and partitioning the filtrate into benzene after addition of sodium chloride solution. The evaporated benzene extract was dissolved in chloroform and chromatographed on Florisil by the Frear-Enos procedure. The first 65 ml. of chloroform eluate (dimethoate fraction) was collected, evaporated, the residue was dissolved in 1 ml. of ether, and injected for analysis. RESULTS AND DISCUSSION
I n Figure 1 are illustrated gas chromatograms of pure standards of Disyston (0,O-diethyl S8-(ethylthio)ethyl phosphorodithioate) and diazinon(0,O - diethyl 0 - (2 - isopropyl - 4 methyl - 6 - pyrimidinyl) phosphorothioate) insecticides in diethyl ether and the reagent blank (ether alone). The interval between sample injection and ignition of the discharge is indicated. Figure 2 shows typical standard curves for the above insecticides, ethion (O,O,O',O' - tetraethyl 8,s' - methylene bisphosphorodithioate), and parathion (0,O - diethyl - 0, p - nitrophenyl thiophosphate). Reproducibility of standard curves was very good. Table I1 lists the detector response of several organophosphorus insecticides
Table II. Detector Response to Organophosphorus Insecticides Sensitivity g. phosphorus/sec.
2535.65A. Electron Microwave Compound power, % emission affinity 9.2 X lo-'* 50 Ciodrin 6.6 X 8.1 x 10-12 90 Diazinon 1.6 X lo-" 50 Dibrom 2.3 X 50 Dimethoate 5.5 x 10-12 26 Disyston 1.9 x 10-11 50 Disyston sulfoxide 2.2 x 10-11 50 Disyston sulfone 2.0 x 10-12 90 Ethion 2.5 X 2.8 X 50 Malathion 8.5 X 1.2 x 10-18 24 Methyl parathion 1.4 X 2.0 x 10-18 90 Parathion 4.6 X 50 Phosdrin 3.3 x 10-11 50 Phosphamidon 1.5 X 7.7 x 10-18 21 Ronnel 2.0 x 10-11 20 Systox 22 6.5 X Thimet 50 7.6 X Thimet oxygen analog 4.5 x 10-18 3.6 x 10-l2 37 Trithion Ciodrin: a (methylbenay134 dimethox phosphinyloxy >cis-crotonate Dibrom: dimethyl-[diethylamidel-cdro-crotonyl (a)]-phos hate Dimethoate: 0,O-dimethyl-S-(N-methyl-carbamoylmethyl) pkosphorodithioate Malathion: S-[ 1,2-6ts(ethoxycarbonyl)ethyl]0,O-dimethyl phosphorodithioate Methyl parathion) 0,O-dimethyl 0,p-nitrophenyl thiophosphate Parathion: 0,O-diethyl 0,p-nitrophenyl thiophosphate Phosdrin: 2-carbomethoxy-1-methylvinyldimethyl phosphate Phosphamidon: 2-chloro-2-diethylcarbamoyl-1-methylvinyldimethyl phosphate Ronnel: O,O-dimethyl-O-(2,4,btrichlorophenyl)hos horothioate Systox: 0,O-diethyl 0-ethylmer,captoeth 1 thiopgospgate Thimet : 0,O-diethyl S-(eth lthiomethyl~~phosphorodithioate Trithion: 0,O-diethyl S-p-c&orophenylthiomethyl phosphorodithioate
1478
ANALYTICAL CHEMISTRY
Oy
20 40 60 80 NANOGRAMS INJECTED
I
100
Figure 2. Standard curves for four insecticides
as determined a t a signal to noise ratio of 3 to 1. The response of several of the compounds to electron affinity detection as determined by Clark (2) is included. The response of the emission spectrometric detector a t 2535.65 A. is in the some range as the sodium thermionic detector ( 5 ) . Differences in emission response are affected by the nature of the bonding and the groups attached to an atom. The length of the carbon chain attached to sulfur appeared to affect markedly the emission response of sulfur as observed by McCormack (6)in a homologous series of monothiol compounds. The low response of Disyston sulfoxide and sulfone as compared to Disyston should be noted. Loss of these compounds by irreversible column adsorption may have occurred. The power settings designated in Table I1 were those found to be optimum for maximum response. Most of the compounds showed larger response a t lower (20-50%) power settings. Several observations regarding response of organophosphorus compounds in relation to operating parameters are noteworthy. At any specific column temperature and flow rate, an optimum power setting exists for maximizing the signal to noise ratio for a given compound. This optimum power setting also varies with the discharge tube geometry and tapered section positioning for a given compound. Eastman (3) has suggested that optimum response may only be realized when the proper impedance (which would be affected by the above parameters) matching between the tapered section and the discharge tube and contents (and therefore maximum microwave power transfer) is obtained. I n general, higher column temperature or faster flow rates
I
1 DISYSTON (0.18FIP.M) IN DOTATOES
ETHION
(0.2 FIP.M.) IN SOIL
CONTROL
I
O?
, 4
12
8
ut-
16
20
24
28
32
MINUTES
%5
zn
Figure 5. Gas chromatograms of ethion added to Canfield silt loam soil and the control
0 6
y
L
4
8
12 .-
5 MINUTES
zc3
Figure 3. Gas chromatograms of Disyston added to potatoes and the control
required higher power settings for maximum response by a given compound. At a given potver setting, response decreased sharply as the bore size or wall thickness of the discharge tube increased. Presumably thick walls absorb more emitted radiation. Tubes with a bore as small as 0.2 mm. were tried but plugging due to column bleeding occurred. The optimum column temperature for maximum response
Figure 4. Gas chromatograms of diazinon added to grapes and the control
varied for different compounds. This may, in part, have been related to the temperature at which partial compound decomposition on the column occurred. I n some instances, lower microwave power greatly reduced solvent tailing which occurred immediately after ignition of the discharge. The time interval between injection and ignition should be kept reasonably constant especially when determining compounds having short retention times. This precaution becomes less critical at lower power.
Increasing column temperature caused a shortening in length of the discharge. This effect would have to be considered if temperature programming was used. The equipment was used for analysis of insecticides in agricultural samples. Figures 3, 4,and 5 show chromatograms of organophosphorus insecticides added to samples prior to extraction and the respective control samples. Figure 6 shows chromatograms and the solution from which they were developed. The flask contains an upper 2-ml. layer of diethyl ether containing the total extractives from 150 grams of alfalfa which was extracted and processed by the procedure of Niessen and Frehse. The upper chromatogram shows the peak corresponding to 88% recovery of 0.19 p.p.m. of dimethoate added to the
0 0 3 6 PPM DIMETHOATE IN LETTUCE TREAT-
DIMETHOATE IN LETTUCE
0.8 J
!
0.7
!
~
I I
OF DIMETHOATE ADDED TO LETTUCE (113 % -
I I
p0.4CONTROL LETTUCE
I
I
3
!P3 0
Z
2a 0.2-
2 I
L Figure 6. Gas chromatograms of dimethoate (0.19 p.p.m.) added to alfalfa, control alfalfa, and the injected ether solution representing the control
Figure 7. Gas chromatograms of dimethoate in field-treated lettuce, the insecticide recovered from lettuce, and the control
0.1 -
1
0
5 IO 15 20 25 DAYS AFTER SPRAYING
I
Figure 8. Disappearance of dimethoate from field-treated lettuce VOL. 37, NO. 12, NOVEMBER 1965
1479
alfalfa prior to extraction. The lower chromatogram represents control alfalfa taken through the procedure. The flask contains the ether solution representing control alfalfa. The ether solution was opaque and viscous, obviously containing a large quantity of dissolved plant substances. Yet the selectivity of the detector at 2535.65 A. for organophosphorus compounds produced the chromatograms showing only the solvent and insecticide peaks. Most of the injected samples which are presented as chromatograms in this paper contained large amounts of extraneous plant materials. I t should be noted that although the response of hydrocarbons observed by LlcCormack et al. (7) at 3883 and 5164 A. were very high (2 X and 1 X 1 0 - 1 4 gram per second, respectively) , injection of green solvent extracts of lettuce into the S.E.30 column at these wavelengths resulted in chromatograms which were remarkably free of major peaks. This implicates the column as an important contributor to selectivity presumably by its irreversible adsorption of many extraneous plant substances.
Figure 7 shows chromatograms of dimethoate in field-sprayed lettuce, and the accompanying insecticide recovery and control samples. The occasional appearance of the solvent peak and those (other than the insecticide) in the chromatograms of these samples are likely due to very weak carbon-carbon or carbon-nitrogen lines near 2535.65 A. which are only detectable when the responsible compound(s) are present in sufficient quantity. Figure 8 illustrates a typical insecticide disappearance curve for dimethoate in the field-sprayed lettuce following emission spectrometric analysis. I n Table I are listed the recoveries of insecticides added to samples (prior to extraction) and the accompanying isolation procedure which was used preceding analysis. The stability and selectivity of the detector allowed these analyses to be made quite routinely. Further studies with organophosphorus and other classes of pesticides will be made to determine the optimum operating parameters for sensitive determination of their residues.
ACKNOWLEDGMENT
The authors thank W. D. Cooke, S. C. Tong, and A. J. McCormack for their helpful suggestions and expert assistance throughout this study. LITERATURE CITED
(1) Anderson, C. A., ;\lacDougall,
D.,
Rept. NO. 8544, pp. 4-5, Chemagro Corp., Kansas City, Mo., 1962. (2) Clark, S. J., “Gas Chromatographic Analysis of Pesticide Residues Using the
Electron Affinity Detector,” JarrellAsh Co., Newtonville, Mass., 1962. (3) Eastman, L. F., Electrical Engineering Dept., Cornell University, Ithaca, N. Y., private communication. (4) Frear, D. E. H., Enos, H. F., J . Agr. Food Chem. 10,477 (1962). (5) Giuffrida, L., J . Assoc. Oflc. Agr. Chemists. 47,294 (1964). (6) McCormack, A. J., “Emission Detectors for Gas Chromatography,” M.S. Thesis, Cornell University, Ithaca, N. Y., 1963. (7) McCormack, A. J., Tong, S. S. C., Cooke, W. D., ANAL.C n m . 37, 1470 (1965). (8) Nelson, R. C., J. Assoc. Ofic. Agr. Chemists. 47, 289 (1964). (9) Kiessen, H., Frehse, H., P$unzenschutz Nachrichten “Buyer” 16, 205 (1963). RECEIVEDfor review April 19, 1965. Accepted June 16, 1965.
Quantitative Spark Source Mass Spectrometry Using Cry os0rptio n Pumping W. L. HARRINGTON, R.
K. SKOGERBOE,
and G. H. MORRISON
Department o f Chemistry, Cornell University, Ithaca, N. Y.
A significant improvement in the quantitative determinaiion of trace impurities in metal samples by spark source mass spectrometry has been achieved through the use of a new cryosorption pump in the source chamber. Tne average precision and accuracy for the simultaneous determination of a lurge variety of elements (metals, nonmetals, and interstitials) in the parts-per-million range was &2070 for a single run, The fast pumping speed and lurge capacity of this simple pump allow rapid pumpdown to low source pressure, maintain lower pressure during sparking, and eliminate hydrocarbon interference with impurity species.
T
IMPORTANCE of spark source mass spectrometry to the field of pure materials research is well established by its ability to detect multiple impurity elements a t the parts-permillion and parts-per-billion level (4, 8, 9). Attempts a t quantitative estimation of such impurities, however, have led to widely divergent results. HE
1480
0
ANALYTICAL CHEMISTRY
Many workers in the field have reported the accuracy of their determinations as 200-30070 relative error (4, 8 , 9, 22). Cooperative tests such as those conducted by the American Society for Testing and Materials ( I ) and the United States Steel Corp. (13) indicate extreme variance of the reported values. I n the few cases where precision and/or accuracy on the order of k30% are reported (6, Y), the results are typically based on a large number of replicate analyses with as many as 100 measurements for a single element. The achievements of this study have allowed the simultaneous estimation of a wide variety of impurity elements (both metals and nonmetals) in National Bureau of Standards (NUS) iron samples with a precision and accuracy of &20% for the more practical case of one analysis consisting of a single graded series of exposures. The success of this work has been due in part to rigid control of analytical procedure and instrument parameters. Most of the improvement, however, has been realized through the use of a new cryosorption pump located in the source
chamber near the sparking sample to lower and maintain constant source pressure, thus reducing or eliminating hydrocarbon interference. EXPERIMENTAL
The experimental facilities and conditions used in this study are summarized in Table I. Emulsion Calibration and Quantitative Calculation. Improved precision in emulsion calibration curves and a n extended range of inteiisity vs. exposure curves have been obtained through t h e use of peak areas rather than peak heights. A simplified method of measuring spectral lines was employed using transmittance areas in conjunction with the Churchill two-line method of emulsion calibration ( 3 ) . The areas of the recorded transmittance peaks nere planimeter-measured and used directly in the preparation of emulsion response curves and in subsequent calculations. It should be noted that these areas differ from the intensity areas used by Owens (12). Transmittance areas are corrected for emulsion response after the area meas-
